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The molecular functions of hepatocyte nuclear factors – In and beyond the liver

  • Hwee Hui Lau
    Affiliations
    Stem Cells and Diabetes Laboratory, Institute of Molecular and Cell Biology, Proteos, Singapore 138673, Singapore
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  • Natasha Hui Jin Ng
    Affiliations
    Stem Cells and Diabetes Laboratory, Institute of Molecular and Cell Biology, Proteos, Singapore 138673, Singapore
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  • Larry Sai Weng Loo
    Affiliations
    Stem Cells and Diabetes Laboratory, Institute of Molecular and Cell Biology, Proteos, Singapore 138673, Singapore

    School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore
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  • Joanita Binte Jasmen
    Affiliations
    Stem Cells and Diabetes Laboratory, Institute of Molecular and Cell Biology, Proteos, Singapore 138673, Singapore
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  • Adrian Kee Keong Teo
    Correspondence
    Corresponding author. Address: Stem Cells and Diabetes Laboratory, Institute of Molecular and Cell Biology, Proteos, Singapore 138673, Singapore.
    Affiliations
    Stem Cells and Diabetes Laboratory, Institute of Molecular and Cell Biology, Proteos, Singapore 138673, Singapore

    School of Biological Sciences, Nanyang Technological University, Singapore 637551, Singapore

    Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117596, Singapore

    Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore 308232, Singapore
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Published:November 23, 2017DOI:https://doi.org/10.1016/j.jhep.2017.11.026

      Summary

      The hepatocyte nuclear factors (HNFs) namely HNF1α/β, FOXA1/2/3, HNF4α/γ and ONECUT1/2 are expressed in a variety of tissues and organs, including the liver, pancreas and kidney. The spatial and temporal manner of HNF expression regulates embryonic development and subsequently the development of multiple tissues during adulthood. Though the HNFs were initially identified individually based on their roles in the liver, numerous studies have now revealed that the HNFs cross-regulate one another and exhibit synergistic relationships in the regulation of tissue development and function. The complex HNF transcriptional regulatory networks have largely been elucidated in rodent models, but less so in human biological systems. Several heterozygous mutations in these HNFs were found to cause diseases in humans but not in rodents, suggesting clear species-specific differences in mutational mechanisms that remain to be uncovered. In this review, we compare and contrast the expression patterns of the HNFs, the HNF cross-regulatory networks and how these liver-enriched transcription factors serve multiple functions in the liver and beyond, extending our focus to the pancreas and kidney. We also summarise the insights gained from both human and rodent studies of mutations in several HNFs that are known to lead to different disease conditions.

      Keywords

      Introduction

      The liver-enriched HNFs are not restricted to the liver and are expressed in a variety of tissues and organs, playing critical roles in regulating tissue development and function.
      The hepatocyte nuclear factors (HNFs) were first identified as liver-enriched transcription factors.
      • Cereghini S.
      Liver-enriched transcription factors and hepatocyte differentiation.
      However, HNFs are also expressed in a variety of other tissues and organs, such as the pancreas and the kidney, playing important roles in regulating the development and functions of these tissues. Studies have successfully identified the binding motifs of HNFs from different families and these binding motifs do not appear to be exclusively active in the liver. A variety of hepatic and non-hepatic genes contain the matched HNF binding motifs in their promoter and enhancer regions, suggesting the likely presence of multiple transcriptional regulatory networks involving HNFs that are not restricted to liver development and function. Importantly, many of the HNFs are involved in complex auto-regulatory and cross-regulatory circuits, likely acting in a combinatorial manner to determine the development of various tissues including the developing embryo, pancreas, kidney and intestines. Herein, we comprehensively summarise the diverse expression patterns and functions of HNFs in the developing embryo and in adulthood, as well as the HNF cross-regulatory network and mutations in these HNF transcription factors that give rise to disease conditions that implicate different organs. In this review, particular attention is placed on the liver, pancreas and kidney, where the roles of HNFs are best studied. We aim to highlight the significance of the HNFs in the liver and beyond, to increase the appreciation of the pleiotropic effects that these factors can have and to encourage a more holistic approach to the understanding of their role in development and disease.

      Molecular structure of the HNF families

      HNFs are classified into four families, namely HNF1, FOXA (or HNF3), HNF4 and ONECUT ([OC] or HNF6), each characterised by distinct regions corresponding to functional domains (summarised in Fig. 1). The HNF1 family comprises of HNF1α and HNF1β, whose DNA-binding domain (DBD) is known to bind to the palindromic consensus sequence GTTAATNATTANC (Fig. 1A). The dimerisation domain at the N-terminus allows both HNF1α and HNF1β to form homodimers or heterodimers.
      • De Simone V.
      • De Magistris L.
      • Lazzaro D.
      • Gerstner J.
      • Monaci P.
      • Nicosia A.
      • et al.
      LFB3, a heterodimer-forming homeoprotein of the LFB1 family, is expressed in specialized epithelia.
      • Rey-Campos J.
      • Chouard T.
      • Yaniv M.
      • Cereghini S.
      VHNF1 is a homeoprotein that activates transcription and forms heterodimers with HNF1.
      The HNF1α and HNF1β genes each encode three isoforms (A, B and C) that appear to have tissue-specific roles.
      • Bach I.
      • Yaniv M.
      More potent transcriptional activators or a transdominant inhibitor of the HNF1 homeoprotein family are generated by alternative RNA processing.
      • Harries L.W.
      • Ellard S.
      • Stride A.
      • Morgan N.G.
      • Hattersley A.T.
      Isomers of the TCF1 gene encoding hepatocyte nuclear factor-1 alpha show differential expression in the pancreas and define the relationship between mutation position and clinical phenotype in monogenic diabetes.
      Figure thumbnail gr1
      Fig. 1Structural domains of HNF families. (A) A schematic diagram of HNF1α and HNF1β. The N-terminus consists of the DD. The DBD of the family is made up of a POU-s and a POU-h. The C-terminus contains the TAD which is less conserved between the two members of the HNF1 family. (B) A schematic diagram representing the subfamily members of Forkhead box, namely FOXA1, FOXA2 and FOXA3. The two TADs are flanking at either ends of each of the transcription factors. The DBD of the family consists of a WH structure with sequences for nuclear localisation. (C) A schematic diagram of the orphan nuclear receptor family HNF4 that comprises HNF4α and HNF4γ. Domain A/B contains the N-terminal AF-1, domain C contains a highly conserved DBD, domain D refers to a hinge region (H) that connects domains C and E, while domain E contains a potential LBD with two zinc fingers that recognise and bind to specific DNA motifs. The LBD also contains AF-2. HNF4 proteins also have a repressor region (domain F) at the C-terminus. (D) A schematic diagram of ONECUT gene family members HNF6 (OC1), OC2 and OC3. A serine/threonine/proline-enriched region (STP Box) is located near the N-terminal end of the OC proteins. Together the CD and the HD form the bipartite DBD near the C-terminus. Diagrams are not drawn to scale. AF-1/2, activation function 1/2; CD, cut domain; DBD, DNA-binding domain; DD, dimerization domain; FOXA, forkhead box A; HD, homeodomain; HNF, hepatocyte nuclear factor; LBD, ligand-binding domain; OC, ONECUT; POU-s, POU-specific domain; POU-h, POU-homeodomain; TAD, transactivation domain; WH, winged helix.
      FOXA (formerly known as HNF3) belongs to the subfamily of the Forkhead box (FOX) proteins, comprising FOXA1, FOXA2 and FOXA3 (formerly HNF3α, HNF3β and HNF3γ).
      • Kaestner K.H.
      • Knöchel W.
      • Martínez D.E.
      Unified nomenclature for the winged helix/forkhead transcription factors.
      • Lai E.
      • Prezioso V.R.
      • Tao W.F.
      • Chen W.S.
      • Darnell J.E.
      Hepatocyte nuclear factor 3 alpha belongs to a gene family in mammals that is homologous to the Drosophila homeotic gene fork head.
      The FOXA family of proteins contain a winged helix (WH) structure (also referred to as the forkhead domain) flanked by sequences required for nuclear localisation (Fig. 1B).
      • Friedman J.R.
      • Kaestner K.H.
      The Foxa family of transcription factors in development and metabolism.
      They also share a highly conserved DBD and bind to the target DNA as a monomer.
      • Rausa F.
      • Samadani U.
      • Ye H.
      • Lim L.
      • Fletcher C.F.
      • Jenkins N.A.
      • et al.
      The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3β in the developing murine liver and pancreas.
      The N- and C-termini of FOXA proteins are conserved and reported to act as the transactivation domains.
      • Pani L.
      • Quian X.B.
      • Clevidence D.
      • Costa R.H.
      The restricted promoter activity of the liver transcription factor hepatocyte nuclear factor 3 beta involves a cell-specific factor and positive autoactivation.
      • Qian X.
      • Costa R.H.
      Analysis of hepatocyte nuclear factor-3 beta protein domains required for transcriptional activation and nuclear targeting.
      HNF4 belongs to the orphan nuclear receptor family and comprises HNF4α and HNF4γ (Fig. 1C). The two transactivation function domains, activation function 1 and 2 (AF-1 and AF-2), located at the N- and C-terminus respectively, activate transcription in a cell type-independent manner.
      • Hadzopoulou-Cladaras M.
      • Kistanova E.
      • Evagelopoulou C.
      • Zeng S.
      • Cladaras C.
      • Ladias J.A.A.
      Functional domains of the nuclear receptor hepatocyte nuclear factor 4.
      • Sladek F.M.
      • Seidel S.D.
      Hepatocyte nuclear factor 4α.
      A ligand-binding domain (LBD) is located adjacent to AF-2 and transactivates genes in a ligand-dependent manner, providing an additional point of control for regulating protein activity. Of note, HNF4 proteins also contain a repressor domain, region F, that has an inhibitory function,
      • Hadzopoulou-Cladaras M.
      • Kistanova E.
      • Evagelopoulou C.
      • Zeng S.
      • Cladaras C.
      • Ladias J.A.A.
      Functional domains of the nuclear receptor hepatocyte nuclear factor 4.
      which has not been characterised in other HNF families. HNF4α is encoded by two developmentally-regulated promoters (P1 and P2), and differential promoter usage and alternative splicing are now known to generate up to twelve isoforms (P1-derived HNF4α1-6 and P2-derived HNF4α7-12) that are expressed in a temporal and tissue-specific manner.
      • Drewes T.
      • Senkel S.
      • Holewa B.
      • Ryffel G.U.
      Human hepatocyte nuclear factor 4 isoforms are encoded by distinct and differentially expressed genes.
      • Huang J.
      • Levitsky L.L.
      • Rhoads D.B.
      Novel P2 promoter-derived HNF4alpha isoforms with different N-terminus generated by alternate exon insertion.
      • Sladek F.M.
      • Ruse M.D.
      • Nepomuceno L.
      • Huang S.-M.
      • Stallcup M.R.
      Modulation of transcriptional activation and coactivator interaction by a splicing variation in the f domain of nuclear receptor hepatocyte nuclear factor 4α1.
      • Thomas H.
      • Jaschkowitz K.
      • Bulman M.
      • Frayling T.M.
      • Mitchell S.M.
      • Roosen S.
      • et al.
      A distant upstream promoter of the HNF-4alpha gene connects the transcription factors involved in maturity-onset diabetes of the young.
      OC1 (or HNF6) belongs to the one cut homeobox gene family. It consists of a single cut domain (CD) and a divergent homeodomain (HD) (Fig. 1D) that form the bipartite DBD.
      • Iyaguchi D.
      • Yao M.
      • Watanabe N.
      • Nishihira J.
      • Tanaka I.
      DNA recognition mechanism of the ONECUT homeodomain of transcription factor HNF-6.
      • Lemaigre F.P.
      • Durviaux S.M.
      • Truong O.
      • Lannoy V.J.
      • Hsuan J.J.
      • Rousseau G.G.
      Hepatocyte nuclear factor 6, a transcription factor that contains a novel type of homeodomain and a single cut domain.
      The cut homeodomain contains sequences that mediate nuclear localisation and transcriptional activation, in conjunction with the N-terminal STP box (serine/threonine/proline-enriched region).
      • Rausa F.
      • Samadani U.
      • Ye H.
      • Lim L.
      • Fletcher C.F.
      • Jenkins N.A.
      • et al.
      The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3β in the developing murine liver and pancreas.
      • Lannoy V.J.
      • Rodolosse A.
      • Pierreux C.E.
      • Rousseau G.G.
      • Lemaigre F.P.
      Transcriptional stimulation by hepatocyte nuclear factor-6: target-specific recruitment of either creb-binding protein (CBP) or p300/CBP-associated factor (p/CAF).
      The members of this family include OC1 and its two paralogs, OC2
      • Jacquemin P.
      • Lannoy V.J.
      • Rousseau G.G.
      • Lemaigre F.P.
      OC-2, a novel mammalian member of the ONECUT class of homeodomain transcription factors whose function in liver partially overlaps with that of hepatocyte nuclear factor-6.
      and OC3.
      • Vanhorenbeeck V.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      OC-3, a Novel Mammalian Member of the ONECUT Class of Transcription Factors.
      It is clear that while the HNF family members harbour common features such as DNA-binding and transactivation capabilities, they are largely defined by distinct, though related, conserved structural domains that account for their functional diversity. The multiple isoforms within each subfamily lend further complexity to their potential to regulate cellular and tissue functions (Fig. 1). The following sections in this review will illustrate overlapping expression patterns among HNF family members and a complex cross-regulatory network connecting several of these HNF isoforms. Studies on rodent gene perturbations and naturally-occurring mutations in humans further reveal their involvement in disease pathophysiology.

      Expression profile of HNFs during mammalian embryonic development and in adulthood

      To-date, the expression patterns of HNFs during development have largely been characterised in rodent tissues, given the limited access to human tissues. These studies have provided important information on the spatial and temporal expression patterns of different HNF families and their isoforms, and thus increased our understanding of the role of these HNFs across multiple tissues.
      The expression patterns of HNF family members during mammalian embryonic development and in adulthood have been better characterised in rodents than in human tissues. In this section, we highlight and compare the spatial and temporal expression patterns of the various HNFs, with a particular focus on the liver, pancreas and kidney. These organs are known to exhibit high expression levels of many of the HNF family members, are often implicated in diseases resulting from dysregulation of HNF proteins, and provide the tissues in which the respective roles of HNFs are most well-studied. Discussion of the role of HNF family members in other organs is beyond the scope of this review; nonetheless, their expression patterns across other mouse and human tissues are summarised (Table 1, Table 2).
      Table 1Tissue-specific expression of HNF family members in the adult mouse.
      Information consolidated from.
      • De Simone V.
      • De Magistris L.
      • Lazzaro D.
      • Gerstner J.
      • Monaci P.
      • Nicosia A.
      • et al.
      LFB3, a heterodimer-forming homeoprotein of the LFB1 family, is expressed in specialized epithelia.
      • Lai E.
      • Prezioso V.R.
      • Tao W.F.
      • Chen W.S.
      • Darnell J.E.
      Hepatocyte nuclear factor 3 alpha belongs to a gene family in mammals that is homologous to the Drosophila homeotic gene fork head.
      • Cereghini S.
      • Ott M.O.
      • Power S.
      • Maury M.
      Expression patterns of vHNF1 and HNF1 homeoproteins in early postimplantation embryos suggest distinct and sequential developmental roles.
      • Ott M.-O.
      • Rey-Campos J.
      • Cereghini S.
      • Yaniv M.
      VHNF1 is expressed in epithelial cells of distinct embryonic origin during development and precedes HNF1 expression.
      • Monaghan A.P.
      • Kaestner K.H.
      • Grau E.
      • Schutz G.
      Postimplantation expression patterns indicate a role for the mouse forkhead/HNF-3 alpha, beta and gamma genes in determination of the definitive endoderm, chordamesoderm and neuroectoderm.
      • Lazzaro D.
      • De Simone V.
      • De Magistris L.
      • Lehtonen E.
      • Cortese R.
      LFB1 and LFB3 homeoproteins are sequentially expressed during kidney development.
      • Ang S.L.
      • Rossant J.
      HNF-3 beta is essential for node and notochord formation in mouse development.
      • Ang S.L.
      • Wierda A.
      • Wong D.
      • Stevens K.A.
      • Cascio S.
      • Rossant J.
      • et al.
      The formation and maintenance of the definitive endoderm lineage in the mouse: involvement of HNF3/forkhead proteins.
      Coloured squares denote expression. *Endocrine tissues refer to thyroid, parathyroid and adrenal glands. Foxa, forkhead box A; Hnf, hepatocyte nuclear factor; Oc, Onecut.
      Table 2Tissue-specific expression of HNF family members in human embryonic and adult tissues.
      Information on embryonic tissue expression is extracted from “An integrative transcriptomic atlas of organogenesis in human embryos”
      • Gerrard D.T.
      • Berry AA.
      • Jennings R.E.
      • Piper Hanley K.
      • Bobola N.
      • Hanley N.A.
      An integrative transcriptomic atlas of organogenesis in human embryos.
      and consolidated from.
      • Harries L.W.
      • Locke J.M.
      • Shields B.
      • Hanley N.A.
      • Hanley K.P.
      • Steele A.
      • et al.
      Mutation carriers is moderated by the expression of HNF4A isoforms from the P1 promoter during fetal development.
      • Kato N.
      • Motoyama T.
      Expression of hepatocyte nuclear factor-1beta in human urogenital tract during the embryonic stage.
      No information is available for ONECUT family members. Information on adult tissue expression is consolidated from
      • Harries L.W.
      • Ellard S.
      • Stride A.
      • Morgan N.G.
      • Hattersley A.T.
      Isomers of the TCF1 gene encoding hepatocyte nuclear factor-1 alpha show differential expression in the pancreas and define the relationship between mutation position and clinical phenotype in monogenic diabetes.
      • Drewes T.
      • Senkel S.
      • Holewa B.
      • Ryffel G.U.
      Human hepatocyte nuclear factor 4 isoforms are encoded by distinct and differentially expressed genes.
      • Harries L.W.
      • Locke J.M.
      • Shields B.
      • Hanley N.A.
      • Hanley K.P.
      • Steele A.
      • et al.
      Mutation carriers is moderated by the expression of HNF4A isoforms from the P1 promoter during fetal development.
      • Kolatsi-Joannou M.
      • Bingham C.
      • Ellard S.
      • Bulman M.P.
      • Allen L.I.S.
      • Hattersley A.T.
      • et al.
      Hepatocyte nuclear factor-1β: a new kindred with renal cysts and diabetes and gene expression in normal human development.
      and adapted from the Protein Atlas database (http://www.proteinatlas.org/). Coloured squares denote expression. Endocrine tissues refer to thyroid, parathyroid and adrenal glands. FOXA, forkhead box A; HNF, hepatocyte nuclear factor; OC, ONECUT.

      Rodent embryonic development and adulthood

      In the mouse, Hnf1β is first detected in the primitive endoderm on embryonic day (E)4.5 and is required for specification of the primitive endoderm lineage.
      • Barbacci E.
      • Reber M.
      • Ott M.O.
      • Breillat C.
      • Huetz F.
      • Cereghini S.
      Variant hepatocyte nuclear factor 1 is required for visceral endoderm specification.
      • Cereghini S.
      • Ott M.O.
      • Power S.
      • Maury M.
      Expression patterns of vHNF1 and HNF1 homeoproteins in early postimplantation embryos suggest distinct and sequential developmental roles.
      The expression of Hnf1β precedes Hnf1α during embryogenesis, as Hnf1α transcripts are first detected in the yolk sac on E8.5.
      • Cereghini S.
      • Ott M.O.
      • Power S.
      • Maury M.
      Expression patterns of vHNF1 and HNF1 homeoproteins in early postimplantation embryos suggest distinct and sequential developmental roles.
      Hnf4α is expressed in the visceral endoderm from E4.5,
      • Chen W.S.
      • Manova K.
      • Weinstein D.C.
      • Duncan S.A.
      • Plump A.S.
      • Prezioso V.R.
      • et al.
      Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
      • Duncan S.A.
      • Manova K.
      • Chen W.S.
      • Hoodless P.
      • Weinstein D.C.
      • Bachvarova R.F.
      • et al.
      Expression of transcription factor HNF-4 in the extraembryonic endoderm, gut, and nephrogenic tissue of the developing mouse embryo: HNF-4 is a marker for primary endoderm in the implanting blastocyst.
      • Li J.
      • Ning G.
      • Duncan S.A.
      Mammalian hepatocyte differentiation requires the transcription factor HNF-4α.
      • Parviz F.
      • Matullo C.
      • Garrison W.D.
      • Savatski L.
      • Adamson J.W.
      • Ning G.
      • et al.
      Hepatocyte nuclear factor 4alpha controls the development of a hepatic epithelium and liver morphogenesis.
      • Taraviras S.
      • Monaghan A.P.
      • Schutz G.
      • Kelsey G.
      Characterization of the mouse HNF-4 gene and its expression during mouse embryogenesis.
      and is subsequently detected in the liver bud and hindgut starting from E8.5.
      • Chen W.S.
      • Manova K.
      • Weinstein D.C.
      • Duncan S.A.
      • Plump A.S.
      • Prezioso V.R.
      • et al.
      Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
      • Duncan S.A.
      • Manova K.
      • Chen W.S.
      • Hoodless P.
      • Weinstein D.C.
      • Bachvarova R.F.
      • et al.
      Expression of transcription factor HNF-4 in the extraembryonic endoderm, gut, and nephrogenic tissue of the developing mouse embryo: HNF-4 is a marker for primary endoderm in the implanting blastocyst.
      • Li J.
      • Ning G.
      • Duncan S.A.
      Mammalian hepatocyte differentiation requires the transcription factor HNF-4α.
      • Parviz F.
      • Matullo C.
      • Garrison W.D.
      • Savatski L.
      • Adamson J.W.
      • Ning G.
      • et al.
      Hepatocyte nuclear factor 4alpha controls the development of a hepatic epithelium and liver morphogenesis.
      • Nammo T.
      • Yamagata K.
      • Tanaka T.
      • Kodama T.
      • Sladek F.M.
      • Fukui K.
      • et al.
      Expression of HNF-4alpha (MODY1), HNF-1beta (MODY5), and HNF-1alpha (MODY3) proteins in the developing mouse pancreas.
      On the other hand, Foxa1, Foxa2 and Foxa3 have distinct temporal expression patterns and appear in partially overlapping domains of the definitive endoderm and notochord.
      Hnf1β transcripts are subsequently detected in the foregut endoderm on E9, from which the liver and pancreas develop. During liver development, both Hnf1α and Hnf1β transcripts can be detected in the liver primordia by E10.5, and they continue to be present in the liver throughout embryonic life.
      • Ott M.-O.
      • Rey-Campos J.
      • Cereghini S.
      • Yaniv M.
      VHNF1 is expressed in epithelial cells of distinct embryonic origin during development and precedes HNF1 expression.
      On E9.5, Foxa1 and Foxa2 are also highly expressed during liver bud formation while Foxa3 is expressed at a lower level. The expression of Foxa1 and Foxa2 then falls between E12.5 and 15.5 before increasing again in the adult liver. In contrast, Foxa3 is initially weakly expressed but its expression increases on E10.5 and remains high throughout liver development.
      • Monaghan A.P.
      • Kaestner K.H.
      • Grau E.
      • Schutz G.
      Postimplantation expression patterns indicate a role for the mouse forkhead/HNF-3 alpha, beta and gamma genes in determination of the definitive endoderm, chordamesoderm and neuroectoderm.
      Similar to the expression profile observed for Foxa1 and Foxa2, Oc1 is detected on E9 when liver differentiation occurs, then disappears transiently between E12.5 and E15, before it continues to be expressed within the extrahepatic biliary system and the liver throughout development.
      • Rausa F.
      • Samadani U.
      • Ye H.
      • Lim L.
      • Fletcher C.F.
      • Jenkins N.A.
      • et al.
      The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3β in the developing murine liver and pancreas.
      • Lannoy V.J.
      • Rodolosse A.
      • Pierreux C.E.
      • Rousseau G.G.
      • Lemaigre F.P.
      Transcriptional stimulation by hepatocyte nuclear factor-6: target-specific recruitment of either creb-binding protein (CBP) or p300/CBP-associated factor (p/CAF).
      • Landry C.
      • Clotman F.
      • Hioki T.
      • Oda H.
      • Picard J.J.
      • Lemaigre F.P.
      • et al.
      HNF-6 is expressed in endoderm derivatives and nervous system of the mouse embryo and participates to the cross-regulatory network of liver-enriched transcription factors.
      In contrast to Foxa and Oc1, high levels of Hnf4 transcripts are localised to the periphery of the liver where hepatocytes develop from E11.5 to E16, but not in the centre where haematopoietic cells differentiate.
      • Taraviras S.
      • Monaghan A.P.
      • Schutz G.
      • Kelsey G.
      Characterization of the mouse HNF-4 gene and its expression during mouse embryogenesis.
      In the mouse liver, there is a controlled switch from P2 (distal) to P1 (proximal) promoter-driven transcription of Hnf4α from foetal life to birth, and P1-derived transcripts continue to be expressed at significantly higher levels in adult liver.
      • Dean S.
      • Tang J.I.
      • Seckl J.R.
      • Nyirenda M.J.
      Developmental and tissue-specific regulation of hepatocyte nuclear factor 4-alpha (HNF4-alpha) isoforms in rodents.
      The early activation of Hnf1β, Foxa1, Foxa2, Hnf4α and Oc1 in the developing liver between E8.5–9.5 suggests their importance in the early commitment towards the hepatoblast lineage, alongside other transcription factors and signalling molecules.
      In the context of pancreatic development, Hnf1β and Hnf4α are expressed by most epithelial cells of the pancreatic bud from E9.5, and in Pdx1+ pancreatic progenitors followed by Ngn3+ (Neurog3) endocrine precursors (∼E12.5) in the early pancreas.
      • Nammo T.
      • Yamagata K.
      • Tanaka T.
      • Kodama T.
      • Sladek F.M.
      • Fukui K.
      • et al.
      Expression of HNF-4alpha (MODY1), HNF-1beta (MODY5), and HNF-1alpha (MODY3) proteins in the developing mouse pancreas.
      In fact, Hnf1β is likely to be a key player in early pancreas morphogenesis as it is expressed in the pre-pancreatic foregut endoderm and in early multipotent pancreatic progenitor cells.
      • De Vas M.G.
      • Kopp J.L.
      • Heliot C.
      • Sander M.
      • Cereghini S.
      • Haumaitre C.
      Hnf1b controls pancreas morphogenesis and the generation of Ngn3+ endocrine progenitors.
      As the pancreas starts to mature (∼E17.5), Hnf4α can be detected in the islet, ductal and acinar cells. In contrast, Hnf1β gene expression is largely restricted to ductal cells.
      • Nammo T.
      • Yamagata K.
      • Tanaka T.
      • Kodama T.
      • Sladek F.M.
      • Fukui K.
      • et al.
      Expression of HNF-4alpha (MODY1), HNF-1beta (MODY5), and HNF-1alpha (MODY3) proteins in the developing mouse pancreas.
      Hnf4γ protein expression in the pancreas is strongest in the endocrine cells and weakest in the cells lining the ducts.
      • Taraviras S.
      • Mantamadiotis T.
      • Dong-Si T.
      • Mincheva A.
      • Lichter P.
      • Drewes T.
      • et al.
      Primary structure, chromosomal mapping, expression and transcriptional activity of murine hepatocyte nuclear factor 4gamma.
      Hnf1α is expressed slightly later in the pancreatic epithelial cells on E10.5, and as pancreatic development continues, Hnf1α expression becomes more confined to both the acinar and developing islet cells, similar to the expression pattern observed for Hnf4α.
      • Nammo T.
      • Yamagata K.
      • Hamaoka R.
      • Zhu Q.
      • Akiyama T.
      • Gonzalez F.
      • et al.
      Expression profile of MODY3/HNF-1α protein in the developing mouse pancreas.
      • Shih D.Q.
      • Stoffel M.
      Dissecting the transcriptional network of pancreatic islets during development and differentiation.
      This suggests a role for Hnf1α in the maintenance of the profile of differentiated islet cells. Similar to Hnf1α, Oc1 is first expressed on E10.5 in the developing pancreas, while Oc2 exhibits biphasic expression, first decreasing after E12.5 and then increasing after E15.5. Conversely, Oc1 expression is high until E16.5, after which it decreases.
      • Jacquemin P.
      • Pierreux C.E.
      • Fierens S.
      • van Eyll J.M.
      • Lemaigre F.P.
      • Rousseau G.G.
      Cloning and embryonic expression pattern of the mouse Onecut transcription factor OC-2.
      On E18, Oc1 is absent from islets
      • Landry C.
      • Clotman F.
      • Hioki T.
      • Oda H.
      • Picard J.J.
      • Lemaigre F.P.
      • et al.
      HNF-6 is expressed in endoderm derivatives and nervous system of the mouse embryo and participates to the cross-regulatory network of liver-enriched transcription factors.
      but is localised to the ductal cells. Unlike Oc1 and Oc2, Oc3 is not detected in the developing liver or pancreas.
      • Vanhorenbeeck V.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      OC-3, a Novel Mammalian Member of the ONECUT Class of Transcription Factors.
      • Jacquemin P.
      • Pierreux C.E.
      • Fierens S.
      • van Eyll J.M.
      • Lemaigre F.P.
      • Rousseau G.G.
      Cloning and embryonic expression pattern of the mouse Onecut transcription factor OC-2.
      Overall, it is clear that the transcription factors display dynamic gene expression patterns due to carefully orchestrated mechanisms for the patterning and eventual development of the liver and pancreas from the definitive endoderm.
      During kidney development, Hnf1β is expressed in the nephrogenic zone of the kidney in newborn rats whereas Hnf1α is absent. Subsequently, Hnf1α transcripts are restricted to the proximal and distal tubules whereas Hnf1β can also be detected in the collecting ducts.
      • Lazzaro D.
      • De Simone V.
      • De Magistris L.
      • Lehtonen E.
      • Cortese R.
      LFB1 and LFB3 homeoproteins are sequentially expressed during kidney development.
      From E10.5, mesonephric tubules of the developing kidney express high levels of Hnf4 transcripts followed by the newly-formed metanephros on E12.5. In the newborn mouse kidney, Hnf4 is strongly expressed in the proximal and distal convoluted tubules of the cortex but is weakly expressed in the loop of Henle.
      • Taraviras S.
      • Monaghan A.P.
      • Schutz G.
      • Kelsey G.
      Characterization of the mouse HNF-4 gene and its expression during mouse embryogenesis.
      There is little information on the involvement of members of the Foxa and Onecut families in the development of the kidney.
      During adulthood, Hnf1α and Hnf1β are both expressed in tissues such as the liver, pancreas, kidney and intestine (Table 1, Table 2). Hnf1β may have broader regulatory roles as it is expressed in organs that lack Hnf1α, such as the thymus, lung, testis and ovary.
      • Cereghini S.
      Liver-enriched transcription factors and hepatocyte differentiation.
      • Reber M.
      • Cereghini S.
      Variant Hepatocyte Nuclear Factor 1 expression in the mouse genital tract.
      Foxa1 and Foxa2 are both highly expressed in the epithelial cells of the developing liver and pancreas but also a number of other tissues in the body (Table 1). Hnf4 transcripts are most abundantly expressed in the adult liver, kidney and intestines, with relatively weaker expression observed in the pancreas, skin and stomach (Table 1).
      • Cattin A.L.
      • Le Beyec J.
      • Barreau F.
      • Saint-Just S.
      • Houllier A.
      • Gonzalez F.J.
      • et al.
      Hepatocyte nuclear factor 4alpha, a key factor for homeostasis, cell architecture, and barrier function of the adult intestinal epithelium.
      • Kanazawa T.
      • Konno A.
      • Hashimoto Y.
      • Kon Y.
      Expression of hepatocyte nuclear factor 4α in developing mice.
      • Miquerol L.
      • Lopez S.
      • Cartier N.
      • Tulliez M.
      • Raymondjean M.
      • Kahn A.
      Expression of the L-type pyruvate kinase gene and the hepatocyte nuclear factor 4 transcription factor in exocrine and endocrine pancreas.
      • Sladek F.M.
      • Zhong W.M.
      • Lai E.
      • Darnell J.E.
      Liver-enriched transcription factor HNF-4 is a novel member of the steroid hormone receptor superfamily.
      In adult mice, only Oc1 is detected at high levels in the pancreas and at very low levels in the testis. Both Oc1 and Oc2 transcripts are highly expressed in the liver.
      • Vanhorenbeeck V.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      OC-3, a Novel Mammalian Member of the ONECUT Class of Transcription Factors.
      Members of the Onecut family are also expressed in the neural system, stomach, biliary system and the intestine (Table 1).

      Limited information exists on human embryonic development and adulthood

      As mentioned previously, HNF1α is expressed as three alternatively processed transcripts whose expression is likely to be regulated during development.
      • Bach I.
      • Yaniv M.
      More potent transcriptional activators or a transdominant inhibitor of the HNF1 homeoprotein family are generated by alternative RNA processing.
      In humans, HNF1α(A) is the major isoform in the adult liver, kidney and foetal pancreas; whereas HNF1α(B) predominates in the adult pancreas.
      • Harries L.W.
      • Ellard S.
      • Stride A.
      • Morgan N.G.
      • Hattersley A.T.
      Isomers of the TCF1 gene encoding hepatocyte nuclear factor-1 alpha show differential expression in the pancreas and define the relationship between mutation position and clinical phenotype in monogenic diabetes.
      This is in contrast to observations in rodents, where Hnf1α(A) predominates in the liver, pancreas and kidney.
      • Harries L.W.
      • Brown J.E.
      • Gloyn A.L.
      Species-specific differences in the expression of the HNF1A, HNF1B and HNF4A genes.
      P1-driven HNF4α isoforms are known to be liver-specific whereas P2-driven HNF4α isoforms are pancreas-specific.
      • Thomas H.
      • Jaschkowitz K.
      • Bulman M.
      • Frayling T.M.
      • Mitchell S.M.
      • Roosen S.
      • et al.
      A distant upstream promoter of the HNF-4alpha gene connects the transcription factors involved in maturity-onset diabetes of the young.
      However, one report had suggested the expression of P1-derived HNF4α transcripts in human pancreatic β cells.
      • Eeckhoute J.
      • Moerman E.
      • Bouckenooghe T.
      • Lukoviak B.
      • Pattou F.
      • Formstecher P.
      • et al.
      Hepatocyte nuclear factor 4 alpha isoforms originated from the P1 promoter are expressed in human pancreatic beta-cells and exhibit stronger transcriptional potentials than P2 promoter-driven isoforms.
      In a more recent study, the expression of HNF4α isoforms in human gestation stage foetal pancreas and adult tissues was more thoroughly examined by isoform-specific real-time PCR.
      • Harries L.W.
      • Locke J.M.
      • Shields B.
      • Hanley N.A.
      • Hanley K.P.
      • Steele A.
      • et al.
      Mutation carriers is moderated by the expression of HNF4A isoforms from the P1 promoter during fetal development.
      Up to 23% of HNF4α expression in 9th–26th week foetal pancreas was found to be of P1 origin, but only P2-driven HNF4α isoforms can be detected in the adult pancreas.
      • Harries L.W.
      • Locke J.M.
      • Shields B.
      • Hanley N.A.
      • Hanley K.P.
      • Steele A.
      • et al.
      Mutation carriers is moderated by the expression of HNF4A isoforms from the P1 promoter during fetal development.
      Immunohistochemistry (IHC) analyses have also shown that P1-driven HNF4α isoforms are expressed in the liver and kidney, whereas P2-driven isoforms are expressed in the pancreas, bile duct and stomach.
      • Tanaka T.
      • Jiang S.
      • Hotta H.
      • Takano K.
      • Iwanari H.
      • Sumi K.
      • et al.
      Dysregulated expression of P1 and P2 promoter-driven hepatocyte nuclear factor-4α in the pathogenesis of human cancer.
      Northern blot analysis revealed that HNF4γ transcripts are weakly expressed in the pancreas, kidney, testis, small intestine and colon, but HNF4α is generally expressed at higher levels in all of these tissues, as well as in the liver.
      • Drewes T.
      • Senkel S.
      • Holewa B.
      • Ryffel G.U.
      Human hepatocyte nuclear factor 4 isoforms are encoded by distinct and differentially expressed genes.
      In two other human studies, a total of 18 foetuses were examined for HNF1β expression in the urogenital tract using reverse transcription PCR, ISH (In situ hybridisation) or IHC. HNF1β is present in mesonephric duct derivatives such as the efferent duct or epididymis in 12th-, 21st- and 22nd-gestational-week-old foetuses, but not in Müllerian duct derivatives, such as the uterus or fallopian tube after the 18th-gestational week.
      • Kolatsi-Joannou M.
      • Bingham C.
      • Ellard S.
      • Bulman M.P.
      • Allen L.I.S.
      • Hattersley A.T.
      • et al.
      Hepatocyte nuclear factor-1β: a new kindred with renal cysts and diabetes and gene expression in normal human development.
      • Kato N.
      • Motoyama T.
      Expression of hepatocyte nuclear factor-1beta in human urogenital tract during the embryonic stage.
      The pattern of expression of HNF family members in human embryonic and various adult tissues is highlighted (Table 2). The former is based on transcriptomic data from various tissues of human embryos at Carnegie Stages 14–22.
      • Gerrard D.T.
      • Berry AA.
      • Jennings R.E.
      • Piper Hanley K.
      • Bobola N.
      • Hanley N.A.
      An integrative transcriptomic atlas of organogenesis in human embryos.
      To date, little has been reported on the expression profiles of FOXA and OC in human development.
      Notably, because of the lack of access to human foetal tissues, there is limited information relating to the expression profile of HNFs during human embryonic development. There continues to be a heavy reliance on the extrapolation from rodent studies. While it is possible to compare the expression profile of HNFs between rodents and humans in adult tissues, the fact that many of these genes are developmental genes first expressed in the embryo implies that HNF gene mutations would have an early impact on tissue development. Given the divergent phenotypes between rodents and humans with heterozygous mutations in several HNF genes, further understanding of the expressions and functions of HNF genes during human embryonic development is critical. This issue will be further discussed in a later section of this review.

      The complex HNF cross-regulatory network in various tissues and organs

      The HNF transcription factors operate in a developmental stage- and tissue-specific manner to mediate cellular development and function. In fact, increasing evidence has shown that several of these HNFs are functionally-related and/or part of a shared regulatory network. In this section, we focus on the cross-regulatory interactions between the HNFs and highlight their roles in regulating different tissues during embryonic development, particularly during the development of the liver, pancreas and kidney. The bulk of these studies have been carried out in rodent models, providing valuable insight into the regulatory circuits that involve HNFs.

      The role of FOXA, HNF1 and HNF4 transcription factors during embryonic development

      Several HNFs are involved in cross-regulatory networks that are functionally-related during embryonic development. The regulation of downstream target genes by specific expression of the HNFs determine cell fate and eventual development of tissues such as the liver or pancreas.
      FOXA proteins are heavily involved in tissue regionalisation within the definitive endoderm. FOXA1 and FOXA2 regulate genes via different mechanisms although they share similar expression patterns during development
      • Duncan S.A.
      • Navas M.A.
      • Dufort D.
      • Rossant J.
      • Stoffel M.
      Regulation of a transcription factor network required for differentiation and metabolism.
      (Fig. 2A). Specifically, FOXA2 is likely to act upstream of the HNF cross-regulatory transcriptional network as Foxa2-null embryos fail to express Foxa1 and also express reduced levels of Hnf4α and Hnf1α.
      • Duncan S.A.
      • Manova K.
      • Chen W.S.
      • Hoodless P.
      • Weinstein D.C.
      • Bachvarova R.F.
      • et al.
      Expression of transcription factor HNF-4 in the extraembryonic endoderm, gut, and nephrogenic tissue of the developing mouse embryo: HNF-4 is a marker for primary endoderm in the implanting blastocyst.
      Furthermore, Foxa1 negatively regulates the expression of Hnf4α and Hnf1α despite sharing identical DNA-binding sites with Foxa2.
      • Duncan S.A.
      • Navas M.A.
      • Dufort D.
      • Rossant J.
      • Stoffel M.
      Regulation of a transcription factor network required for differentiation and metabolism.
      FOXA1 uniquely achieves its repressor function by competing for binding targets with FOXA2. Insulin purportedly upregulates Foxa2 while reducing Foxa1 expression, resulting in a timely regulation of downstream targets.
      • Duncan S.A.
      • Navas M.A.
      • Dufort D.
      • Rossant J.
      • Stoffel M.
      Regulation of a transcription factor network required for differentiation and metabolism.
      Figure thumbnail gr2
      Fig. 2Schematic outlining key HNF cross-regulatory networks. (A) HNF regulatory network in the developing embryo. FOXA2 acts upstream of FOXA1 to regulate its initial expression in the embryo. FOXA1 negatively regulates the gene expression of P1-driven HNF4α and HNF1α. FOXA1 achieves its repressor function by competing for binding targets with FOXA2. FOXA3 is positively regulated by HNF1α and HNF1β. HNF1β synergises with GATA6 to strongly regulate expression of P1-driven HNF4α. (B) HNF regulatory network in the liver. FOXA1, FOXA2 and GATA4 are involved in the initial specification of liver progenitors. FOXA2 further self-regulates its own expression. OC1 positively regulates HNF1β and HNF4α. In turn, HNF1β binds to the promoter of HNF4α and HNF1α to regulate their expression. OC1, HNF1β and HNF1α regulate the expression of HNF4α. P1 promoter-driven HNF4α suppresses the expression of P2 promoter-driven HNF4α in the adult liver. (C) HNF regulatory network in the pancreas: HNF1β activates OC1 expression in pancreatic precursor cells. OC1 is involved in initiating the onset of PDX1 expression for pancreatic cell fate specification. OC1 regulates pancreatic endocrine differentiation via NGN3 (NEUROG3) expression. OC1 also binds directly to the FOXA2 promoter. P2 promoter-driven HNF4α and HNF1α form a cross-regulatory loop, forming a complex with p300 to promote downstream gene expression. HNF1α is also involved in regulating the expression of HNF4α, HNF4γ and FOXA3 transcripts in islets. FOXA, forkhead box A; HNF, hepatocyte nuclear factor; OC, ONECUT.
      Foxa3 is involved later during development as its expression is positively regulated by HNF1β and HNF1α proteins.
      • Hiemisch H.
      • Schütz G.
      • Kaestner K.H.
      Transcriptional regulation in endoderm development: characterization of an enhancer controlling Hnf3g expression by transgenesis and targeted mutagenesis.
      HNF1β likely regulates Foxa3 expression in early gut endoderm and liver primordium, before HNF1α participates in liver development, consistent with the observed order of their expression patterns in these tissues
      • Ott M.-O.
      • Rey-Campos J.
      • Cereghini S.
      • Yaniv M.
      VHNF1 is expressed in epithelial cells of distinct embryonic origin during development and precedes HNF1 expression.
      • Monaghan A.P.
      • Kaestner K.H.
      • Grau E.
      • Schutz G.
      Postimplantation expression patterns indicate a role for the mouse forkhead/HNF-3 alpha, beta and gamma genes in determination of the definitive endoderm, chordamesoderm and neuroectoderm.
      • Hiemisch H.
      • Schütz G.
      • Kaestner K.H.
      Transcriptional regulation in endoderm development: characterization of an enhancer controlling Hnf3g expression by transgenesis and targeted mutagenesis.
      (Fig. 2A). As Hnf4α precedes Hnf1α expression during early embryonic development, Hnf1α is not likely to participate in the initial activation of Hnf4α. Conversely, HNF1β was found to be a strong inducer of Hnf4α when acting together with GATA6 even though it binds to the same motifs as HNF1α.
      • Hatzis P.
      • Talianidis I.
      Regulatory mechanisms controlling human hepatocyte nuclear factor 4α gene expression.

      Complex interplay amongst the HNFs in the stepwise regulation of liver development

      In the anterior endoderm, Foxa1 and Foxa2 act together with Gata4 to induce Albumin expression, providing the initial specification for liver differentiation.
      • Si-Tayeb K.
      • Lemaigre F.P.
      • Duncan S.A.
      Organogenesis and development of the liver.
      Studies performed in HepG2 cells indicate that FOXA2 binds onto its own promoter to autoregulate its expression. Interestingly, after liver progenitor specification, development of later hepatic differentiation stages is independent of FOXA.
      • Li Z.
      • White P.
      • Tuteja G.
      • Rubins N.
      • Sackett S.
      • Kaestner K.H.
      Foxa1 and Foxa2 regulate bile duct development in mice.
      The liver primordium forms on E8.5–9, beginning with HNF1β–induced liver differentiation.
      • Cereghini S.
      • Ott M.O.
      • Power S.
      • Maury M.
      Expression patterns of vHNF1 and HNF1 homeoproteins in early postimplantation embryos suggest distinct and sequential developmental roles.
      • Lazzaro D.
      • De Simone V.
      • De Magistris L.
      • Lehtonen E.
      • Cortese R.
      LFB1 and LFB3 homeoproteins are sequentially expressed during kidney development.
      HNF1β also cooperates with GATA6 to strongly activate the HNF4α promoter
      • Hatzis P.
      • Talianidis I.
      Regulatory mechanisms controlling human hepatocyte nuclear factor 4α gene expression.
      (Fig. 2B). As liver differentiation occurs, the expression of Hnf1β decreases while that of Hnf1α increases. The balance between HNF1β and HNF1α levels may determine the composition of the transcription initiation complex formed on the HNF4α promoter.
      • Hatzis P.
      • Talianidis I.
      Regulatory mechanisms controlling human hepatocyte nuclear factor 4α gene expression.
      There appears to be a redundancy between Hnf1α and Hnf1β in the regulation of Hnf4α expression as shown in Hnf1α-null mice.
      • Pontoglio M.
      • Barra J.
      • Hadchouel M.
      • Doyen A.
      • Kress C.
      • Bach J.P.
      • et al.
      Hepatocyte nuclear factor 1 inactivation results in hepatic dysfunction, phenylketonuria, and renal Fanconi syndrome.
      However, by adulthood, Hnf1β is no longer expressed
      • Schrem H.
      • Klempnauer J.
      • Borlak J.
      Liver-enriched transcription factors in liver function and development. Part II: the C/EBPs and D site-binding protein in cell cycle control, carcinogenesis, circadian gene regulation, liver regeneration, apoptosis, and liver-specific gene regulation.
      whereas HNF1α continues to cooperate with OC1 to regulate HNF4α gene expression in human liver cells.
      • Hatzis P.
      • Talianidis I.
      Regulatory mechanisms controlling human hepatocyte nuclear factor 4α gene expression.
      (Fig. 2B). The mutagenesis of HNF1α or OC1 binding sites within the HNF4α promoter reduces promoter activity, while the overexpression of these transcription factors activates HNF4α expression.
      Genome-wide promoter occupancy studies (using chromatin immunoprecipitation microarray; ChIP-on-chip) performed in adult human hepatocytes have provided insight into how the liver-specific HNF cross-regulatory network formed by HNF1α, HNF4α and OC1 regulates hepatocyte function
      • Odom D.T.
      • Zizlsperger N.
      • Gordon D.B.
      • Bell G.W.
      • Rinaldi N.J.
      • Murray H.L.
      • et al.
      Control of pancreas and liver gene expression by HNF transcription factors.
      (Fig. 2B). Interestingly, HNF4α was found to bind to more than 40% of the promoters of actively-transcribed genes. It also occupied most of the promoters bound by HNF1α and OC1,
      • Odom D.T.
      • Zizlsperger N.
      • Gordon D.B.
      • Bell G.W.
      • Rinaldi N.J.
      • Murray H.L.
      • et al.
      Control of pancreas and liver gene expression by HNF transcription factors.
      suggesting that HNF4α plays a central role in the transcriptional programme in the liver.
      • Kuo C.J.
      • Conley P.B.
      • Chen L.
      • Sladek F.M.
      • Darnell J.E.
      • Crabtree G.R.
      A transcriptional hierarchy involved in mammalian cell-type specification.
      In addition, other studies have also ascertained that HNF4α binds onto the proximal promoter of the HNF1α gene
      • Tian J.M.
      • Schibler U.
      Tissue-specific expression of the gene encoding hepatocyte nuclear factor 1 may involve hepatocyte nuclear factor 4.
      and that Hnf4α-null hepatocytes express reduced Hnf1α transcripts.
      • Li J.
      • Ning G.
      • Duncan S.A.
      Mammalian hepatocyte differentiation requires the transcription factor HNF-4α.
      In vitro studies performed in HepG2 cells show that a point mutation in the HNF4α binding site of the HNF1α promoter reduces HNF4α binding, leading to reduced activation of HNF1α gene expression.
      • Lausen J.
      • Thomas H.
      • Lemm I.
      • Bulman M.
      • Borgschulze M.
      • Lingott A.
      • et al.
      Naturally occurring mutations in the human HNF4α gene impair the function of the transcription factor to a varying degree.
      HNF4α is known to bind to the HNF1α promoter via an evolutionarily conserved cis sequence element, regulating its transcription in hepatocytes.
      • Kuo C.J.
      • Conley P.B.
      • Chen L.
      • Sladek F.M.
      • Darnell J.E.
      • Crabtree G.R.
      A transcriptional hierarchy involved in mammalian cell-type specification.
      HNF1α also autoregulates its own expression in hepatocytes.
      • Minra N.
      • Tanaka K.
      Analysis of the rat hepatocyte nuclear factor (HNF) 1 gene promoter: synergistic activation by HNF4 and HNF1 proteins.
      In the liver, Oc1 and Oc2 have redundant roles in the differentiation of both hepatocytes and biliary cells.
      • Clotman F.
      • Jacquemin P.
      • Plumb-Rudewiez N.
      • Pierreux C.E.
      • Van der Smissen P.
      • Dietz H.C.
      • et al.
      Control of liver cell fate decision by a gradient of TGFβ signaling modulated by Onecut transcription factors.
      Oc1 stimulates Hnf1β expression in the liver for intrahepatic bile duct morphogenesis.
      • Clotman F.
      • Lannoy V.J.
      • Reber M.
      • Cereghini S.
      • Cassiman D.
      • Jacquemin P.
      • et al.
      The onecut transcription factor HNF6 is required for normal development of the biliary tract.
      This is consistent with the observation that Oc1-null mice exhibit diminished Hnf1β expression, failed development of the gallbladder, and severe abnormalities in both extra- and intrahepatic bile ducts
      • Clotman F.
      • Lannoy V.J.
      • Reber M.
      • Cereghini S.
      • Cassiman D.
      • Jacquemin P.
      • et al.
      The onecut transcription factor HNF6 is required for normal development of the biliary tract.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      (Fig. 2B). Studies in Oc1-null/Oc2-null mice further revealed that Oc1 and Hnf1 regulation of Hnf4α7 expression (P2 promoter-driven form) is suppressed by Hnf4α1 (P1 promoter-driven form) later in liver maturation.
      • Briançon N.
      • Bailly A.
      • Clotman F.
      • Jacquemin P.
      • Lemaigre F.P.
      • Weiss M.C.
      Expression of the α7 Isoform of Hepatocyte Nuclear Factor (HNF) 4 Is Activated by HNF6/OC-2 and HNF1 and Repressed by HNF4α1 in the Liver.

      The HNF1α-HNF4α and OC1-HNF1β cross-regulatory network in the pancreas

      In the pancreas, Hnf1α is known to regulate the expression of Hnf4α. Hnf1α-null mice display reduced expression of Hnf4α, Hnf4γ and Foxa3 transcripts in the islets.
      • Boj S.F.
      • Párrizas M.
      • Maestro M.A.
      • Ferrer J.
      A transcription factor regulatory circuit in differentiated pancreatic cells.
      • Shih D.Q.
      • Screenan S.
      • Munoz K.N.
      • Philipson L.
      • Pontoglio M.
      • Yaniv M.
      • et al.
      Loss of HNF-1α function in mice leads to abnormal expression of genes involved in pancreatic islet development and metabolism.
      Specifically, HNF1α regulates the P2 promoter of the HNF4α gene in the pancreas.
      • Thomas H.
      • Jaschkowitz K.
      • Bulman M.
      • Frayling T.M.
      • Mitchell S.M.
      • Roosen S.
      • et al.
      A distant upstream promoter of the HNF-4alpha gene connects the transcription factors involved in maturity-onset diabetes of the young.
      • Boj S.F.
      • Párrizas M.
      • Maestro M.A.
      • Ferrer J.
      A transcription factor regulatory circuit in differentiated pancreatic cells.
      A naturally-occurring mutation in the human HNF4α P2 promoter, which abolishes binding by HNF1α, is known to result in maturity onset diabetes of the young 1 (MODY1). The significance of this HNF1α-HNF4α gene regulation is reflected by similarities in phenotypes caused by the disruption of Hnf4α transcription by Hnf1α and loss-of-function Hnf1α mutations.
      • Hansen S.K.
      • Párrizas M.
      • Jensen M.L.
      • Pruhova S.
      • Ek J.
      • Boj S.F.
      • et al.
      Genetic evidence that HNF-1α–dependent transcriptional control of HNF-4α is essential for human pancreatic β cell function.
      Conversely, in the liver, expression of the P1 promoter-driven form of Hnf4α is not activated by Hnf1α.
      • Thomas H.
      • Jaschkowitz K.
      • Bulman M.
      • Frayling T.M.
      • Mitchell S.M.
      • Roosen S.
      • et al.
      A distant upstream promoter of the HNF-4alpha gene connects the transcription factors involved in maturity-onset diabetes of the young.
      • Boj S.F.
      • Párrizas M.
      • Maestro M.A.
      • Ferrer J.
      A transcription factor regulatory circuit in differentiated pancreatic cells.
      • Jiang S.
      • Tanaka T.
      • Iwanari H.
      • Hotta H.
      • Yamashita H.
      • Kumakura J.
      • et al.
      Expression and localization of P1 promoter-driven hepatocyte nuclear factor-4α (HNF4α) isoforms in human and rats.
      These studies reflect tissue-specific differences in the HNF1α-HNF4α-mediated transcriptional regulatory network (Fig. 2B, C).
      The cross-regulatory loop between the two transcription factors is made even more complex as HNF4α is also an essential activator of HNF1α gene expression (Fig. 2B). The AF-2 module and LBD of HNF4α collectively mediate the regulation of HNF1α gene expression 73. HNF4α can facilitate the recruitment of transcriptional coactivators such as p300 to enhance the activation of HNF1α expression. In fact, direct protein-protein interaction also occurs as HNF4α can bind cooperatively with HNF1α, thereby improving the docking of p300 onto the HNF4α-HNF1α complex, regulating downstream target gene expression in a synergistic manner.
      • Eeckhoute J.
      • Formstecher P.
      • Laine B.
      Hepatocyte nuclear factor 4α enhances the hepatocyte nuclear factor 1α-mediated activation of transcription.
      Moreover, studies have shown that HNF4α and HNF1α share common transcriptomic signatures in the pancreatic islets and that their functional activities are interdependent, though this is likely to be target-specific.
      • Boj S.F.
      • Petrov D.
      • Ferrer J.
      Epistasis of transcriptomes reveals synergism between transcriptional activators Hnf1α and Hnf4α.
      This is important for our understanding of how defects in either HNF4α or HNF1α can lead to islet dysfunction and diabetes (discussed in a later section).
      Oc1 and Oc2 exert partially redundant functions in dorsal and ventral pancreas organogenesis.
      • Vanhorenbeeck V.
      • Jenny M.
      • Cornut J.-F.
      • Gradwohl G.
      • Lemaigre F.P.
      • Rousseau G.G.
      • et al.
      Role of the Onecut transcription factors in pancreas morphogenesis and in pancreatic and enteric endocrine differentiation.
      The Onecut family members are also involved in initiating expression of the key pancreatic transcription factor Pdx1. In contrast to observations in liver cells, the expression of Oc1 in the pancreatic precursor cells is activated by Hnf1β, leading to the expression of Pdx1, which is critical for the specification of pancreatic cell fate.
      • Poll A.V.
      • Pierreux C.E.
      • Lokmane L.
      • Haumaitre C.
      • Achouri Y.
      • Jacquemin P.
      • et al.
      A vHNF1/TCF2-HNF6 cascade regulates the transcription factor network that controls generation of pancreatic precursor cells.
      At later stages of development, Oc1 regulates pancreatic endocrine differentiation via Ngn3 expression.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      The Onecut transcription factor HNF-6 (OC-1) is required for timely specification of the pancreas and acts upstream of Pdx-1 in the specification cascade.
      OC1 also binds to the proximal promoter region of FOXA2 to regulate its expression in the gut endoderm
      • Samadani U.
      • Costa R.H.
      The transcriptional activator hepatocyte nuclear factor 6 regulates liver gene expression.
      (Fig. 2C). Evidently, OC1 and FOXA2 are uniquely co-expressed to regulate cellular identity in hepatocytes, pancreatic and intestinal cells.
      • Rausa F.
      • Samadani U.
      • Ye H.
      • Lim L.
      • Fletcher C.F.
      • Jenkins N.A.
      • et al.
      The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3β in the developing murine liver and pancreas.

      Complex differential regulation of HNF4α isoforms in the liver and pancreas

      The multiple isoforms of HNF4α generated via alternative splicing, highlight the regulatory complexity of HNF4 expression and function. The consensus is that isoforms HNF4α1-3 derived from the HNF4A P1 promoter are predominantly expressed in adult liver (also in kidney) while isoforms HNF4α7-9 derived from the P2 promoter are active in both embryonic liver and in pancreatic β cells
      • Thomas H.
      • Jaschkowitz K.
      • Bulman M.
      • Frayling T.M.
      • Mitchell S.M.
      • Roosen S.
      • et al.
      A distant upstream promoter of the HNF-4alpha gene connects the transcription factors involved in maturity-onset diabetes of the young.
      (Fig. 3). Binding sites for HNF1, OC1 and GATA6 have all been reported on the P1 promoter.
      • Kyrmizi I.
      • Hatzis P.
      • Katrakili N.
      • Tronche F.
      • Gonzalez F.J.
      • Talianidis I.
      Plasticity and expanding complexity of the hepatic transcription factor network during liver development.
      In the embryonic liver, it is suggested that HNF1β cooperates with GATA6 to activate HNF4α1 expression 55 (Figs. 2A, 3). However, in the adult liver, synergism between FOXA2 and OC1 drives HNF4α1 expression 79 (Figs. 2B, 3).
      Figure thumbnail gr3
      Fig. 3Complex differential regulation of HNF4α isoforms in the liver and pancreas. HNF1, OC1, GATA6 and FOXA2 are involved in activating P1 promoter-driven HNF4α expression in the embryonic liver. HNF1β cooperates with GATA6 to activate P1-HNF4α while in the adult liver, synergism between FOXA2 and OC1 drives P1-HNF4α expression. Similarly, P2-HNF4α expression is driven by OC1, HNF1β (embryonic liver) and HNF1α (adult liver). The HNF4α P2 promoter contains binding sites for transcription factors in which their mutations lead to MODY, namely HNF1α (MODY3), HNF1β (MODY5) and PDX1 (MODY4). FOXA, forkhead box A; HNF, hepatocyte nuclear factor; MODY, maturity onset diabetes of the young; OC, ONECUT.
      The alternative P2 promoter, which gives rise to HNF4α7, can be activated by HNF1β and OC1 in the embryonic liver.
      • Kyrmizi I.
      • Hatzis P.
      • Katrakili N.
      • Tronche F.
      • Gonzalez F.J.
      • Talianidis I.
      Plasticity and expanding complexity of the hepatic transcription factor network during liver development.
      Whilst, in the pancreas, the P2 promoter is activated by pancreatic transcription factors HNF1α, OC1 and PDX1 (Fig. 3). The striking developmental and tissue-specific variation in the expression of HNF4α isoforms can play a role in moderating disease outcome depending on the precise location of the disease-causing mutation and its consequential effects on isoform expression and activity.
      • Harries L.W.
      • Locke J.M.
      • Shields B.
      • Hanley N.A.
      • Hanley K.P.
      • Steele A.
      • et al.
      Mutation carriers is moderated by the expression of HNF4A isoforms from the P1 promoter during fetal development.

      HNF1β and HNF1α function in the kidney

      Hnf1β is expressed in the kidney and controls the expression of cystic genes such as Pkhd1, Pkd2 and Umod.
      • Coffinier C.
      • Barra J.
      • Babinet C.
      • Yaniv M.
      Expression of the vHNF1/HNF1β homeoprotein gene during mouse organogenesis.
      • Gresh L.
      • Fischer E.
      • Reimann A.
      • Tanguy M.
      • Garbay S.
      • Shao X.
      • et al.
      A transcriptional network in polycystic kidney disease.
      Hnf1β directly regulates the expression of Pkhd1 in tubular epithelial cells that are involved in tubulogenesis in the kidney,
      • Hiesberger T.
      • Shao X.
      • Gourley E.
      • Reimann A.
      • Pontoglio M.
      • Igarashi P.
      Role of the hepatocyte nuclear factor-1β (HNF-1β) C-terminal domain in Pkhd1 (ARPKD) gene transcription and renal cystogenesis.
      • Igarashi P.
      • Shao X.
      • McNally B.T.
      • Hiesberger T.
      Roles of HNF-1beta in kidney development and congenital cystic diseases.
      and Socs3 which is essential for renal repair.
      • Faguer S.
      • Mayeur N.
      • Casemayou A.
      • Pageaud A.L.
      • Courtellemont C.
      • Cartery C.
      • et al.
      Hnf-1beta transcription factor is an early hif-1alpha-independent marker of epithelial hypoxia and controls renal repair.
      Although Hnf1α is expressed in the developing kidney and plays a role in kidney development, it is also a key regulator of numerous essential metabolic processes during adulthood. These include maintaining glucose homeostasis through regulation of the glucose-6-phosphatase system and the glucose transporter, as well as regulating other transporters that impact on the metabolism of amino acids because of impaired renal reabsorption.
      • Pontoglio M.
      • Barra J.
      • Hadchouel M.
      • Doyen A.
      • Kress C.
      • Bach J.P.
      • et al.
      Hepatocyte nuclear factor 1 inactivation results in hepatic dysfunction, phenylketonuria, and renal Fanconi syndrome.
      • Bonzo J.A.
      • Patterson A.D.
      • Krausz K.W.
      • Gonzalez F.J.
      Metabolomics identifies novel Hnf1alpha-dependent physiological pathways in vivo.
      • Hiraiwa H.
      • Pan C.-J.
      • Lin B.
      • Akiyama T.E.
      • Gonzalez F.J.
      • Chou J.Y.
      A molecular link between the common phenotypes of type 1 glycogen storage disease and HNF1α-null Mice.
      • Pontoglio M.
      • Prie D.
      • Cheret C.
      • Doyen A.
      • Leroy C.
      • Froguel P.
      • et al.
      HNF1alpha controls renal glucose reabsorption in mouse and man.
      For instance, Hnf1α-null mice express lower levels of Npt1 and Npt4 genes, members of the sodium/phosphate cotransporter family. Npt1 is likely to be a direct target of HNF1α.
      • Cheret C.
      • Doyen A.
      • Yaniv M.
      • Pontoglio M.
      Hepatocyte nuclear factor 1 alpha controls renal expression of the Npt1-Npt4 anionic transporter locus.
      As these observations have been obtained from knockout mouse models, the physiological impacts of HNF1A mutations on kidney function in humans will require further in-depth investigation.
      Overall, it is clear that the HNFs contribute to a complex cross-regulatory network exhibiting both specificity and redundancy across different tissue types. Nevertheless, most of these insights were gained from studies in rodents, and it is important to understand if the same applies to humans. This is important for the understanding of the roles of HNFs in disease pathophysiology, as heterozygous mutations in humans result in diseases that are not recapitulated in rodent models.

      The role of mutations in the HNF family members in health and disease

      Mutations in several HNF transcription factors are well-known to cause MODY, a rare autosomal dominantly inherited form of diabetes that often occurs before 25 years of age, typically leading to dysfunctional pancreatic β cells with accompanying liver and/or kidney defects. These MODY genes include HNF1α (MODY3), HNF1β (MODY5) and HNF4α (MODY1)
      • Kyrmizi I.
      • Hatzis P.
      • Katrakili N.
      • Tronche F.
      • Gonzalez F.J.
      • Talianidis I.
      Plasticity and expanding complexity of the hepatic transcription factor network during liver development.
      • Fajans S.S.
      • Bell G.I.
      • Polonsky K.S.
      Molecular mechanisms and clinical pathophysiology of maturity-onset diabetes of the young.
      (Table 3, Table 4). In the previous section, we discussed the complex HNF cross-regulatory transcriptional network that is activated in a developmental stage- and tissue-specific manner, focusing on the liver, pancreas and kidney. The multifactorial roles of these HNFs imply that a single inactivating mutation is likely to affect the function of more than one tissue (liver and beyond). Much of the existing knowledge on the functional mechanisms underlying the HNFs has been obtained from rodent studies because of the inaccessibility of human tissues. However, rare genetic mutations identified in HNF genes that result in pathophysiological conditions have provided a ‘natural’ human disease model for the study of gene function. Here, we compare the phenotypes that arise from the perturbations in HNF genes in mice and humans. We also highlight the mechanistic insights gained from rodent studies and the limitations of rodent models, which may not necessarily recapitulate human HNF physiology (Table 3, Table 4).
      Table 3Summary of HNF mutations and the respective phenotypes observed in the affected organ(s) in mice.
      Hnf mutations in mice (genotype)Phenotype(s)
      Hnf1
      Hnf1α (+/−)-
      Hnf1α (−/−)Develop severe diabetes 2 weeks after birth; develop liver and reproductive system dysfunction, rendering the mice sterile.
      • Lee Y.-H.
      • Sauer B.
      • Gonzalez F.J.
      Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1α knockout mouse.
      ,
      • Pontoglio M.
      • Sreenan S.
      • Roe M.
      • Pugh W.
      • Ostrega D.
      • Doyen A.
      • et al.
      Defective insulin secretion in hepatocyte nuclear factor 1alpha-deficient mice.
      Reduced β cell mass and develop renal Fanconi syndrome.
      • Pontoglio M.
      • Pausa M.
      • Doyen A.
      • Viollet B.
      • Yaniv M.
      • Tedesco F.
      Hepatocyte nuclear factor 1α controls the expression of terminal complement genes.
      Establish Laron-type dwarfism; the mice showed no signs of renal dysfunction.
      • Lee Y.-H.
      • Sauer B.
      • Gonzalez F.J.
      Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1α knockout mouse.
      Display reduced expression of Glut2 (Slc2a2) glucose transporter and L-type pyruvate kinase (pklr) genes specifically in pancreatic insulin-producing cells.
      • Parrizas M.
      • Maestro M.A.
      • Boj S.F.
      • Paniagua A.
      • Casamitjana R.
      • Gomis R.
      • et al.
      Hepatic nuclear factor 1-alpha directs nucleosomal hyperacetylation to its tissue-specific transcriptional targets.
      Hnf1β (+/−)-
      Hnf1β (−/−)Die at E5.5-E6 due to abnormal or absent extraembryonic endoderm.
      • Barbacci E.
      • Reber M.
      • Ott M.O.
      • Breillat C.
      • Huetz F.
      • Cereghini S.
      Variant hepatocyte nuclear factor 1 is required for visceral endoderm specification.
      Hnf1β (Tetraploid complementation)Fail to develop ventral pancreas and do not form endocrine cells.
      • Haumaitre C.
      • Barbacci E.
      • Jenny M.
      • Ott M.O.
      • Gradwohl G.
      • Cereghini S.
      Lack of TCF2/vHNF1 in mice leads to pancreas agenesis.
      No hepatic bud formation, defective hepatic specification of the entire ventral endoderm,
      • Lokmane L.
      • Haumaitre C.
      • Garcia-Villalba P.
      • Anselme I.
      • Schneider-Maunoury S.
      • Cereghini S.
      Crucial role of vHNF1 in vertebrate hepatic specification.
      affected organs include lung, gall bladder and ventral pancreas. Ureter bud branching morphogenesis disruption and absence of nephrogenesis induction.
      • Lokmane L.
      • Heliot C.
      • Garcia-Villalba P.
      • Fabre M.
      • Cereghini S.
      vHNF1 functions in distinct regulatory circuits to control ureteric bud branching and early nephrogenesis.
      Hnf1β (Targeted deletion)Neonatal cholestasis and jaundice due to an abnormal development of gall bladder and intrahepatic bile ducts.
      • Coffinier C.
      • Gresh L.
      • Fiette L.
      • Tronche F.
      • Schütz G.
      • Babinet C.
      • et al.
      Bile system morphogenesis defects and liver dysfunction upon targeted deletion of HNF1β.
      Foxa
      Foxa1 (+/−)-
      Foxa1 (−/−)Hypoglycaemic and die between postnatal day 2 to 12.
      • Friedman J.R.
      • Kaestner K.H.
      The Foxa family of transcription factors in development and metabolism.
      • Kaestner K.H.
      • Katz J.
      • Liu Y.
      • Drucker D.J.
      • Schütz G.
      Inactivation of the winged helix transcription factor HNF3α affects glucose homeostasis and islet glucagon gene expression in vivo.
      Develop nephrogenic diabetes insipidus shortly after birth.
      • Behr R.
      • Brestelli J.
      • Fulmer J.T.
      • Miyawaki N.
      • Kleyman T.R.
      • Kaestner K.H.
      Mild nephrogenic diabetes insipidus caused by foxa1 deficiency.
      Develop HCC in a sexual dimorphic manner.
      • Li Z.
      • Tuteja G.
      • Schug J.
      • Kaestner K.H.
      Foxa1 and Foxa2 are essential for sexual dimorphism in liver cancer.
      No mature luminal epithelial cells observed in the prostate.
      • Gao N.
      • Ishii K.
      • Mirosevich J.
      • Kuwajima S.
      • Oppenheimer S.R.
      • Roberts R.L.
      • et al.
      Forkhead box A1 regulates prostate ductal morphogenesis and promotes epithelial cell maturation.
      Foxa2 (+/−)-
      Foxa2 (−/−)Embryonic lethal after gastrulation, mice established absence of organized node and notochord formation and subsequent defects in dorsal-ventral patterning in the neural tube.
      • Ang S.L.
      • Rossant J.
      HNF-3 beta is essential for node and notochord formation in mouse development.
      Develop HCC in a sexual dimorphic manner.
      • Li Z.
      • Tuteja G.
      • Schug J.
      • Kaestner K.H.
      Foxa1 and Foxa2 are essential for sexual dimorphism in liver cancer.
      Foxa2 (Targeted deletion)Targeted deletion of Foxa2 in β cells results in hypoglycaemia and relative hyperinsulinemia condition.
      • Lantz K.A.
      • Vatamaniuk M.Z.
      • Brestelli J.E.
      • Friedman J.R.
      • Matschinsky F.M.
      • Kaestner K.H.
      Foxa2 regulates multiple pathways of insulin secretion.
      Foxa3 (+/− or −/−)Display reduced male fertility and increased germ cell apoptosis.
      • Behr R.
      • Brestelli J.
      • Fulmer J.T.
      • Miyawaki N.
      • Kleyman T.R.
      • Kaestner K.H.
      Mild nephrogenic diabetes insipidus caused by foxa1 deficiency.
      Hnf4
      Hnf4α (+/−)-
      Hnf4α (−/−)Embryonic lethal.
      • Chen W.S.
      • Manova K.
      • Weinstein D.C.
      • Duncan S.A.
      • Plump A.S.
      • Prezioso V.R.
      • et al.
      Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
      Hnf4α (Tetraploid complementation)Hnf4α-null foetal liver failed to express a large array of genes that are associated with mature hepatocyte function
      • Li J.
      • Ning G.
      • Duncan S.A.
      Mammalian hepatocyte differentiation requires the transcription factor HNF-4α.
      Hnf4α (Targeted deletion)Deletion of Hnf4α in the liver results in steatosis, severe disruption of gluconeogenesis and death at 6–8 weeks of age.
      • Chen W.S.
      • Manova K.
      • Weinstein D.C.
      • Duncan S.A.
      • Plump A.S.
      • Prezioso V.R.
      • et al.
      Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
      • Hayhurst G.P.
      • Lee Y.-H.
      • Lambert G.
      • Ward J.M.
      • Gonzalez F.J.
      Hepatocyte nuclear factor 4α (Nuclear Receptor 2A1) is essential for maintenance of hepatic gene expression and lipid homeostasis.
      Leads to HCC. Knockdown of Hnf4α in the kidney results in apoptosis in the condensed mesenchyme.
      • Kanazawa T.
      • Konno A.
      • Hashimoto Y.
      • Kon Y.
      Hepatocyte nuclear factor 4 alpha is related to survival of the condensed mesenchyme in the developing mouse kidney.
      Hnf4γ (−/−)Appears to have neurological defects, with reduced locomotor activity at night.
      • Gerdin A.K.
      • Surve V.V.
      • Jönsson M.
      • Bjursell M.
      • Björkman M.
      • Edenro A.
      • et al.
      Phenotypic screening of hepatocyte nuclear factor (HNF) 4-γ receptor knockout mice.
      Onecut
      Oc1 (−/−)

      Oc2 (−/−)
      Severe defects in the liver and die before postnatal day 10.
      • Clotman F.
      • Lannoy V.J.
      • Reber M.
      • Cereghini S.
      • Cassiman D.
      • Jacquemin P.
      • et al.
      The onecut transcription factor HNF6 is required for normal development of the biliary tract.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      • Margagliotti S.
      • Clotman F.
      • Pierreux C.E.
      • Beaudry J.-B.
      • Jacquemin P.
      • Rousseau G.G.
      • et al.
      The Onecut transcription factors HNF-6/OC-1 and OC-2 regulate early liver expansion by controlling hepatoblast migration.
      Perturbed islet morphology
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      The Onecut transcription factor HNF-6 (OC-1) is required for timely specification of the pancreas and acts upstream of Pdx-1 in the specification cascade.
      and severe diabetes reported.
      • Clotman F.
      • Lannoy V.J.
      • Reber M.
      • Cereghini S.
      • Cassiman D.
      • Jacquemin P.
      • et al.
      The onecut transcription factor HNF6 is required for normal development of the biliary tract.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      Pancreatic hypoplasia.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      The Onecut transcription factor HNF-6 (OC-1) is required for timely specification of the pancreas and acts upstream of Pdx-1 in the specification cascade.
      Hindlimb paresis at birth.
      • Audouard E.
      • Schakman O.
      • René F.
      • Huettl R.-E.
      • Huber A.B.
      • Loeffler J.-P.
      • et al.
      The onecut transcription factor HNF-6 regulates in motor neurons the formation of the neuromuscular junctions.
      (-) indicates no difference observed in mutants compared to the healthy wild-type littermates. Foxa, forkhead box A; HCC, hepatocellular carcinoma; Hnf, hepatocyte nuclear factor; Oc, Onecut.
      Table 4Summary of the HNF mutations and the respective phenotypes reported in the affected organ(s) in humans.
      HNF mutations in humans (genotype)Phenotype(s)
      HNF1
      HNF1α
      There is no reported homozygous mutation in HNF1α, HNF1β and HNF4α. FOXA, forkhead box A; HNF, hepatocyte nuclear factor; MODY, maturity onset diabetes of the young; OC, ONECUT.
      (+/−)
      MODY3.
      • Fajans S.S.
      • Bell G.I.
      • Polonsky K.S.
      Molecular mechanisms and clinical pathophysiology of maturity-onset diabetes of the young.
      Some MODY3 patients also suffer from renal dysplasia, growth hormone deficiency and hypothyroidism. Patients display infantile uterus and unidentifiable ovary phenotypes, leading to infertility.
      • Simms R.J.
      • Sayer J.A.
      • Quinton R.
      • Walker M.
      • Ellard S.
      • Goodship T.H.J.
      Monogenic diabetes, renal dysplasia and hypopituitarism: a patient with a mutation.
      Patients develop hepatocellular adenoma.
      • Bioulac-Sage P.
      • Balabaud C.
      • Zucman-Rossi J.
      Subtype classification of hepatocellular adenoma.
      • Bioulac-Sage P.
      • Laumonier H.
      • Couchy G.
      • Le Bail B.
      • Sa Cunha A.
      • Rullier A.
      • et al.
      Hepatocellular adenoma management and phenotypic classification: The Bordeaux experience.
      HNF1β
      There is no reported homozygous mutation in HNF1α, HNF1β and HNF4α. FOXA, forkhead box A; HNF, hepatocyte nuclear factor; MODY, maturity onset diabetes of the young; OC, ONECUT.
      (+/−)
      Patients establish partial pancreatic agenesis-mediated MODY5, renal cystic disease, occasional genital tract abnormalities and abnormal liver function.
      • Beards F.
      • Frayling T.
      • Bulman M.
      • Horikawa Y.
      • Allen L.
      • Appleton M.
      • et al.
      Mutations in hepatocyte nuclear factor 1beta are not a common cause of maturity-onset diabetes of the young in the U.K..
      • Bingham C.
      • Hattersley A.T.
      Renal cysts and diabetes syndrome resulting from mutations in hepatocyte nuclear factor-1β.
      • Edghill E.L.
      • Bingham C.
      • Ellard S.
      • Hattersley A.T.
      Mutations in hepatocyte nuclear factor-1β and their related phenotypes.
      • Gonc E.N.
      • Ozturk B.B.
      • Haldorsen I.S.
      • Molnes J.
      • Immervoll H.
      • Raeder H.
      • et al.
      HNF1B mutation in a Turkish child with renal and exocrine pancreas insufficiency, diabetes and liver disease.
      FOXA
      FOXA1 (+/−)Associated with breast and prostate cancer in humans.
      • Robinson J.L.L.
      • Holmes K.A.
      • Carroll J.S.
      FOXA1 mutations in hormone-dependent cancers.
      FOXA1 or FOXA2 (+/−)Patients develop hepatocellular carcinoma in a sexual dimorphic manner, with higher occurrence in males.
      • Li Z.
      • Tuteja G.
      • Schug J.
      • Kaestner K.H.
      Foxa1 and Foxa2 are essential for sexual dimorphism in liver cancer.
      FOXA3 (−)No reported cases of mutation leading to human disease yet.
      HNF4
      HNF4α
      There is no reported homozygous mutation in HNF1α, HNF1β and HNF4α. FOXA, forkhead box A; HNF, hepatocyte nuclear factor; MODY, maturity onset diabetes of the young; OC, ONECUT.
      (+/−)
      MODY1.
      • Frayling T.M.
      • Evans J.C.
      • Bulman M.P.
      • Pearson E.
      • Allen L.
      • Owen K.
      • et al.
      Beta-cell genes and diabetes: molecular and clinical characterization of mutations in transcription factors.
      Patients develop typical dominant Fanconi syndrome in addition to a β cell phenotype.
      • Hamilton A.J.
      • Bingham C.
      • McDonald T.J.
      • Cook P.R.
      • Caswell R.C.
      • Weedon M.N.
      • et al.
      The HNF4A R76W mutation causes atypical dominant Fanconi syndrome in addition to a β cell phenotype.
      HNF4γ (−)No reported cases of HNF4γ polymorphisms leading to human disease yet.
      HNF6
      HNF6 (−)No reported cases of HNF6 polymorphisms leading to human disease yet.
      * There is no reported homozygous mutation in HNF1α, HNF1β and HNF4α. FOXA, forkhead box A; HNF, hepatocyte nuclear factor; MODY, maturity onset diabetes of the young; OC, ONECUT.

      HNF1α mutations in mice and men

      Heterozygous HNF1α mutations in humans give rise to MODY3, the most common form of MODY. Patients with MODY3 exhibit progressive failure in glycaemic control with impaired β cell function as the primary cause of diabetes.
      • Fajans S.S.
      • Bell G.I.
      • Polonsky K.S.
      Molecular mechanisms and clinical pathophysiology of maturity-onset diabetes of the young.
      • Lehto M.
      • Tuomi T.
      • Mahtani M.M.
      • Widen E.
      • Forsblom C.
      • Sarelin L.
      • et al.
      Characterization of the MODY3 phenotype. Early-onset diabetes caused by an insulin secretion defect.
      A considerable number of patients also develop microvascular and macrovascular complications often seen in patients with type 1 and 2 diabetes (T1D and T2D).
      • Steele A.M.
      • Shields B.M.
      • Shepherd M.
      • Ellard S.
      • Hattersley A.T.
      • Pearson E.R.
      Increased all-cause and cardiovascular mortality in monogenic diabetes as a result of mutations in the HNF1A gene.
      In MODY3, the progressive loss of the insulin secretory capacity can be treated with sulfonylureas with excellent results.
      • Pearson E.R.
      • Liddell W.G.
      • Shepherd M.
      • Corrall R.J.
      • Hattersley A.T.
      Sensitivity to sulphonylureas in patients with hepatocyte nuclear factor-1alpha gene mutations: evidence for pharmacogenetics in diabetes.
      • Sovik O.
      • Njolstad P.
      • Folling I.
      • Sagen J.
      • Cockburn B.N.
      • Bell G.I.
      Hyperexcitability to sulphonylurea in MODY3.
      In fact, MODY is often misdiagnosed as T1D or T2D. In contrast, heterozygous Hnf1α knockout mice do not exhibit a disease phenotype, whereas homozygous Hnf1α knockout mice exhibit stunted growth, reduced size and only weigh ∼50–60% of their wild-type counterparts by five weeks.
      • Lee Y.-H.
      • Sauer B.
      • Gonzalez F.J.
      Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1α knockout mouse.
      Hnf1α-null mice also exhibit abnormal glucose-stimulated insulin secretion and develop diabetes two weeks after birth.
      • Lee Y.-H.
      • Sauer B.
      • Gonzalez F.J.
      Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1α knockout mouse.
      • Pontoglio M.
      • Sreenan S.
      • Roe M.
      • Pugh W.
      • Ostrega D.
      • Doyen A.
      • et al.
      Defective insulin secretion in hepatocyte nuclear factor 1alpha-deficient mice.
      These mice have reduced β cell mass and express low levels of insulin and insulin-like growth factor-1 (Igf1).
      • Servitja J.-M.
      • Pignatelli M.
      • Maestro M.Á.
      • Cardalda C.
      • Boj S.F.
      • Lozano J.
      • et al.
      Hnf1α (MODY3) controls tissue-specific transcriptional programs and exerts opposed effects on cell growth in pancreatic islets and liver.
      Some patients with MODY3 also suffer from renal dysplasia, growth hormone deficiency and hypothyroidism. Other abnormalities include infantile uterus and unidentifiable ovaries, leading to infertility – a phenotype also observed in Hnf1α-null mice.
      • Lee Y.-H.
      • Sauer B.
      • Gonzalez F.J.
      Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1α knockout mouse.
      • Simms R.J.
      • Sayer J.A.
      • Quinton R.
      • Walker M.
      • Ellard S.
      • Goodship T.H.J.
      Monogenic diabetes, renal dysplasia and hypopituitarism: a patient with a mutation.
      Hnf1α-null mice also develop abnormalities in the reproductive system, rendering them sterile.
      • Lee Y.-H.
      • Sauer B.
      • Gonzalez F.J.
      Laron dwarfism and non-insulin-dependent diabetes mellitus in the Hnf-1α knockout mouse.
      • Pontoglio M.
      • Sreenan S.
      • Roe M.
      • Pugh W.
      • Ostrega D.
      • Doyen A.
      • et al.
      Defective insulin secretion in hepatocyte nuclear factor 1alpha-deficient mice.
      One reason for the observed glycosuria (excretion of glucose into the urine) could be a low renal threshold for glucose, possibly because of decreased expression of SGLT2 (SLC5A2) and decreased glucose reabsorption in the proximal tubules.
      • Pontoglio M.
      • Prie D.
      • Cheret C.
      • Doyen A.
      • Leroy C.
      • Froguel P.
      • et al.
      HNF1alpha controls renal glucose reabsorption in mouse and man.
      In the kidney, HNF1α binds to the SGLT2 promoter.
      • Pontoglio M.
      • Prie D.
      • Cheret C.
      • Doyen A.
      • Leroy C.
      • Froguel P.
      • et al.
      HNF1alpha controls renal glucose reabsorption in mouse and man.
      Hnf1α-null mice express reduced Sglt2 transcripts in the tubular cells.
      • Pontoglio M.
      • Prie D.
      • Cheret C.
      • Doyen A.
      • Leroy C.
      • Froguel P.
      • et al.
      HNF1alpha controls renal glucose reabsorption in mouse and man.
      Complete ablation of HNF1α results in severe renal proximal tubular dysfunction in the kidney, which can be likened to the clinical phenotype observed in human patients with Fanconi syndrome.
      • Pontoglio M.
      • Barra J.
      • Hadchouel M.
      • Doyen A.
      • Kress C.
      • Bach J.P.
      • et al.
      Hepatocyte nuclear factor 1 inactivation results in hepatic dysfunction, phenylketonuria, and renal Fanconi syndrome.
      However, Lee et al. (1998) reported that the Hnf1α-null mice show no signs of renal dysfunction, which is quite surprising as it is in contrast with previous results showing that HNF1α inactivation leads to hepatic dysfunction, phenylketonuria and renal Fanconi syndrome.
      • Pontoglio M.
      • Barra J.
      • Hadchouel M.
      • Doyen A.
      • Kress C.
      • Bach J.P.
      • et al.
      Hepatocyte nuclear factor 1 inactivation results in hepatic dysfunction, phenylketonuria, and renal Fanconi syndrome.
      HNF1α mutations have also been reported in 35–40% of patients with hepatocellular adenoma (HCA).
      • Kim H.
      • Jang J.-J.
      • Kim D.-S.
      • Yeom B.W.
      • Won N.H.
      Clinicopathological analysis of hepatocellular adenoma according to new bordeaux classification: report of eight korean cases.
      In contrast to MODY3, bi-allelic inactivation of HNF1α has been identified in tumour tissue and this is often a result of two inactivating somatic mutations, and on rare occasions one germline mutation with a second inactivating mutation. These results suggest that HNF1α could play a role as a tumour suppressor gene in the liver.
      • Bioulac-Sage P.
      • Balabaud C.
      • Zucman-Rossi J.
      Subtype classification of hepatocellular adenoma.
      • Bluteau O.
      • Jeannot E.
      • Bioulac-Sage P.
      • Marques J.M.
      • Blanc J.F.
      • Bui H.
      • et al.
      Bi-allelic inactivation of TCF1 in hepatic adenomas.
      Individuals with germline HNF1α mutations develop HCA at a younger age and usually have a family history of liver adenomatosis.
      • Bioulac-Sage P.
      • Balabaud C.
      • Zucman-Rossi J.
      Subtype classification of hepatocellular adenoma.
      • Bioulac-Sage P.
      • Laumonier H.
      • Couchy G.
      • Le Bail B.
      • Sa Cunha A.
      • Rullier A.
      • et al.
      Hepatocellular adenoma management and phenotypic classification: The Bordeaux experience.
      Furthermore, HNF1α mutations result in increased fatty acid synthesis and perturb fatty acid transportation, leading to lipid accumulation.
      • Rebouissou S.
      • Imbeaud S.
      • Balabaud C.
      • Boulanger V.
      • Bertrand-Michel J.
      • Terce F.
      • et al.
      HNF1alpha inactivation promotes lipogenesis in human hepatocellular adenoma independently of SREBP-1 and carbohydrate-response element-binding protein (ChREBP) activation.
      Hence, it is not surprising that intratumoural steatosis has been observed in HNF1α mutation-induced HCA.
      • Bioulac-Sage P.
      • Laumonier H.
      • Couchy G.
      • Le Bail B.
      • Sa Cunha A.
      • Rullier A.
      • et al.
      Hepatocellular adenoma management and phenotypic classification: The Bordeaux experience.
      • van Aalten S.M.
      • Thomeer M.G.J.
      • Terkivatan T.
      • Dwarkasing R.S.
      • Verheij J.
      • de Man R.A.
      • et al.
      Hepatocellular adenomas: correlation of MR imaging findings with pathologic subtype classification.

      HNF1β mutations in mice and men

      Individuals with a heterozygous HNF1β inactivating mutation develop MODY5, a relatively severe condition. It is characterised by disorders in multiple organs, from lower birth weight (by ∼800 g) resulting from reduced insulin, to partial pancreatic agenesis, exocrine dysfunction, renal cystic disease, occasional genital tract abnormalities and abnormal liver function.
      • Beards F.
      • Frayling T.
      • Bulman M.
      • Horikawa Y.
      • Allen L.
      • Appleton M.
      • et al.
      Mutations in hepatocyte nuclear factor 1beta are not a common cause of maturity-onset diabetes of the young in the U.K..
      • Bingham C.
      • Hattersley A.T.
      Renal cysts and diabetes syndrome resulting from mutations in hepatocyte nuclear factor-1β.
      • Edghill E.L.
      • Bingham C.
      • Ellard S.
      • Hattersley A.T.
      Mutations in hepatocyte nuclear factor-1β and their related phenotypes.
      • Gonc E.N.
      • Ozturk B.B.
      • Haldorsen I.S.
      • Molnes J.
      • Immervoll H.
      • Raeder H.
      • et al.
      HNF1B mutation in a Turkish child with renal and exocrine pancreas insufficiency, diabetes and liver disease.
      • Haldorsen I.S.
      • Vesterhus M.
      • Ræder H.
      • Jensen D.K.
      • Søvik O.
      • Molven A.
      • et al.
      Lack of pancreatic body and tail in HNF1B mutation carriers.
      It is clear that HNF1β plays a critical role in pancreas organogenesis although gene dosage differs between humans and mice.
      • Haumaitre C.
      • Fabre M.
      • Cormier S.
      • Baumann C.
      • Delezoide A.-L.
      • Cereghini S.
      Severe pancreas hypoplasia and multicystic renal dysplasia in two human fetuses carrying novel HNF1β/MODY5 mutations.
      Hnf1β-null embryos die at the blastocyst stage (E3.5) because of abnormal or absent extraembryonic endoderm.
      • Barbacci E.
      • Reber M.
      • Ott M.O.
      • Breillat C.
      • Huetz F.
      • Cereghini S.
      Variant hepatocyte nuclear factor 1 is required for visceral endoderm specification.
      Hnf1β-null embryos rescued via tetraploid complementation do not have a detectable ventral pancreas, form only a small dorsal pancreas, fail to express Ngn3 and do not successfully form endocrine cells.
      • Haumaitre C.
      • Barbacci E.
      • Jenny M.
      • Ott M.O.
      • Gradwohl G.
      • Cereghini S.
      Lack of TCF2/vHNF1 in mice leads to pancreas agenesis.
      In contrast to humans, mice that are heterozygous for the Hnf1β mutation exhibit no apparent phenotype.
      Patients with MODY5 typically develop kidney abnormalities, such as bilateral renal cysts, renal dysplasia and/or familial hypoplasia glomerulocystic kidney disease.
      • Bingham C.
      • Ellard S.
      • Allen L.
      • Bulman M.
      • Shepherd M.
      • Frayling T.
      • et al.
      Abnormal nephron development associated with a frameshift mutation in the transcription factor hepatocyte nuclear factor-1 beta.
      • Edghill E.L.
      • Oram R.A.
      • Owens M.
      • Stals K.L.
      • Harries L.W.
      • Hattersley A.T.
      • et al.
      Hepatocyte nuclear factor-1beta gene deletions–a common cause of renal disease.
      This can ultimately lead to end-stage renal failure. Conditional knockout of Hnf1β in young mice (at postnatal P1-P10 stages) also results in the development of polycystic kidneys, whereas knockout of Hnf1β at P10 or later results in significantly delayed cyst formation.
      • Gresh L.
      • Fischer E.
      • Reimann A.
      • Tanguy M.
      • Garbay S.
      • Shao X.
      • et al.
      A transcriptional network in polycystic kidney disease.
      • Verdeguer F.
      • Le Corre S.
      • Fischer E.
      • Callens C.
      • Garbay S.
      • Doyen A.
      • et al.
      A mitotic transcriptional switch in polycystic kidney disease.
      Rare cases of genital tract abnormalities, such as rudimentary uterus, vaginal aplasia, bicornuate uterus and double vagina have also been observed in patients with MODY5.
      • Bingham C.
      • Ellard S.
      • Cole T.R.
      • Jones K.E.
      • Allen L.I.
      • Goodship J.A.
      • et al.
      Solitary functioning kidney and diverse genital tract malformations associated with hepatocyte nuclear factor-1beta mutations.
      • Iwasaki N.
      • Babazono T.
      • Tomonaga O.
      • Ogata M.
      • Yokokawa H.
      • Iwamoto Y.
      Mutations in the hepatocyte nuclear factor-1β (MODY5) gene are not a major factor contributing to end-stage renal disease in Japanese people with diabetes mellitus.
      • Lindner T.H.
      • Njolstad P.R.
      • Horikawa Y.
      • Bostad L.
      • Bell G.I.
      • Sovik O.
      A novel syndrome of diabetes mellitus, renal dysfunction and genital malformation associated with a partial deletion of the pseudo-POU domain of hepatocyte nuclear factor-1beta.
      Hnf1β-null mice also show defects in Wolffian duct tube development and as a result, a lack of Müllerian duct formation.
      • Lokmane L.
      • Heliot C.
      • Garcia-Villalba P.
      • Fabre M.
      • Cereghini S.
      VHNF1 functions in distinct regulatory circuits to control ureteric bud branching and early nephrogenesis.
      Patients with MODY5 also present with liver deficiencies, such as neonatal cholestasis, adult-onset cholestasis,
      • Roelandt P.
      • Antoniou A.
      • Libbrecht L.
      • Van Steenbergen W.
      • Laleman W.
      • Verslype C.
      • et al.
      HNF1B deficiency causes ciliary defects in human cholangiocytes.
      hypercholesterolaemia, neonatal jaundice, lack of intrahepatic bile ducts and liver dysfunction.
      • Beckers D.
      • Bellanne-Chantelot C.
      • Maes M.
      Neonatal cholestatic jaundice as the first symptom of a mutation in the hepatocyte nuclear factor-1beta gene (HNF-1beta).
      • Raile K.
      • Klopocki E.
      • Holder M.
      • Wessel T.
      • Galler A.
      • Deiss D.
      • et al.
      Expanded clinical spectrum in hepatocyte nuclear factor 1b-maturity-onset diabetes of the young.
      In mice, targeted inactivation of Hnf1β in hepatocytes and bile duct cells results in the development of severe neonatal cholestasis and jaundice because of abnormal development of the gall bladder and intrahepatic bile ducts, affecting bile acid transport and metabolism.
      • Coffinier C.
      • Gresh L.
      • Fiette L.
      • Tronche F.
      • Schütz G.
      • Babinet C.
      • et al.
      Bile system morphogenesis defects and liver dysfunction upon targeted deletion of HNF1β.

      FOXA mutations in mice and men

      The few studies that reported on FOXA mutations in humans were typically linked to cancer, but not the development or function of the liver, pancreas or kidney. FOXA1 mutations have been linked to human cancers, such as acute myeloid leukaemia, oesophageal, lung, thyroid, pancreatic, breast and prostate cancer.
      • Lin L.
      • Miller C.T.
      • Contreras J.I.
      • Prescott M.S.
      • Dagenais S.L.
      • Wu R.
      • et al.
      The hepatocyte nuclear factor 3 α gene, HNF3α, on chromosome band 14q13 is amplified and overexpressed in esophageal and lung adenocarcinomas.
      • Neben K.
      • Schnittger S.
      • Brors B.
      • Tews B.
      • Kokocinski F.
      • Haferlach T.
      • et al.
      Distinct gene expression patterns associated with FLT3- and NRAS-activating mutations in acute myeloid leukemia with normal karyotype.
      • Nucera C.
      • Eeckhoute J.
      • Finn S.
      • Carroll J.S.
      • Ligon A.H.
      • Priolo C.
      • et al.
      FOXA1 is a potential oncogene in anaplastic thyroid carcinoma.
      • Song L.
      • Xu Z.
      • Zhang C.
      • Qiao X.
      • Huang C.
      Up-regulation of the HSP72 by Foxa1 in MCF-7 human breast cancer cell line.
      • Robinson J.L.L.
      • Holmes K.A.
      • Carroll J.S.
      FOXA1 mutations in hormone-dependent cancers.
      Single nucleotide polymorphisms in FOXA1 and FOXA2 are associated with hepatocellular carcinoma (HCC) in a sexual dimorphic manner, with a significantly higher incidence in males. This phenomenon is replicated in Foxa1- and Foxa2-deficient mice, which suggested that ERα (oestrogen receptor)-dependent gene regulation in female mice prevents cancer formation, while AR (androgen receptor)-mediated regulation promotes it. This is in part linked to the regulation of downstream targets of ER and AR by FOXA1 and FOXA2.
      • Li Z.
      • Tuteja G.
      • Schug J.
      • Kaestner K.H.
      Foxa1 and Foxa2 are essential for sexual dimorphism in liver cancer.
      In lung cancer, inactivation of the anti-proliferative protein C/EBPα has resulted in the inhibition of FOXA2 expression. Restoration of FOXA2 expression causes proliferative arrest and apoptosis of the tumour cells.
      • Halmos B.
      • Huettner C.S.
      • Kocher O.
      • Ferenczi K.
      • Karp D.D.
      • Tenen D.G.
      Down-regulation and antiproliferative role of C/EBPα in lung cancer.
      These expression patterns are suggestive of FOXA1 and FOXA2 acting as pro- or anti-tumourigenic genes. We are not aware of any studies linking mutations in FOXA3 to human disease.
      In contrast to the naturally-occurring FOXA mutations linked to cancer in humans, the phenotypes in mice are quite different. Foxa1-null embryos are phenotypically indistinguishable from wild-type embryos while Foxa2-null embryos are embryonic lethal.
      • Ang S.L.
      • Rossant J.
      HNF-3 beta is essential for node and notochord formation in mouse development.
      • Weinstein J.
      • Jacobsen F.W.
      • Hsu-Chen J.
      • Wu T.
      • Baum L.G.
      A novel mammalian protein, p55CDC, present in dividing cells is associated with protein kinase activity and has homology to the Saccharomyces cerevisiae cell division cycle proteins Cdc20 and Cdc4.
      Foxa1-null mice exhibit electrolyte imbalance, become dehydrated, and develop nephrogenic diabetes insipidus shortly after birth.
      • Behr R.
      • Brestelli J.
      • Fulmer J.T.
      • Miyawaki N.
      • Kleyman T.R.
      • Kaestner K.H.
      Mild nephrogenic diabetes insipidus caused by foxa1 deficiency.
      Foxa1- and Foxa2-null embryos fail to form the liver bud in the foregut endoderm.
      • Lee C.S.
      • Friedman J.R.
      • Fulmer J.T.
      • Kaestner K.H.
      The initiation of liver development is dependent on Foxa transcription factors.
      However, the ablation of Foxa1 or Foxa2 after liver specification does not affect hepatocyte development.
      • Bioulac-Sage P.
      • Laumonier H.
      • Couchy G.
      • Le Bail B.
      • Sa Cunha A.
      • Rullier A.
      • et al.
      Hepatocellular adenoma management and phenotypic classification: The Bordeaux experience.
      While Foxa1 and Foxa2 play critical roles in liver specification and metabolism, Foxa3 is important for hepatic glucose homeostasis as Foxa3-null mice exhibit reduced expression of the glucose transporter Glut2, resulting in hypoglycaemia during prolonged fasting.
      • Friedman J.R.
      • Kaestner K.H.
      The Foxa family of transcription factors in development and metabolism.
      • Bioulac-Sage P.
      • Laumonier H.
      • Couchy G.
      • Le Bail B.
      • Sa Cunha A.
      • Rullier A.
      • et al.
      Hepatocellular adenoma management and phenotypic classification: The Bordeaux experience.
      • Lee C.S.
      • Friedman J.R.
      • Fulmer J.T.
      • Kaestner K.H.
      The initiation of liver development is dependent on Foxa transcription factors.
      • Shen W.
      • Scearce L.M.
      • Brestelli J.E.
      • Sund N.J.
      • Kaestner K.H.
      Foxa3 (Hepatocyte Nuclear Factor 3γ) is required for the regulation of hepatic GLUT2 expression and the maintenance of glucose homeostasis during a prolonged fast.
      In the context of the pancreas, Foxa1-null mice have a 70% reduction in proglucagon expression, making them severely hypoglycaemic and resulting in death between postnatal day 2–12.
      • Friedman J.R.
      • Kaestner K.H.
      The Foxa family of transcription factors in development and metabolism.
      • Kaestner K.H.
      • Katz J.
      • Liu Y.
      • Drucker D.J.
      • Schütz G.
      Inactivation of the winged helix transcription factor HNF3α affects glucose homeostasis and islet glucagon gene expression in vivo.
      Targeted deletion of Foxa2 in pancreatic β cells results in hypoglycaemia and relative hyperinsulinemia. These β cells not only fail to secrete insulin upon glucose challenge but secrete insulin inappropriately in response to amino acids.
      • Lantz K.A.
      • Vatamaniuk M.Z.
      • Brestelli J.E.
      • Friedman J.R.
      • Matschinsky F.M.
      • Kaestner K.H.
      Foxa2 regulates multiple pathways of insulin secretion.
      It remains to be determined whether perturbations in FOXA genes in humans will affect foregut, liver and pancreas development or function.

      HNF4α mutations in mice and men

      Inactivating mutations in HNF4α result in MODY1, which gives rise to insulin secretory defects similar to those in MODY3.
      • Fajans S.S.
      • Bell G.I.
      • Polonsky K.S.
      Molecular mechanisms and clinical pathophysiology of maturity-onset diabetes of the young.
      • Byrne M.M.
      • Sturis J.
      • Menzel S.
      • Yamagata K.
      • Fajans S.S.
      • Dronsfield M.J.
      • et al.
      Altered insulin secretory responses to glucose in diabetic and nondiabetic subjects with mutations in the diabetes susceptibility gene MODY3 on chromosome 12.
      • Yamagata K.
      • Furuta H.
      • Oda N.
      • Kaisaki P.J.
      • Menzel S.
      • Cox N.J.
      • et al.
      Mutations in the hepatocyte nuclear factor-4α gene in maturity-onset diabetes of the young (MODY1).
      Patients with MODY1 also exhibit liver disorders, such as reduced HDL cholesterol, apolipoprotein A1 and A2, and triglyceride levels, as well as increased LDL cholesterol levels.
      • Pearson E.R.
      • Pruhova S.
      • Tack C.J.
      • Johansen A.
      • Castleden H.A.J.
      • Lumb P.J.
      • et al.
      Molecular genetics and phenotypic characteristics of MODY caused by hepatocyte nuclear factor 4α mutations in a large European collection.
      In studies of liver-specific Hnf4α-null mice, Hnf4α was found to be dispensable for the early development of the liver, but important for regulating many genes involved in hepatic function, such as the apolipoproteins and metabolic proteins (APOA1, APOB, PAH, FABP1).
      • Li J.
      • Ning G.
      • Duncan S.A.
      Mammalian hepatocyte differentiation requires the transcription factor HNF-4α.
      The loss of Hnf4α also affects the development of the hepatic epithelium,
      • Parviz F.
      • Matullo C.
      • Garrison W.D.
      • Savatski L.
      • Adamson J.W.
      • Ning G.
      • et al.
      Hepatocyte nuclear factor 4alpha controls the development of a hepatic epithelium and liver morphogenesis.
      leading to steatosis, severe disruption of gluconeogenesis and death at six to eight weeks of age.
      • Chen W.S.
      • Manova K.
      • Weinstein D.C.
      • Duncan S.A.
      • Plump A.S.
      • Prezioso V.R.
      • et al.
      Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
      • Hayhurst G.P.
      • Lee Y.-H.
      • Lambert G.
      • Ward J.M.
      • Gonzalez F.J.
      Hepatocyte nuclear factor 4α (Nuclear Receptor 2A1) is essential for maintenance of hepatic gene expression and lipid homeostasis.
      Since Hnf4α plays a critical role in rodent liver homeostasis, it is not surprising that mutations in the human HNF4α gene can also contribute to HCC. In many cases of HCC, HNF4α gene expression has been shown to be downregulated.
      • Lazarevich N.L.
      • Cheremnova O.A.
      • Varga E.V.
      • Ovchinnikov D.A.
      • Kudrjavtseva E.I.
      • Morozova O.V.
      • et al.
      Progression of HCC in mice is associated with a downregulation in the expression of hepatocyte nuclear factors.
      • Ning Y.
      • Riggins R.B.
      • Mulla J.E.
      • Chung H.
      • Zwart A.
      • Clarke R.
      Interferon gamma restores breast cancer sensitivity to fulvestrant by regulating STAT1, IRF1, NFκB, BCL2 family members, and signaling to caspase-dependent apoptosis.
      Concordantly, overexpression of Hnf4α in mouse HCC models confers protection against carcinogenesis and metastasis. Therefore, Hnf4α possibly acts as a tumour suppressor gene.
      • Ning Y.
      • Riggins R.B.
      • Mulla J.E.
      • Chung H.
      • Zwart A.
      • Clarke R.
      Interferon gamma restores breast cancer sensitivity to fulvestrant by regulating STAT1, IRF1, NFκB, BCL2 family members, and signaling to caspase-dependent apoptosis.
      • Yin X.-M.
      • Ding W.-X.
      • Gao W.
      Autophagy in the liver.
      While β cell-specific Hnf4α-null mice appear to be indistinguishable from their healthy littermates and do not develop diabetes, they exhibit altered cholesterol and triglyceride profiles,
      • Hayhurst G.P.
      • Lee Y.-H.
      • Lambert G.
      • Ward J.M.
      • Gonzalez F.J.
      Hepatocyte nuclear factor 4α (Nuclear Receptor 2A1) is essential for maintenance of hepatic gene expression and lipid homeostasis.
      dysregulation of insulin secretion,
      • Gupta R.K.
      • Vatamaniuk M.Z.
      • Lee C.S.
      • Flaschen R.C.
      • Fulmer J.T.
      • Matschinsky F.M.
      • et al.
      The MODY1 gene HNF-4alpha regulates selected genes involved in insulin secretion.
      and are hyperglycaemic during an intraperitoneal glucose tolerance test with no change in insulin sensitivity.
      • Miura A.
      • Yamagata K.
      • Kakei M.
      • Hatakeyama H.
      • Takahashi N.
      • Fukui K.
      • et al.
      Hepatocyte nuclear factor-4alpha is essential for glucose-stimulated insulin secretion by pancreatic beta-cells.
      In contrast, patients with MODY1 and heterozygous HNF4α mutations typically exhibit an increase in birth weight (macrosomia), transient hypoglycaemia and/or diazoxide-responsive hyperinsulinemia at birth.
      • Pearson E.R.
      • Boj S.F.
      • Steele A.M.
      • Barrett T.
      • Stals K.
      • Shield J.P.
      • et al.
      Macrosomia and hyperinsulinaemic hypoglycaemia in patients with heterozygous mutations in the HNF4A Gene.
      They subsequently display progressive hyperglycaemia associated with impaired insulin secretion that worsens with time.
      • Frayling T.M.
      • Evans J.C.
      • Bulman M.P.
      • Pearson E.
      • Allen L.
      • Owen K.
      • et al.
      Beta-cell genes and diabetes: molecular and clinical characterization of mutations in transcription factors.
      Since HNF1α regulates the expression of the HNF4α gene, this could explain the similarity in clinical phenotype between MODY1 and MODY3.
      • Yamagata K.
      • Furuta H.
      • Oda N.
      • Kaisaki P.J.
      • Menzel S.
      • Cox N.J.
      • et al.
      Mutations in the hepatocyte nuclear factor-4α gene in maturity-onset diabetes of the young (MODY1).
      In contrast to HNF4α mutations, HNF4γ gene mutations or polymorphisms have yet to be linked to any human diseases.

      Genetic variants in HNF1α, HNF1β and HNF4α are associated with T2D risk

      Genetic variants in HNF1α, HNF1β and HNF4α that are likely to be less penetrant have also been associated with the risk of polygenic forms of diabetes (T2D) in genome-wide association studies (GWAS). Variants in HNF1α that have been associated with T2D risk include common missense variants I27L and A98V and low-frequency variant E508K, that may result in the impairment of insulin secretion because of changes in HNF1α transcriptional activity.
      • Holmkvist J.
      • Cervin C.
      • Lyssenko V.
      • Winckler W.
      • Anevski D.
      • Cilio C.
      • et al.
      Common variants in HNF-1 alpha and risk of type 2 diabetes.
      • Marcil V.
      • Sinnett D.
      • Seidman E.
      • Boudreau F.
      • Gendron F.P.
      • Beaulieu J.F.
      • et al.
      Association between genetic variants in the HNF4A gene and childhood-onset Crohn/'s disease.
      • Voight B.F.
      • Scott L.J.
      • Steinthorsdottir V.
      • Morris A.P.
      • Dina C.
      • Welch R.P.
      • et al.
      Twelve type 2 diabetes susceptibility loci identified through large-scale association analysis.
      Further detailed studies of non-diabetic cohorts have also shown that associated variants in HNF1α are likely to play a role in the regulation of insulinogenic index.
      • Dimas A.S.
      • Lagou V.
      • Barker A.
      • Knowles J.W.
      • Magi R.
      • Hivert M.F.
      • et al.
      Impact of type 2 diabetes susceptibility variants on quantitative glycemic traits reveals mechanistic heterogeneity.
      • Huyghe J.R.
      • Jackson A.U.
      • Fogarty M.P.
      • Buchkovich M.L.
      • Stancakova A.
      • Stringham H.M.
      • et al.
      Exome array analysis identifies new loci and low-frequency variants influencing insulin processing and secretion.
      In HNF1β, a number of non-coding variants have previously been associated with T2D.
      • Gudmundsson J.
      • Sulem P.
      • Steinthorsdottir V.
      • Bergthorsson J.T.
      • Thorleifsson G.
      • Manolescu A.
      • et al.
      Two variants on chromosome 17 confer prostate cancer risk, and the one in TCF2 protects against type 2 diabetes.
      • Winckler W.
      • Burtt N.P.
      • Holmkvist J.
      • Cervin C.
      • de Bakker P.I.
      • Sun M.
      • et al.
      Association of common variation in the HNF1alpha gene region with risk of type 2 diabetes.
      One of these variants in the 3′UTR was located in a miRNA-binding site in HNF1β and was found to impact on the action of two miRNAs (hsa-miR-214-5p/has-miR-550a-5p) to influence T2D susceptibility.
      • Jafar-Mohammadi B.
      • Groves C.J.
      • Gjesing A.P.
      • Herrera B.M.
      • Winckler W.
      • Stringham H.M.
      • et al.
      A role for coding functional variants in HNF4A in type 2 diabetes susceptibility.
      HNF1β variation could impact on T2D risk by affecting insulin sensitivity.
      • Dimas A.S.
      • Lagou V.
      • Barker A.
      • Knowles J.W.
      • Magi R.
      • Hivert M.F.
      • et al.
      Impact of type 2 diabetes susceptibility variants on quantitative glycemic traits reveals mechanistic heterogeneity.
      In HNF4α, a low-frequency missense variant T139I has been reported to be associated with T2D in multiple large-scale studies and is distinct from the common non-coding GWAS signal.
      • Fuchsberger C.
      • Flannick J.
      • Teslovich T.M.
      • Mahajan A.
      • Agarwala V.
      • Gaulton K.J.
      • et al.
      The genetic architecture of type 2 diabetes.
      • Gaulton K.J.
      • Ferreira T.
      • Lee Y.
      • Raimondo A.
      • Magi R.
      • Reschen M.E.
      • et al.
      Genetic fine mapping and genomic annotation defines causal mechanisms at type 2 diabetes susceptibility loci.
      • Scott R.A.
      • Scott L.J.
      • Magi R.
      • Marullo L.
      • Gaulton K.J.
      • Kaakinen M.
      • et al.
      An expanded genome-wide association study of type 2 diabetes in Europeans.
      Many studies reporting an association between HNF1α, HNF1β, HNF4α and T2D in multiple ancestry groups have not been described here because they are out of scope. However, the evidence implies that disruption of the function of these factors not only impairs pancreatic islet development and function (in the case of MODY) but also contributes to the susceptibility to T2D.

      ONECUT mutations in mice and men

      There is little information describing OC1 mutations in humans that cause developmental defects in tissues or organs. The expression level of OC1 appears to be decreased concomitantly with its target genes in pancreatic intraepithelial neoplasms and invasive pancreatic ductal adenocarcinoma.
      • Jiang X.
      • Zhang W.
      • Kayed H.
      • Zheng P.
      • Giese N.A.
      • Friess H.
      • et al.
      Loss of ONECUT1 expression in human pancreatic cancer cells.
      This implicates the involvement of OC1 as a tumour suppressor gene in pancreatic cancer. However, more definitive studies are required.
      Oc1- and Oc2-null mice develop severe defects in the liver and biliary system, leading to liver failure and death before postnatal day 10.
      • Clotman F.
      • Lannoy V.J.
      • Reber M.
      • Cereghini S.
      • Cassiman D.
      • Jacquemin P.
      • et al.
      The onecut transcription factor HNF6 is required for normal development of the biliary tract.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      • Margagliotti S.
      • Clotman F.
      • Pierreux C.E.
      • Beaudry J.-B.
      • Jacquemin P.
      • Rousseau G.G.
      • et al.
      The Onecut transcription factors HNF-6/OC-1 and OC-2 regulate early liver expansion by controlling hepatoblast migration.
      The migration of hepatoblasts in the septum transversum and differentiation of hepatoblasts into hepatocytes and biliary lineages is affected, leading to hybrid cells that display characteristics of both cell types.
      • Clotman F.
      • Jacquemin P.
      • Plumb-Rudewiez N.
      • Pierreux C.E.
      • Van der Smissen P.
      • Dietz H.C.
      • et al.
      Control of liver cell fate decision by a gradient of TGFβ signaling modulated by Onecut transcription factors.
      • Margagliotti S.
      • Clotman F.
      • Pierreux C.E.
      • Beaudry J.-B.
      • Jacquemin P.
      • Rousseau G.G.
      • et al.
      The Onecut transcription factors HNF-6/OC-1 and OC-2 regulate early liver expansion by controlling hepatoblast migration.
      The loss of Oc1 in hepatoblasts results in cholestatic liver injury characterised by extensive hepatic necrosis and fibrosis.
      • Vanderpool C.
      • Sparks E.E.
      • Huppert K.A.
      • Gannon M.
      • Means A.L.
      • Huppert S.S.
      Genetic interactions between hepatocyte nuclear factor-6 and notch signaling regulate mouse intrahepatic bile duct development in vivo.
      These liver defects result in a high incidence of early postnatal lethality.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      Oc1- and Oc2-null mice that survive have pancreatic developmental defects and exhibit dysfunctional islets, complete absence of a gallbladder and abnormal extrahepatic bile ducts, leading to severe diabetes.
      • Clotman F.
      • Lannoy V.J.
      • Reber M.
      • Cereghini S.
      • Cassiman D.
      • Jacquemin P.
      • et al.
      The onecut transcription factor HNF6 is required for normal development of the biliary tract.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      As Oc1 regulates Pdx1 expression, it is not surprising that pancreatic hypoplasia occurs in Oc1-null mice because of defects in early pancreas specification.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      The Onecut transcription factor HNF-6 (OC-1) is required for timely specification of the pancreas and acts upstream of Pdx-1 in the specification cascade.
      Oc1-null embryos also lack the expression of Ngn3, a key pancreatic endocrine progenitor marker essential for endocrine specification. The lack of Ngn3 expression results in the impairment of pancreatic endocrine cell differentiation.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      Although there are few endocrine cells at birth in Oc1-null murine pancreas, islets eventually develop. However, both islet morphology and duct morphogenesis are perturbed.
      • Jacquemin P.
      • Durviaux S.M.
      • Jensen J.
      • Godfraind C.
      • Gradwohl G.
      • Guillemot F.
      • et al.
      Transcription factor hepatocyte nuclear factor 6 regulates pancreatic endocrine cell differentiation and controls expression of the proendocrine gene ngn3.
      • Jacquemin P.
      • Lemaigre F.P.
      • Rousseau G.G.
      The Onecut transcription factor HNF-6 (OC-1) is required for timely specification of the pancreas and acts upstream of Pdx-1 in the specification cascade.
      It is currently unclear why OC1 mutations in humans have not been reported to affect human liver or pancreas development.

      Conclusion

      The HNF family members have diverse spatial and temporal expression patterns that are made even more complex by the different isoforms present in multiple tissues and organs, from the earliest stages of embryonic development through to adulthood. Our comparisons between the expression profiles of HNFs in rodents and humans reflect the value of rodent studies in providing biological insight into gene expression and function, given the relative inaccessibility of human tissue. Though rodent models serve as a critical foundation upon which we can start to build our knowledge of HNF biology, the differences in disease phenotypes caused by heterozygous inactivating mutations in humans and rodents reveal the limitations of rodent studies, which do not always translate to the human context. In this respect, human induced pluripotent stem cell models, especially those that are derived from patients, are a promising tool for the analysis of human-specific disease mechanisms during the development of different tissues.
      • DeLaForest A.
      • Nagaoka M.
      • Si-Tayeb K.
      • Noto F.K.
      • Konopka G.
      • Battle M.A.
      • et al.
      HNF4A is essential for specification of hepatic progenitors from human pluripotent stem cells.
      • Teo A.K.
      • Lau H.H.
      • Valdez I.A.
      • Dirice E.
      • Tjora E.
      • Raeder H.
      • et al.
      Early developmental perturbations in a human stem cell model of MODY5/HNF1B pancreatic hypoplasia.
      Mutations in HNFs contribute to disorders that implicate not just the liver but also the pancreas and kidney. While rodent models provide a strong basis for understanding disease development caused by perturbation of HNF gene function, there are key differences in phenotypes arising from heterozygous mutations in humans and rodents. In this respect, human stem cell models could provide a more suitable tool for analysing human-specific disease mechanisms.
      Though many HNF proteins and their isoforms share a high degree of homology, they may play both interdependent as well as distinct roles depending on their expression levels and the tissue(s) involved. The complexity is reflected in the cross-regulatory circuits formed by the HNF proteins that specify tissue identity and function. While studies have now begun to tease out the molecular roles of each of these factors, future work may be aimed at comparing the genome-wide binding targets and downstream effectors of the different HNF isoforms at different developmental stages and in different tissues, to achieve better resolution.
      While HNFs are highly enriched in the liver and play critical regulatory roles in liver development and function, it is evident that mutations in HNFs also give rise to disorders related to the pancreas and kidney (Table 4, Fig. 4). In many instances, the precise molecular mechanisms by which these mutations affect the transcriptional networks in specific tissues or organs remain to be clarified in humans. It is still unclear if the high expression levels of HNFs in the liver result in redundancy and compensation that protect patients from severe liver disorders, despite heterozygous inactivating mutations. Further investigations into the molecular functions of HNFs within the liver and beyond will not only shed light on the variability of clinical phenotypes arising from the different mutations (within and between gene families) but may also uncover novel pathways to tackle human diseases that implicate HNF dysregulation.
      Figure thumbnail gr4
      Fig. 4Overview of key implications in the liver, pancreas and kidney due to dysregulation of HNF proteins. Mutations in HNF1α have been associated with HCA, MODY3 and renal dysplasia. Patients with HNF1β mutation display abnormal liver function, partial pancreatic agenesis-mediated MODY5 and renal cystic disease. FOXA1 and FOXA2 mutations have been reported to be associated with HCC in a sexual dimorphic manner. Patients with HNF4α mutation develop MODY1 and Fanconi syndrome. FOXA, forkhead box A; HCA, hepatocellular adenoma; HCC, hepatocellular carcinoma; HNF, hepatocyte nuclear factor; MODY, maturity onset diabetes of the young; OC, ONECUT.

      Financial support

      L.S.W.L. is supported by the A∗STAR Graduate Academy ( AGA ). A.K.K.T. is supported by the Institute of Molecular and Cell Biology ( IMCB ), A∗STAR, NHG-KTPH SIG/14033, the NUHS-CG Metabolic In-Vitro Core Seed Funding, the JCO Career Development Award (CDA) 15302FG148, A∗STAR and the NMRC OF-YIRG.

      Conflict of interest

      The authors declare no conflicts of interest that pertain to this work.
      Please refer to the accompanying ICMJE disclosure forms for further details.

      Authors’ contributions

      HHL and JBJ prepared the figures and tables. HHL, NHJN, LSWL, JBJ and AKKT wrote the manuscript.

      Acknowledgements

      We thank members of the Teo laboratory for the critical reading of this manuscript.

      Supplementary data

      References

      Author names in bold designate shared co-first authorship

        • Cereghini S.
        Liver-enriched transcription factors and hepatocyte differentiation.
        FASEB J. 1996; 10: 267-282
        • De Simone V.
        • De Magistris L.
        • Lazzaro D.
        • Gerstner J.
        • Monaci P.
        • Nicosia A.
        • et al.
        LFB3, a heterodimer-forming homeoprotein of the LFB1 family, is expressed in specialized epithelia.
        EMBO J. 1991; 10: 1435-1443
        • Rey-Campos J.
        • Chouard T.
        • Yaniv M.
        • Cereghini S.
        VHNF1 is a homeoprotein that activates transcription and forms heterodimers with HNF1.
        EMBO J. 1991; 10: 1445-1457
        • Bach I.
        • Yaniv M.
        More potent transcriptional activators or a transdominant inhibitor of the HNF1 homeoprotein family are generated by alternative RNA processing.
        EMBO J. 1993; 12: 4229-4242
        • Harries L.W.
        • Ellard S.
        • Stride A.
        • Morgan N.G.
        • Hattersley A.T.
        Isomers of the TCF1 gene encoding hepatocyte nuclear factor-1 alpha show differential expression in the pancreas and define the relationship between mutation position and clinical phenotype in monogenic diabetes.
        Hum Mol Genet. 2006; 15: 2216-2224
        • Kaestner K.H.
        • Knöchel W.
        • Martínez D.E.
        Unified nomenclature for the winged helix/forkhead transcription factors.
        Genes Dev. 2000; 14: 142-146
        • Lai E.
        • Prezioso V.R.
        • Tao W.F.
        • Chen W.S.
        • Darnell J.E.
        Hepatocyte nuclear factor 3 alpha belongs to a gene family in mammals that is homologous to the Drosophila homeotic gene fork head.
        Genes Dev. 1991; 5: 416-427
        • Friedman J.R.
        • Kaestner K.H.
        The Foxa family of transcription factors in development and metabolism.
        Cell Mol Life Sci. 2006; 63: 2317-2328
        • Rausa F.
        • Samadani U.
        • Ye H.
        • Lim L.
        • Fletcher C.F.
        • Jenkins N.A.
        • et al.
        The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3β in the developing murine liver and pancreas.
        Dev Biol. 1997; 192: 228-246
        • Pani L.
        • Quian X.B.
        • Clevidence D.
        • Costa R.H.
        The restricted promoter activity of the liver transcription factor hepatocyte nuclear factor 3 beta involves a cell-specific factor and positive autoactivation.
        Mol Cell Biol. 1992; 12: 552-562
        • Qian X.
        • Costa R.H.
        Analysis of hepatocyte nuclear factor-3 beta protein domains required for transcriptional activation and nuclear targeting.
        Nucleic Acids Res. 1995; 23: 1184-1191
        • Hadzopoulou-Cladaras M.
        • Kistanova E.
        • Evagelopoulou C.
        • Zeng S.
        • Cladaras C.
        • Ladias J.A.A.
        Functional domains of the nuclear receptor hepatocyte nuclear factor 4.
        J Biol Chem. 1997; 272: 539-550
        • Sladek F.M.
        • Seidel S.D.
        Hepatocyte nuclear factor 4α.
        in: Burris T.B. McCabe E.R.B. Nuclear Receptors and Genetic Disease. Academic Press, San Diego2001
        • Drewes T.
        • Senkel S.
        • Holewa B.
        • Ryffel G.U.
        Human hepatocyte nuclear factor 4 isoforms are encoded by distinct and differentially expressed genes.
        Mol Cell Biol. 1996; 16
        • Huang J.
        • Levitsky L.L.
        • Rhoads D.B.
        Novel P2 promoter-derived HNF4alpha isoforms with different N-terminus generated by alternate exon insertion.
        Exp Cell Res. 2009; 315: 1200-1211
        • Sladek F.M.
        • Ruse M.D.
        • Nepomuceno L.
        • Huang S.-M.
        • Stallcup M.R.
        Modulation of transcriptional activation and coactivator interaction by a splicing variation in the f domain of nuclear receptor hepatocyte nuclear factor 4α1.
        Mol Cell Biol. 1999; 19: 6509-6522
        • Thomas H.
        • Jaschkowitz K.
        • Bulman M.
        • Frayling T.M.
        • Mitchell S.M.
        • Roosen S.
        • et al.
        A distant upstream promoter of the HNF-4alpha gene connects the transcription factors involved in maturity-onset diabetes of the young.
        Hum Mol Genet. 2001; 10: 2089-2097
        • Iyaguchi D.
        • Yao M.
        • Watanabe N.
        • Nishihira J.
        • Tanaka I.
        DNA recognition mechanism of the ONECUT homeodomain of transcription factor HNF-6.
        Structure. 2007; 15: 75-83
        • Lemaigre F.P.
        • Durviaux S.M.
        • Truong O.
        • Lannoy V.J.
        • Hsuan J.J.
        • Rousseau G.G.
        Hepatocyte nuclear factor 6, a transcription factor that contains a novel type of homeodomain and a single cut domain.
        PNAS. 1996; 93: 9460-9464
        • Lannoy V.J.
        • Rodolosse A.
        • Pierreux C.E.
        • Rousseau G.G.
        • Lemaigre F.P.
        Transcriptional stimulation by hepatocyte nuclear factor-6: target-specific recruitment of either creb-binding protein (CBP) or p300/CBP-associated factor (p/CAF).
        J Biol Chem. 2000; 275: 22098-22103
        • Jacquemin P.
        • Lannoy V.J.
        • Rousseau G.G.
        • Lemaigre F.P.
        OC-2, a novel mammalian member of the ONECUT class of homeodomain transcription factors whose function in liver partially overlaps with that of hepatocyte nuclear factor-6.
        J Biol Chem. 1999; 274: 2665-2671
        • Vanhorenbeeck V.
        • Jacquemin P.
        • Lemaigre F.P.
        • Rousseau G.G.
        OC-3, a Novel Mammalian Member of the ONECUT Class of Transcription Factors.
        Biochem Biophys Res Commun. 2002; 292: 848-854
        • Barbacci E.
        • Reber M.
        • Ott M.O.
        • Breillat C.
        • Huetz F.
        • Cereghini S.
        Variant hepatocyte nuclear factor 1 is required for visceral endoderm specification.
        Development. 1999; 126: 4795-4805
        • Cereghini S.
        • Ott M.O.
        • Power S.
        • Maury M.
        Expression patterns of vHNF1 and HNF1 homeoproteins in early postimplantation embryos suggest distinct and sequential developmental roles.
        Development. 1992; 116: 783-797
        • Chen W.S.
        • Manova K.
        • Weinstein D.C.
        • Duncan S.A.
        • Plump A.S.
        • Prezioso V.R.
        • et al.
        Disruption of the HNF-4 gene, expressed in visceral endoderm, leads to cell death in embryonic ectoderm and impaired gastrulation of mouse embryos.
        Genes Dev. 1994; 8
        • Duncan S.A.
        • Manova K.
        • Chen W.S.
        • Hoodless P.
        • Weinstein D.C.
        • Bachvarova R.F.
        • et al.
        Expression of transcription factor HNF-4 in the extraembryonic endoderm, gut, and nephrogenic tissue of the developing mouse embryo: HNF-4 is a marker for primary endoderm in the implanting blastocyst.
        PNAS. 1994; 91: 7598-7602
        • Li J.
        • Ning G.
        • Duncan S.A.
        Mammalian hepatocyte differentiation requires the transcription factor HNF-4α.
        Genes Dev. 2000; 14: 464-474
        • Parviz F.
        • Matullo C.
        • Garrison W.D.
        • Savatski L.
        • Adamson J.W.
        • Ning G.
        • et al.
        Hepatocyte nuclear factor 4alpha controls the development of a hepatic epithelium and liver morphogenesis.
        Nat Genet. 2003; 34: 292-296
        • Taraviras S.
        • Monaghan A.P.
        • Schutz G.
        • Kelsey G.
        Characterization of the mouse HNF-4 gene and its expression during mouse embryogenesis.
        Mech Dev. 1994; 48
        • Nammo T.
        • Yamagata K.
        • Tanaka T.
        • Kodama T.
        • Sladek F.M.
        • Fukui K.
        • et al.
        Expression of HNF-4alpha (MODY1), HNF-1beta (MODY5), and HNF-1alpha (MODY3) proteins in the developing mouse pancreas.
        Gene Expr Patterns. 2008; 8: 96-106
        • Ott M.-O.
        • Rey-Campos J.
        • Cereghini S.
        • Yaniv M.
        VHNF1 is expressed in epithelial cells of distinct embryonic origin during development and precedes HNF1 expression.
        Mech Dev. 1991; 36: 47-58
        • Monaghan A.P.
        • Kaestner K.H.
        • Grau E.
        • Schutz G.
        Postimplantation expression patterns indicate a role for the mouse forkhead/HNF-3 alpha, beta and gamma genes in determination of the definitive endoderm, chordamesoderm and neuroectoderm.
        Development. 1993; 119: 567-578
        • Landry C.
        • Clotman F.
        • Hioki T.
        • Oda H.
        • Picard J.J.
        • Lemaigre F.P.
        • et al.
        HNF-6 is expressed in endoderm derivatives and nervous system of the mouse embryo and participates to the cross-regulatory network of liver-enriched transcription factors.
        Dev Biol. 1997; 192: 247-257
        • Dean S.
        • Tang J.I.
        • Seckl J.R.
        • Nyirenda M.J.
        Developmental and tissue-specific regulation of hepatocyte nuclear factor 4-alpha (HNF4-alpha) isoforms in rodents.
        Gene Expr. 2010; 14: 337-344
        • De Vas M.G.
        • Kopp J.L.
        • Heliot C.
        • Sander M.
        • Cereghini S.
        • Haumaitre C.
        Hnf1b controls pancreas morphogenesis and the generation of Ngn3+ endocrine progenitors.
        Development. 2015; 142: 871-882
        • Taraviras S.
        • Mantamadiotis T.
        • Dong-Si T.
        • Mincheva A.
        • Lichter P.
        • Drewes T.
        • et al.
        Primary structure, chromosomal mapping, expression and transcriptional activity of murine hepatocyte nuclear factor 4gamma.
        Biochim Biophys Acta. 2000; 1490: 21-32
        • Nammo T.
        • Yamagata K.
        • Hamaoka R.
        • Zhu Q.
        • Akiyama T.
        • Gonzalez F.
        • et al.
        Expression profile of MODY3/HNF-1α protein in the developing mouse pancreas.
        Diabetologia. 2002; 45: 1142-1153
        • Shih D.Q.
        • Stoffel M.
        Dissecting the transcriptional network of pancreatic islets during development and differentiation.
        Proc Natl Acad Sci U S A. 2001; 98: 14189-14191
        • Jacquemin P.
        • Pierreux C.E.
        • Fierens S.
        • van Eyll J.M.
        • Lemaigre F.P.
        • Rousseau G.G.
        Cloning and embryonic expression pattern of the mouse Onecut transcription factor OC-2.
        Gene Expr Patterns. 2003; 3: 639-644