Structural Basis of Natural Promoter Recognition by a Unique Nuclear Receptor, HNF4α

Abstract

HNF4␣ (hepatocyte nuclear factor 4␣) plays an essential role in the development and function of vertebrate organs, including hepatocytes and pancreatic ␤-cells by regulating expression of multiple genes involved in organ development, nutrient transport, and diverse metabolic pathways. As such, HNF4␣ is a culprit gene product for a monogenic and dominantly inherited form of diabetes, known as maturity onset diabetes of the young (MODY). As a unique member of the nuclear receptor superfamily, HNF4␣ recognizes target genes containing two hexanucleotide direct repeat DNA-response elements separated by one base pair (DR1) by exclusively forming a cooperative homodimer. We describe here the 2.0 Å crystal structure of human HNF4␣ DNA binding domain in complex with a high affinity promoter element of another MODY gene, HNF1␣, which reveals the molecular basis of unique target gene selection/recognition, DNA binding cooperativity, and dysfunction caused by diabetes-causing mutations. The predicted effects of MODY mutations have been tested by a set of biochemical and functional studies, which show that, in contrast to other MODY gene products, the subtle disruption of HNF4␣ molecular function can cause significant effects in afflicted MODY patients.HNF4␣ is a novel member (NR2A1) of the nuclear receptor (NR) 2 family (1) and plays a crucial role in the development and function of vital organs. For example, HNF4␣ is essential for development of the liver (2), colon (3), and pancreas (4), and deletion of the HNF4␣ gene from the mouse genome results in embryonic lethality due to the failure to undergo normal gastrulation (5, 6). Conditional targeted gene disruption of HFN4␣ results in marked metabolic disregulation and increased mortality (7-9). In addition, HNF4␣ is known to activate a wide variety of genes involved in glucose, fatty acid, cholesterol, and amino acid metabolism in the liver, kidney, intestine, and pancreas (10 -12), including counterpart transcription factors, such as HNF1␣ (13), HNF6 (14), and pregnane X receptor (15), that in turn control additional liver and ␤-cell-specific target genes. Further underscoring the importance of HNF4␣ in pancreatic ␤-cells, mutations on HNF4␣ in humans are directly linked to the onset of MODY1 (maturity onset diabetes of the young 1) (Fig. 1A), one of the most common monogenic causes of diabetes, mainly characterized by impairment of glucosestimulated insulin secretion from the ␤-cells (16). A direct link between HNF4␣ and glucose-stimulated insulin secretion by pancreatic ␤-cells has been proven by cellular studies (17,18) and pancreatic ␤-cell-specific HNF4␣ knock-out mice (17). A recent genome-wide expression profiling study revealed that both HNF1␣ and HNF4␣ are the master regulators of the ␤-cells directly affecting their physiology (19,20). HNF4␣ is functionally very closely related to HNF1␣, since they crossregulate each other and form a common network of transcription factors that controls the development and function of hepatocyte and pancreatic islets (21, 22). A...

Proteomics Reveals a Physical and Functional Link between Hepatocyte Nuclear Factor 4α and Transcription Factor IID

“…Thus, residues 42-165, which fall within the DNA-binding domain, contribute to TBP interaction, whereas residues 108 -115 are particularly important. Residues 108 -115 correspond to conserved ␣-helix III, which is immediately C-terminal to the second zinc finger of the DNA-binding domain (61). Notably, the DNA-binding domains are almost identical in HNF4␣ and HNF4␥, which we also identified as a TBP-interacting protein in our MudPIT analyses, whereas the AF-1 regions of HNF4␣ and HNF4␥ are highly divergent.…”

“…Regions C and D include the zinc finger and hinge regions (60), which together make up the HNF4␣ DNA-binding domain (61). Region E contains the ligand-binding, dimerization, and ligand-dependent AF-2 transactivation domains, whereas region F is a C-terminal negative regulatory domain that appears to modulate HNF4␣ interaction with coregulators and/or fatty acid ligands ( Fig.…”

“…Whether this reflects additional interaction(s) between one or more of the TAF subunits with HNF4␣ remains to be determined. (61). H, hinge region; LBD, ligand-binding domain; NRD, negative regulatory domain.…”

Multiple Binding Modes between HNF4α and the LXXLL Motifs of PGC-1α Lead to Full Activation

“…Transient Transfection and Luciferase Reporter Assays-The luciferase constructs were prepared as described previously (54). Briefly, the HNF4␣-binding element (Ϫ64 to Ϫ52) from the human Hnf1␣ promoter (Ϫ298 to the first AGT) was subcloned into the firefly luciferase reporter vector pGL3-Basis (Promega) (pGL3-HNF1␣).…”

Genomic Control Processes in Adult Body Part Formation

“…Sources: For references to linkages that had been established earlier, see caption to predecessor of this GRN (Davidson, 2006, p. 179). More recent sources of data supporting the linkages shown in this GRN model, both positive and negative, are as follows: PTF1 to ptf1a, PTF1 to exocrine genes, PTF1 to RBP-JL (Beres et al, 2006); (ptf1a, RBP-JL) to pdx1 (Miyatsuka et al, 2007;Wiebe et al, 2007); (mafa, mafb) to pdx1 (Vanhoose et al, 2008); (foxa1, foxa2) to pdx1 (Gao et al, 2008); (foxa, nkx2.2, pdx1) to mafa (Raum et al, 2006); glis3 to mafa, glis3 to insulin, glis3 to pdx1, glis3 to ngn3 (Fernandez-Zapico et al, 2009;Kang et al, 2009); (islet1, hnf4a) to insulin, (Eeckhoute et al, 2006;Zhang et al, 2009);(meis, pbx1, pbx2) to pax6 (Zhang et al, 2006;Delporte et al, 2008); nkx6.1 to hnf1α (Donelan et al, 2010); (nkx2.2, ngn3) to neuroD (Anderson et al, 2009); (pdx1, gata4) to gata4 (Rojas et al, 2009);(nkx6.1, pdx1, pax6) to mafa (Raum et al, 2010); foxa to gata4 (Rojas et al, 2010); pax6 to mafb, pax6 to c-maf, pax6 to neuroD1, and (mafb, neuroD1, foxa1, foxa2) to glucagon (Gosmain et al, 2010); islet1 to arx (Liu et al, 2011); dac to cyc21 (Kalousova et al, 2010); hnf1a to mafa ; pdx1 to pax6 (Delporte et al, 2008); ngn3 to ia1 (Mellitzer et al, 2006); hnf4a to hnf1α (Lu et al, 2008); pax6 to glucagon (Grapp et al, 2009); pbx1 to glucagon (Liu et al, 2006); (pax6, cmaf) to glucagon ; nk6.1 to glucagon (Gauthier et al, 2007); brn4 to glucagon, (pdx1, mafa, neuroD) to pander (Burkhardt et al, 2008); alx3 to insulin (Mirasierra and Vallejo, 2006;Mirasierra et al, ...…”