Orphan nuclear receptor SHP interacts with and represses hepatocyte nuclear factor-6 (HNF-6) transactivation.
SHP (small heterodimer partner; NR0B2) is an atypical orphan NR (nuclear receptor) that functions as a transcriptional co-repressor by interacting with a diverse set of NRs and transcriptional factors. HNF-6 (hepatocyte nuclear factor-6) is a key regulatory factor in pancreatic development, endocrine differentiation and the formation of the biliary tract, as well as glucose metabolism. In this study, we have investigated the function of SHP as a putative repressor of HNF-6. Using transient transfection assays, we have shown that SHP represses the transcriptional activity of HNF-6. Confocal microscopy revealed that both SHP and HNF-6 co-localize in the nuclei of cells. SHP physically interacted with HNF-6 in protein-protein association assays in vitro. EMSAs (electrophoretic mobility-shift assays) and ChIP (chromatin immunoprecipitation) assays demonstrated that SHP inhibits the DNA-binding activity of HNF-6 to an HNF-6-response element consensus sequence, and the HNF-6 target region of the endogenous G6Pase (glucose 6-phosphatase) promoter respectively. Northern blot analysis of HNF-6 target genes in cells infected with adenoviral vectors for SHP and SHP siRNAs (small inhibitory RNAs) indicated that SHP represses the expression of endogenous G6Pase and PEPCK (phosphoenolpyruvate carboxykinase). Our results suggest that HNF-6 is a novel target of SHP in the regulation of gluconeogenesis.
Lee
Yong-Soo
YS
Hormone Research Center, School of Biological Sciences and Technology, Chonnam National University, Gwangju 500-757, Republic of Korea.
Kim
Don-Kyu
DK
Kim
Yong Deuk
YD
Park
Ki Cheol
KC
Shong
Minho
M
Seong
Hyun-A
HA
Ha
Hyun Jung
HJ
Choi
Hueng-Sik
HS
D015153:Blotting, Western; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D005952:Glucose-6-Phosphatase / Q000235:genetics / Q000378:metabolism; D051559:Hepatocyte Nuclear Factor 6 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D018613:Microscopy, Confocal; D010730:Phosphoenolpyruvate Carboxylase / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015533:Transcriptional Activation
D051559:Hepatocyte Nuclear Factor 6; D034741:RNA, Small Interfering; D018160:Receptors, Cytoplasmic and Nuclear; C099713:nuclear receptor subfamily 0, group B, member 2; D005952:Glucose-6-Phosphatase; D010730:Phosphoenolpyruvate Carboxylase