A Prospero-related homeodomain protein is a novel co-regulator of hepatocyte nuclear factor 4alpha that regulates the cholesterol 7alpha-hydroxylase gene.
Prox1, an early specific marker for developing liver and pancreas in foregut endoderm has recently been shown to interact with alpha-fetoprotein transcription factor and repress cholesterol 7alpha-hydroxylase (CYP7A1) gene transcription. Using a yeast two-hybrid assay, we found that Prox1 strongly and specifically interacted with hepatocyte nuclear factor (HNF)4alpha, an important transactivator of the human CYP7A1 gene in bile acid synthesis and phosphoenolpyruvate carboxykinase (PEPCK) gene in gluconeogenesis. A real time PCR assay detected Prox1 mRNA expression in human primary hepatocytes and HepG2 cells. Reporter assay, GST pull-down, co-immunoprecipitation, and yeast two-hybrid assays identified a specific interaction between the N-terminal LXXLL motif of Prox1 and the activation function 2 domain of HNF4alpha. Prox1 strongly inhibited HNF4alpha and peroxisome proliferators-activated receptor gamma coactivator-1alpha co-activation of the CYP7A1 and PEPCK genes. Knock down of the endogenous Prox1 by small interfering RNA resulted in significant increase of CYP7A1 and PEPCK mRNA expression and the rate of bile acid synthesis in HepG2 cells. These results suggest that Prox1 is a novel co-regulator of HNF4alpha that may play a key role in the regulation of bile acid synthesis and gluconeogenesis in the liver.
Song
Kwang-Hoon
KH
Department of Microbiology, Immunology and Biochemistry, Northeastern Ohio University College of Medicine, 4209 State Route 44, Rootstown, OH 44272, USA.
Li
Tiangang
T
Chiang
John Y L
JY
D000368:Aged; D020816:Amino Acid Motifs; D001647:Bile Acids and Salts / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured / Q000378:metabolism; D002790:Cholesterol 7-alpha-Hydroxylase / Q000737:chemistry* / Q000235:genetics*; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D005943:Gluconeogenesis; D005982:Glutathione Transferase / Q000378:metabolism; D051557:Hepatocyte Nuclear Factor 4 / Q000378:metabolism / Q000502:physiology*; D022781:Hepatocytes / Q000378:metabolism; D018398:Homeodomain Proteins / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D047468:Immunoprecipitation; D008099:Liver / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D019764:Phosphoenolpyruvate Carboxykinase (ATP) / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D020218:Response Elements; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D025521:Tumor Suppressor Proteins; D020798:Two-Hybrid System Techniques; D000098551:Prospero-Related Homeobox 1 Protein
D001647:Bile Acids and Salts; C496592:HNF4A protein, human; D051557:Hepatocyte Nuclear Factor 4; D018398:Homeodomain Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D025521:Tumor Suppressor Proteins; D000098551:Prospero-Related Homeobox 1 Protein; D008156:Luciferases; D002790:Cholesterol 7-alpha-Hydroxylase; D005982:Glutathione Transferase; D019764:Phosphoenolpyruvate Carboxykinase (ATP)