Elevated Levels of Hepatocyte Nuclear Factor 3β in Mouse Hepatocytes Influence Expression of Genes Involved in Bile Acid and Glucose Homeostasis

Rausa, Francisco M.; Tan, Yongjun; Zhou, Heping; Yoo, Kyung Whan; Stolz, Donna B.; Watkins, Simon C.; Franks, Roberta; Unterman, Terry G.; Costa, Robert H.

doi:10.1128/mcb.20.21.8264-8282.2000

Cited by 100 publications

(139 citation statements)

References 84 publications

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“…Tissue blotting of the transgenic mice by anti-V5 antibody showed strong expression of the transgenic protein, V5-BRE, in liver, weak expression in the ear skin and none in other tissues (Supplementary Figure 6a). Immunohistochemical staining revealed mosaic hepatocyte expression pattern (Supplementary Figures 6b and c) characteristic of transgenes driven by this TTR promoter fragment (Rausa et al, 2000;Yu et al, 2007). These transgenic mice showed no detectable abnormalities in liver histology and overall phenotype.…”

Section: Significant Overexpression Of Bre In Human Hepatocellular Camentioning

confidence: 93%

BRE is an antiapoptotic protein in vivo and overexpressed in human hepatocellular carcinoma

Chan

Ching

et al. 2007

Oncogene

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BRE binds to the cytoplasmic domains of tumor necrosis factor receptor-1 and Fas, and in cell lines can attenuate death receptor-initiated apoptosis by inhibiting t-BIDinduced activation of the mitochondrial apoptotic pathway. Overexpression of BRE by transfection can also attenuate intrinsic apoptosis and promote growth of the transfected Lewis lung carcinoma line in mice. There is, however, a complete lack of in vivo data about the protein.Here, we report that by using our BRE-specific monoclonal antibody on the immunohistochemistry of 123 specimens of human hepatocellular carcinoma (HCC), significant differences in BRE expression levels between the paired tumoral and non-tumoral regions (Po2.2eÀ16) were found. Marked overexpression of BRE was detected in majority of the tumors, whereas most non-tumoral regions expressed the same low level of the protein as in normal livers. To investigate whether BRE overexpression could promote cell survival in vivo, liver-specific transgenic BRE mice were generated and found to be significantly resistant to Fas-mediated lethal hepatic apoptosis. The transgenic model also revealed post-transcriptional regulation of Bre level in the liver, which was not observed in HCC and non-HCC cell lines. Indeed, all cell lines analysed express high levels of BRE. In conclusion, BRE is antiapoptotic in vivo, and may promote tumorigenesis when overexpressed.

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Section: Significant Overexpression Of Bre In Human Hepatocellular Camentioning

confidence: 93%

BRE is an antiapoptotic protein in vivo and overexpressed in human hepatocellular carcinoma

Chan

Ching

et al. 2007

Oncogene

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“…Eluted proteins were resolved via SDS-PAGE and transferred to nitrocellulose membranes for Western blot analysis with either monoclonal antibody specific to GFP antibody (Clontech, Palo Alto, CA, for GFP-HNF6), HA epitope tag (Invitrogen, Carlsbad, CA, for HA-C/EBP␣), V5 epitope tag (Invitrogen, for V5-HNF6), or C/EBP␤ (sc-150 [C-19]; Santa Cruz Biotechnology, Santa Cruz, CA) and detected as previously described. 19,21,27 For coimmunoprecipitation assays, 750 g of total protein extract from quiescent mouse liver (0h) or regenerating mouse liver was immunoprecipitated with HNF6 27 or rabbit serum (Vector Laboratories, Burlingame, CA) and then subjected to Western analysis with rabbit anti-C/ EBP␣ antibody (sc-61; Santa Cruz Biotechnology), C/EBP␤ antibody (sc-150 [C-19]; Santa Cruz Biotechnology) or rabbit anti-CBP C-terminal antibody (Upstate, Lake Placid, NY). Peroxidase-conjugated ImmunoPure Recombinant Protein A/G (Pierce, Rockford, IL) was used to identify antibody-antigen bands and was detected via enhanced chemiluminescence (ECL-ϩ Amersham Pharmacia Biotech, Piscataway, NJ) followed by autoradiography.…”

Section: Methodsmentioning

confidence: 99%

C/EBPα and HNF6 protein complex formation stimulates HNF6-dependent transcription by CBP coactivator recruitment in HepG2 cells

et al. 2006

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We previously demonstrated that formation of complexes between the DNA-binding domains of hepatocyte nuclear factor 6 (HNF6) and forkhead box a2 (Foxa2) proteins stimulated Foxa2 transcriptional activity. Here, we used HepG2 cell cotransfection assays to demonstrate that HNF6 transcriptional activity was stimulated by CCAAT/enhancer-binding protein ␣ (C/EBP␣), but not by the related C/EBP␤ or C/EBP␦ proteins. Formation of the C/EBP␣-HNF6 protein complex required the HNF6 cut domain and the C/EBP␣ activation domain (AD) 1/AD2 sequences. This C/EBP␣-HNF6 transcriptional synergy required both the N-terminal HNF6 polyhistidine and serine/threonine/proline box sequences, as well as the C/EBP␣ AD1/AD2 sequences, the latter of which are known to recruit the CREB binding protein (CBP) transcriptional coactivator. Consistent with these findings, adenovirus E1A-mediated inhibition of p300/CBP histone acetyltransferase activity abrogated C/EBP␣-HNF6 transcriptional synergy in cotransfection assays. Co-immunoprecipitation assays with liver protein extracts demonstrate an association between the HNF6 and C/EBP␣ transcription factors and the CBP coactivator protein in vivo. Furthermore, chromatin immunoprecipitation assays with hepatoma cells demonstrated that increased levels of both C/EBP␣ and HNF6 proteins were required to stimulate association of these transcription factors and the CBP coactivator protein with the endogenous mouse Foxa2 promoter region. In conclusion, formation of the C/EBP␣-HNF6 protein complex stimulates recruitment of the CBP coactivator protein for expression of Foxa2, a transcription factor critical for regulating expression of hepatic gluconeogenic genes during fasting. (HEPATOLOGY 2006;43:276-286.) T ransfection studies in hepatoma cell lines demonstrated that synergistic transcriptional activation of hepatocyte-specific genes requires simultaneous binding of multiple hepatocyte nuclear factor to their promoter/enhancer regions. 1 However, the mechanisms by which these hepatocyte nuclear factor transcription factors functionally interact to elicit transcriptional synergy have yet to be deciphered. The CCAAT/enhancer Abbreviations: HNF6, hepatocyte nuclear factor 6; Foxa2, forkhead box a2; C/EBP␣, CCAAT/enhancer-binding protein

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“…11 The HNF-6 protein-DNA complexes were resolved from the unbound probe via electrophoresis on a nondenaturing gel and detected with autoradiography. Specificity of the binding was evaluated via coincubation with 100-fold molar excess of unlabeled oligocompetitor, mouse HNF-6 specific-antibody, 18 or preimmune serum.…”

Section: Methodsmentioning

confidence: 99%

“…17 Syntheses for antisense mouse HNF-6, HNF-4␣, C/EBP␣, FoxA2 (also known as HNF-3␤), insulinlike growth factor 1, and cyclophilin riboprobes have been described. 18 The mouse CYP7A1 ribonuclease protection assay (RPA) probe plasmid was obtained via reverse transcription polymerase chain reaction of mouse liver RNA (using primers 5Ј-GCA AGG ATC CTA CTT CTG CGA AGG CAT TTG G-3Ј and 5Ј-GCC GGA ATT CAA ACA TCA CTC GGT AGC AGA A-3Ј) and cloned into the pBluescript SK(ϩ) template, and an antisense RNA probe was synthesized from a BamHI digested template using T7 RNA polymerase. Expression levels were determined after exposure of the gel to phosphoimaging screens overnight.…”

Section: Methodsmentioning

confidence: 99%

In vivo regulation of murine CYP7A1 by HNF-6: A novel mechanism for diminished CYP7A1 expression in biliary obstruction

et al. 2004

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Disruption of the enterohepatic bile acid circulation during biliary tract obstruction leads to profound perturbation of the cholesterol and bile acid metabolic pathways. Several families of nuclear receptor proteins have been shown to modulate this critical process by regulating hepatic cholesterol catabolism and bile acid synthesis through the transcriptional control of cholesterol 7-␣ hydroxylase (CYP7A1). Hepatocyte nuclear factor (HNF) 6 (also known as OC-1) is a member of the ONECUT family of transcription factors that activate numerous hepatic target genes essential to liver function. We have previously shown that hepatic expression of mouse HNF-6 messenger RNA (mRNA) and protein significantly decrease following bile duct ligation. Because CYP7A1 contains potential HNF-6 binding sites in its promoter region, we tested the hypothesis that HNF-6 transcriptionally regulates CYP7A1. Following bile duct ligation, we demonstrated that diminished HNF-6 mRNA levels correlate with a reduction in CYP7A1 mRNA expression. Increasing hepatic levels of HNF-6 either by infection with recombinant adenovirus vector expressing HNF-6 cDNA by growth hormone treatment leads to an induction of CYP7A1 mRNA. To directly evaluate if HNF-6 is a transcriptional activator for CYP7A1, we used deletional and mutational analyses of CYP7A1 promoter sequences and defined sequences ؊206/؊194 to be critical for CYP7A1 transcriptional stimulation by HNF-6 in cotransfection assays. In conclusion, the HNF-6 protein is a component of the complex network of hepatic transcription factors that regulates the expression of hepatic genes essential for bile acid homeostasis and cholesterol/lipid metabolism in normal and pathological conditions.

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Elevated Levels of Hepatocyte Nuclear Factor 3β in Mouse Hepatocytes Influence Expression of Genes Involved in Bile Acid and Glucose Homeostasis

Cited by 100 publications

References 84 publications

BRE is an antiapoptotic protein in vivo and overexpressed in human hepatocellular carcinoma

BRE is an antiapoptotic protein in vivo and overexpressed in human hepatocellular carcinoma

C/EBPα and HNF6 protein complex formation stimulates HNF6-dependent transcription by CBP coactivator recruitment in HepG2 cells

In vivo regulation of murine CYP7A1 by HNF-6: A novel mechanism for diminished CYP7A1 expression in biliary obstruction

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