Beta-catenin regulates vitamin C biosynthesis and cell survival in murine liver.
Because the Wnt/beta-catenin pathway plays multiple roles in liver pathobiology, it is critical to identify gene targets that mediate such diverse effects. Here we report a novel role of beta-catenin in controlling ascorbic acid biosynthesis in murine liver through regulation of expression of regucalcin or senescence marker protein 30 and L-gulonolactone oxidase. Reverse transcription-PCR, Western blotting, and immunohistochemistry demonstrate decreased regucalcin expression in beta-catenin-null livers and greater expression in beta-catenin overexpressing transgenic livers, HepG2 hepatoma cells (contain constitutively active beta-catenin), regenerating livers, and in hepatocellular cancer tissues that exhibit beta-catenin activation. Interestingly, coprecipitation and immunofluorescence studies also demonstrate an association of beta-catenin and regucalcin. Luciferase reporter and chromatin immunoprecipitation assays verified a functional TCF-4-binding site located between -163 and -157 (CTTTGCA) on the regucalcin promoter to be critical for regulation by beta-catenin. Significantly lower serum ascorbate levels were observed in beta-catenin knock-out mice secondary to decreased expression of regucalcin and also of L-gulonolactone oxidase, the penultimate and last (also rate-limiting) steps in the synthesis of ascorbic acid, respectively. These mice also show enhanced basal hepatocyte apoptosis. To test if ascorbate deficiency secondary to beta-catenin loss and regucalcin decrease was contributing to apoptosis, beta-catenin-null hepatocytes or regucalcin small interfering RNA-transfected HepG2 cells were cultured, which exhibited significant apoptosis that was alleviated by the addition of ascorbic acid. Thus, through regucalcin and L-gulonolactone oxidase expression, beta-catenin regulates vitamin C biosynthesis in murine liver, which in turn may be one of the mechanisms contributing to the role of beta-catenin in cell survival.
Nejak-Bowen
Kari N
KN
Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15216.
Zeng
Gang
G
Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15216.
Tan
Xinping
X
Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15216.
Cieply
Benjamin
B
Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15216.
Monga
Satdarshan P
SP
Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15216; Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15216. Electronic address: smonga@pitt.edu.
D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D001205:Ascorbic Acid / Q000096:biosynthesis*; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D050544:L-Gulonolactone Oxidase / Q000235:genetics / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D000073940:Transcription Factor 4; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
D000975:Antioxidants; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D002135:Calcium-Binding Proteins; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C513323:RGN protein, human; D034741:RNA, Small Interfering; C513324:Rgn protein, mouse; C532096:TCF4 protein, human; D000073940:Transcription Factor 4; D014157:Transcription Factors; D051176:beta Catenin; D050544:L-Gulonolactone Oxidase; D001205:Ascorbic Acid
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