β-Catenin Regulates Vitamin C Biosynthesis and Cell Survival in Murine Liver

• Published in: The Journal of biological chemistry 2009-10, Vol.284 (41), p.28115-28127
• Main Authors: Nejak-Bowen, Kari N., Zeng, Gang, Tan, Xinping, Cieply, Benjamin, Monga, Satdarshan P.
• Format: Article
• Language: English
• Subjects: Animals; Antioxidants - metabolism; Ascorbic Acid - biosynthesis; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; beta Catenin - genetics; beta Catenin - metabolism; Calcium-Binding Proteins - genetics; Calcium-Binding Proteins - metabolism; Carcinoma, Hepatocellular - genetics; Carcinoma, Hepatocellular - metabolism; Cell Line, Tumor; Cell Survival - physiology; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Gene Expression Regulation; Hepatocytes - cytology; Hepatocytes - metabolism; Humans; Intracellular Signaling Peptides and Proteins - genetics; Intracellular Signaling Peptides and Proteins - metabolism; L-Gulonolactone Oxidase - genetics; L-Gulonolactone Oxidase - metabolism; Liver - cytology; Liver - metabolism; Liver Neoplasms - genetics; Liver Neoplasms - metabolism; Metabolism and Bioenergetics; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Promoter Regions, Genetic; RNA, Small Interfering - genetics; RNA, Small Interfering - metabolism; Transcription Factor 4; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M109.047258

Because the Wnt/β-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 β-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 β-catenin-null livers and greater expression in β-catenin overexpressing transgenic livers, HepG2 hepatoma cells (contain constitutively active β-catenin), regenerating livers, and in hepatocellular cancer tissues that exhibit β-catenin activation. Interestingly, coprecipitation and immunofluorescence studies also demonstrate an association of β-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 β-catenin. Significantly lower serum ascorbate levels were observed in β-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 β-catenin loss and regucalcin decrease was contributing to apoptosis, β-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, β-catenin regulates vitamin C biosynthesis in murine liver, which in turn may be one of the mechanisms contributing to the role of β-catenin in cell survival.