Inactivation of the human vitamin D receptor by caspase-3.
Calcitriol actions are mediated by the vitamin D receptor (VDR), a nuclear transcription factor of the steroid-retinoid-thyroid nuclear receptor gene superfamily. Calcitriol inhibits the growth of many cells including cancer cells by inducing cell cycle arrest. In some cancer cell lines, calcitriol also induces apoptosis. In the LNCaP prostate cancer cell line, induction of apoptosis and caspase-3/7 activities by staurosporine (STS) abolished [(3)H]1,25-dihydroxy vitamin D(3) binding and VDR protein, suggesting that the VDR may be targeted for inactivation by caspases during apoptosis. A potential caspase-3 site (D(195)MMD(198)S) was identified in the human VDR ligand-binding domain. Mutations D195A, D198A, and S199A were generated in the putative capase-3 cleavage site. In transfected COS-7 cells, STS treatment resulted in the cleavage of the wild-type (WT) VDR and S199A mutant VDR but not the D195A or D198A mutants. In in vitro assays, the WT VDR and S199A mutant VDR were cleaved by caspase-3, although the D195A and D198A mutants were resistant to caspase-3. In vitro, the WT VDR was also cleaved by caspase-6 and caspase-7 and in extracts of STS-treated LNCaP cells. In STS-treated LNCaP cells and human skin fibroblasts, the proteasome inhibitor MG-132 protected the VDR caspase cleavage fragment from further degradation by the 26S proteasome. The rat VDR that does not contain the caspase-3 cleavage site was not cleaved in STS-treated COS-7 cells. In conclusion, our results demonstrate that the human VDR is a target of caspase-3 and suggest that activation of caspase-3 may limit VDR activity.
Malloy
Peter J
PJ
Division of Endocrinology, Gerontology, and Metabolism, Department of Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA. malloy@cmgm.Stanford.edu
Feldman
David
D
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D001665:Binding Sites; D019556:COS Cells; D053148:Caspase 3 / Q000378:metabolism*; D002522:Chlorocebus aethiops; D006801:Humans; D008954:Models, Biological; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000502:physiology; D018167:Receptors, Calcitriol / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014407:Tumor Cells, Cultured
D050505:Mutant Proteins; D018167:Receptors, Calcitriol; D053148:Caspase 3