Abstract 3216: CREG1, an epigenetically regulated gene, cooperates with p16INK4a in cellular senescence

Abstract

Abstract By passing cellular senescence, an irreversible growth arrest of cells that is activated in normal cells upon shortening of telomere and other cellular stress, to become immortal is one of the prerequisites for carcinogenesis. Using spontaneously immortal Li-Fraumeni Syndrome (LFS) fibroblasts, we found that CREG1 (Cellular Repressor of E1A-stimulated Genes1) is one of the genes whose expression fit the criteria of senescence-associated genes, decrease expression during immortalization and increase in senescence. Moreover, we found that epigenetic mechanism regulates CREG1 expression. CREG1 is a secreted glycoprotein that was shown to bind RB-family pocket proteins and inhibits E2F transactivation activity. Therefore, we hypothesize that CREG1 plays a role in cellular senescence involving p16INK4a/Rb pathway. Ectopic expression of CREG1 in the immortal LFS cell lines decreases cell proliferation but does not directly induce senescence. We confirmed this in osteosarcoma and fibrosarcoma cancer cell lines commonly seen in Li-Fraumeni Syndrome. Because CREG1 was shown to interact with RB pocket proteins in vitro, we tested that growth suppression by CREG1 may depend on the phosphorylation status of Rb. We found that p16INK4a is also downregulated in immortal cells and that coexpression of CREG1 and p16INK4a, and inhibitor of CDK and Rb phosphorylation, has a greater effect than CREG1 and p16INK4a alone to reduce cell growth, induce cell cycle arrest and cellular senescence in immortal LFS, osteosarcoma and fibrosarcoma cell lines. Moreover, cooperation of CREG1 and p16INK4a inhibits the expression of cyclin A and cyclin B by inhibiting promoter activity thereby decreasing mRNA and protein levels; these proteins are required for S-phase entry and G2/M transition. Whether CREG1 directly involved in transcriptional repression of cyclin A and B mediated by p16INK4a will be confirmed by ChIP assay. In conclusion, we are the first to find that CREG1 enhances p16INK4a-induced senescence by transcriptional repression of cell cycle mediated genes. Analysis of how CREG1 and its cooperation with p16INK4a regulate cell cycle and cellular senescence may provide better understanding of cellular immortalization, an early step in human tumorigenesis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3216.