Abstract 2945: Methyl CpG binding protein MeCP2 interacts with the stress-response transcription coactivator LEDGF/p75 and modulates its survival and transcription functions
Authors: Lai Sum Leoh, Leslimar Rios-Colon, Shannalee Martinez, Sandy Tungteakkhun, Maria Filippova, Llano Manuel, Rik Gijsbers, Zeger Debyser, Carlos A. Casiano
Journal: Cancer Research (2010)

Abstract The lens epithelium derived growth factor p75 (LEDGF/p75) is an emerging oncoprotein identified as a key player in the cellular response to oxidative stress. LEDGF/p75 is induced by oxidative stress and promotes resistance to stress-induced cell death presumably by transcriptionally activating specific stress and survival genes. LEDGF/p75 has a splice variant, p52, which induces cell death when overexpressed. We have shown elevated expression of LEDGF/p75 in advanced stage prostatic adenocarcinomas, most likely as a result of increased oxidative stress in the prostate tumor microenvironment. To understand the mechanisms by which LEDGF/p75 transcriptionally activate stress genes and promotes protection against oxidative stress-induced cell death, it was necessary to identify endogenous cellular interacting partners of this protein. Screening of a transcription factor protein array identified the Methyl CpG binding Protein 2 (MeCP2) as a possible interacting partner of LEDGF/p75. MeCP2 is mutated in patients with Rett syndrome, both activate and repress transcription and has been implicated in prostate tumor growth. This study was designed to validate MeCP2 as an interacting partner of LEDFG/p75 and to explore its influence on LEDGF/p75's function. To examine the interaction of MeCP2 and LEDGF, pull down assays were performed with purified GST-MeCP2 incubated with His-LEDGF/p75 or His-p52, and with GST-MeCP2 or GST-p75 incubated with U2OS cell lysates. The results suggested binding of MeCP2 to both LEDGF/p75 and p52. Co-immunoprecipitation assays using U2OS cell lysates revealed binding of proteins, detected by immunoblotting using specific anti-LEDGF/p75 or MeCP2 antibodies, suggesting interaction of both proteins in the cellular microenvironment. We also examined the intracellular localization of both proteins in U2OS cells by immunofluorescence microscopy. Both MeCP2 and LEDGF/p75 co-localized in the chromatin, showing a similar dense fine speckled staining pattern. To examine if MeCP2 influences LEDGF/p75's transcriptional activity, luciferase reporter assays were performed. Even though LEDGF/p75, p52 and MeCP2 transactivated the Hsp27 promoter (pr) individually, co-expression of LEDGF/p75 with MeCP2 increased Hsp27pr activity, while co-expression of LEDGF/p52 with MeCP2 decreased activity. In addition, LEDGF/p75 overexpression protected PC3 cells from tert-butyl hydrogen peroxide (TBHP) treatment; but siRNA knockdown of MeCP2 in these cells sensitized cells to TBHP treatment compared to control siRNA. In conclusion, our data suggest that LEDGF/p75 and MeCP2 are interacting partners and MeCP2 influences LEDGF/p75's transcription and survival function. Future studies will explore modulating the interaction between LEDGF/p75 and MeCP2 to sensitize prostate cancer cells to therapy-induced cell death. 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 2945.