The human AP-endonuclease (APE1/Ref-1), an essential multifunctional protein involved in repair

The human AP-endonuclease (APE1/Ref-1), an essential multifunctional protein involved in repair of oxidative DNA damage as well as in transcriptional regulation, is often overexpressed in tumor cells. of p21. Interestingly, APE1 and AP4 showed mutual dependence for p21 repression. Moreover, ectopic p53 in p53-null cells inhibited AP4s association with APE1, their binding to the promoter and p21 repression. These results together establish APE1s role as a co-activator or co-repressor of p21 gene, dependent on p53 status. It is thus likely that APE1 overexpression and inactivation of p53, often observed in tumor cells, promote tumor cell proliferation by constitutively downregulating p21 expression. Introduction The mammalian apurinic/apyrimidinic-endonuclease (APE1/Ref-1), a ubiquitous and multifunctional protein, was initially characterized as a central player in DNA base excision repair (BER) pathway [1], [2], [3]. APE1 repairs apurinic/apyrimidinic (AP) sites and DNA single-strand breaks induced by reactive oxygen species and alkylating agents, and their oxidation products generated in the genome either spontaneously or after excision of oxidized and alkylated bases by DNA glycosylases. APE1 may also function in the nucleotide incision repair pathway by incising DNA 5 to oxidatively damaged bases including 5,6-dihydrothymidine and alpha-2deoxyadenosine [4], [5]. Apart from its key role in DNA BER, mammalian APE1 possesses two unique and apparently distinct transcriptional regulatory functions. It acts as a reductive activator of several transcription factors including c-Jun, p53, NF-B p50, HIF1- etc and was independently named redox effector factor-1 (Ref-1) [6], [7], [8], [9]. APE1 could also act as a redox-independent and in cultured cells at Lys 6/7 by p300 [13] and deacetylated by SIRT1 [14]. We showed that acetylation of APE1 stimulates formation of the nCaRE-B complex in PTH promoter which contains hnRNP-L and HDAC1 [13]. Furthermore, others in collaboration with us also reported the involvement of APE1 in the transcriptional regulation of Bax [15], Egr-1-mediated PTEN [16] and IL-6-inducible STAT3-mediated acute-phase reactant gene expression [17]. APE1 was found to be essential in mouse and also in cultured human and mouse cells [18], [19], [20]. We also showed that both acetyl acceptor Lys 6/7 and the active His 309 residue in APE1 are essential for cell survival [20]. APE1 was found to buy Gastrodin (Gastrodine) be ubiquitinated by MDM2 at specific N-terminal Lys residues [21] and phosphorylated by CDK5 at Thr 233 [22] which enhanced its ubiquitination and also modulates its gene regulatory functions [23]. APE1 is often buy Gastrodin (Gastrodine) overexpressed in tumor tissues and cancer cells of diverse origin including ovarian, cervical, prostate, glioma, head and neck, germ-cell, non-small-cell lung carcinoma etc and its overexpression is associated with tumor cells resistance to various anticancer drugs [8], [24], [25], [26]. Targeted knockdown of APE1 in mammalian cells or its functional impairment enhances apoptosis, inhibits cell proliferation and sensitizes cells to a variety of genotoxic agents eg. MMS, H2O2, bleomycin, TMZ, BCNU, etoposide, cisplatin and doxorubicin etc. [6], [25], [27], [28], [29], [30], [31], [32], [33]. We made a novel observation on APE1s transcriptional regulatory function and associated drug resistance. We showed Rabbit Polyclonal to AurB/C (phospho-Thr236/202) that APE1 and its acetylated form activate multiple drug resistance gene MDR1 by interacting with buy Gastrodin (Gastrodine) p300/YB-1 and RNA pol II and promoting their recruitment to the MDR1 promoter [34]. The tumor suppressor p53 is buy Gastrodin (Gastrodine) a transcriptional activator that plays an essential role in DNA damage response by inducing cell cycle arrest, senescence and/or apoptosis [35], [36]. p53 triggers cell cycle arrest at G1 phase by transactivating cyclin-dependent kinase inhibitor 1A (CDKN1A)/p21 gene which causes cell cycle arrest and suppresses cell proliferation [37], [38], [39]. Earlier studies showed that APE1 stimulates DNA binding activity of p53 by both redox-dependent and independent mechanisms [40] and its transactivation function in p21 regulation [41]. On the other hand, global gene expression profile analyses by Tell group in collaboration with us identified p21 as one of the genes that is upregulated in APE1-knowkdown HeLa cells in which p53 is nonfunctional [42]. Subsequently, Jiang also documented upregulation of p21 expression and impairment of cell cycle progression and proliferation by silencing APE1 in pancreatic cell lines [43]. We addressed this paradox by analyzing the role of APE1 in p21 expression in p53-expressing colon carcinoma HCT116 cells (HCT116WT) and its isogenic p53-null cells (HCT116p53null). We report here that APE1 functions as a constitutive co-repressor for p21 gene by its association with transcription factor AP4. However, APE1s co-repressor function is overridden in the presence of p53 which after binding to APE1 activates the p21 gene..