The Modified Human DNA Repair EnzymeO⁶-Methylguanine-DNA Methyltransferase Is a Negative Regulator of Estrogen Receptor-Mediated Transcription upon Alkylation DNA Damage

Abstract

Cell proliferation requires precise control to prevent mutations from replication of (unrepaired) damaged DNA in cells exposed spontaneously to mutagens. Here we show that the modified human DNA repair enzyme O 6 -methylguanine-DNA methyltransferase (R-MGMT), formed from the suicidal repair of the mutagenic O 6 -alkylguanine (6RG) lesions by MGMT in the cells exposed to alkylating carcinogens, functions in such control by preventing the estrogen receptor (ER) from transcription activation that mediates cell proliferation. This function is in contrast to the phosphotriester repair domain of bacterial ADA protein, which acts merely as a transcription activator for its own synthesis upon repair of phosphotriester lesions. First, MGMT, which is constitutively present at active transcription sites, coprecipitates with the transcription integrator CREBbinding protein CBP/p300 but not R-MGMT. Second, R-MGMT, which adopts an altered conformation, utilizes its exposed VLWKLLKVV peptide domain (codons 98 to 106) to bind ER. This binding blocks ER from association with the LXXLL motif of its coactivator, steroid receptor coactivator-1, and thus represses ER effectively from carrying out transcription that regulates cell growth. Thus, through a change in conformation upon repair of the 6RG lesion, MGMT switches from a DNA repair factor to a transcription regulator (R-MGMT), enabling the cell to sense as well as respond to mutagens. These results have implications in chemotherapy and provide insights into the mechanisms for linking transcription suppression with transcription-coupled DNA repair.Exposure to environmental mutagens, such as UV irradiation and N-nitroso compounds, accounts for 80% of the human cancer incidence (30). The effectiveness of our cells' attempts to repair the DNA lesions inflicted by mutagens on our DNA before DNA replication is fundamentally linked to manifestation of the disease through this etiological pathway. The p53 protein is critical here for maintaining genomic integrity, since its induction upon DNA damage enables the cell to acquire sufficient time to repair the damaged DNA by halting cell cycle progression through its effector, the cell cycle-dependent kinase inhibitor p21 WAFI (11,15). However, p53 appears to be only a downstream effector of this DNA damage response pathway in the cell, since cellular factors, such as the hChk1 and hChk2 (human homologs of the yeast RAD53 and CDS1 proteins), are shown to stabilize p53 through phosphorylation upon exposure to mutagens (6,14,35). While much is known about cell regulation where external stimuli are transduced via the membrane receptors and kinase cascades to activate the nuclear DNA (8), knowledge of reciprocal pathways through which DNA, when it is damaged, signals cellular response through immediate factors remains circumstantial.The high-fidelity property of DNA and RNA polymerases enables them to serve as important signaling factors for the integrity of the DNA as they are arrested at the bulky DNA lesions inflicted by mutagens (33) whil...

Promoter Methylation and Polymorphisms of the<i> MGMT</i> Gene in Glioblastomas: A Population-Based Study

“…Upon the repair of O 6 -methylguanine adducts, MGMT changes its conformation to expose its LXXLL motif and binds the estrogen receptor (ER), blocking ER from binding transcription coactivators containing the same motif [59] . MGMT 143Val may affect the regulation of ER-dependent signaling, as MGMT Ile143Val lies in close proximity to the conserved ER-interacting helix.…”

Hypermethylation of CpG island in O6-methylguanine-DNA methyltransferase gene was associated withK-rasG to A mutation in colorectal tumor

“…MGMT removes alkyl adducts from the O 6 position of guanine, and then inactivates the proliferation of cells with damaged DNA. Therefore, it can protect cells from alkylating damages and maintain genomic integrity [15] . Our results suggest that the epigenetic alteration in MGMT promoter may confer an increased risk of alkylating agentinduced colorectal carcinogenesis.…”

Targeting O 6-methylguanine-DNA methyltransferase with specific inhibitors as a strategy in cancer therapy

“…Thus, in adherent cells, no effect on cell growth is seen whether they express low or very high MGMT levels, indicating that MGMT itself is not linked to regulation of proliferation. In addition, although MGMT is frequently upregulated in cells in more aggressively growing tumours [22], there is no evidence that MGMT has a direct stimulatory or inhibitory effect on tumour growth, although there is one report that the methylated (inactivated) MGMT can bind to the oestrogen receptor and affects growth rate [23]. It has also been shown that, in CD34 ?…”