PKB/Akt激對MNNG誘發之細胞毒性與致突變性的影響

• Main Authors: Wei-Lei Yang, 楊偉磊
• Other Authors: Jia-Ling Yang
• Format: Others
• Language: zh-TW
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/92686705955143154514

碩士 === 國立清華大學 === 生命科學系 === 89 === Alkylating agent N-methyl-N’-nitro-N-nitroso-guanidine (MNNG) forms a structurally diverse population of alkyl-DNA adducts. The O6-methylguanine induced by MNNG is the most significant adduct causing cytotoxicity and mutagenicity. This adduct is repaired by O6-methylguanine-DNA methyltransferase (MGMT) in cells. Previous studies have indicated that alkylating agents can also induce signal transduction pathways, suggesting that signal transduction pathways may be associated with DNA repair systems. We have adopted the MGMT highly expressed Chinese hamster ovary cells (phenotype Mer+, AGT) and the MGMT deficient parental cells (phenotype Mer-, CHOM) to investigate whether MGMT is involved in the regulation of signaling pathways induced by MNNG. We have also investigated the roles of MNNG-induced signals in cytotoxicity and genotoxicity. CHOM cells exhibit higher levels of endogenous phospho-Akt than AGT cells. MNNG significantly induced phospho-Akt in AGT cells, conversely it decreased the levels of phospho-Akt by in CHOM cells. These phenomena are also observed in Mer+ and Mer- human fibroblasts HFW and GM0011, respectively. MNNG did not elicit ERK and p38 MAPK signals nor did it alter the expression of PTEN, a PI3 kinase phosphatase, in both AGT and CHOM cells. Co- administrating Wortmannin or LY294002, PI3-kinase inhibitors, increased cytotoxicity and decreased mutagenicity induced by MNNG in AGT cells. Okadaic acid, a protein phosphatase inhibitor, markedly elevated the levels of phospho-Akt, and decreased cytotoxicity and mutagenicity induced by MNNG in CHOM cells. The results suggest that in Mer+ cells the activity of Akt may be negatively regulated by MGMT, which must conduct DNA repair upon MNNG exposure and thereby leave the Akt negative control complex to allow increase of Akt activity. In Mer- cells, low MGMT enables the endogenous Akt activity maintained at high levels, which may be decreased by other negative control mechanism such as phosphatase elicited upon MNNG exposure. Akt signal protects MNNG-induced cytotoxicity in both Mer+ and Mer- cells. However, in Mer+ cells Akt activity may elicit "error-prone" repair or replication pathways and subsequently increase mutation frequency. Conversely, supplement Akt activity can decrease mutagenesis in Mer- cells. The results suggest that Akt signal transduction pathway is a key regulator in controlling DNA repair, replication and mutagenesis in MNNG-treated cells.