BNIP3 is Essential for Mediating 6-thioguanine-and 5-fluorouracil-induced Autophagy Following DNA Mismatch Repair Processing

DNA mismatch repair (MMR) processes the chemically-induced mispairs following treatment with clinically important nucleoside analogs such as 6-thioguanine (6-TG) and 5-fluorouracil (5-FU). MMR processing of these drugs has been implicated in activation of a prolonged G(2)/M cell cycle arrest for repair and later induction of apoptosis and/or autophagy for irreparable DNA damage. In this study, we investigated the role of BNIP3 in the activation of autophagy and the temporal relationship between a G(2)/M cell cycle arrest and the activation of BNIP3-mediated autophagy following MMR processing of 6-TG and 5-FU. We found that BNIP3 protein levels are up-regulated in a MLH1 (MMR(+))-dependent manner following 6-TG and 5-FU treatment. Subsequent siRNA-mediated BNIP3 knockdown abrogates 6-TG -induced autophagy. We also found that p53 knockdown or inhibition of mTOR activity by rapamycin cotreatment impairs 6-TG and 5-FU-induced up-regulation of BNIP3 protein levels and autophagy. Furthermore, suppression of Chk1 expression and a subsequent reduction in 6-TG-induced G(2)/M cell cycle arrest by Chk1 siRNA promotes the extent of 6-TG-induced autophagy. These findings suggest that BNIP3 mediates 6-TG- and 5-FU induced autophagy in a p53- and mTOR-dependent manner. Additionally, the duration of Chk1-activated G(2)/M cell cycle arrest determines the level of autophagy following MMR processing of these nucleoside analogs.