Mitochondrial dynamics and quality control mechanisms in chlorpyrifos-induced cytotoxicity

Abstract

Mitochondrial dynamics (fission/fusion) and mitophagy are recognized as two critical processes underlying mitochondrial homeostasis. Mitochondrial dysfunction has been implicated in mitochondrial fission and mitophagy. However, little is known about the mechanism by which mitochondrial fission and mitophagy modulates mitochondrial damage in response to chlorpyrifos (CPF)-induced toxicity. CPF treatment activated the mitogen-activated protein kinases (MAPKs) and induced intrinsic apoptosis by activating caspase-9, caspase-3, and cytochrome c release in human neuroblastoma SH-SY5Y cells. We found that cytosolic DRP1 was translocated to the mitochondria in CPF-treated cells and was phosphorylated at Ser616. DRP1 was required for p53 to translocate to the mitochondria under CPF-induced oxidative stress. Treating cells with mitochondrial-division inhibitor-1 (mdivi-1), which blocks DRP1 translocation, abrogated activation of cleaved caspase-3. Specifically, pretreating cells with mdivi-1 inhibited BAX translocation to the mitochondria by blocking p53 signaling. Recent studies have demonstrated that mitochondrial fission is linked to mitophagy in damaged mitochondria. Therefore, we elucidated the clearance mechanism of damaged mitochondria in CPF-induced toxicity. CPF induces mitochondrial damage and subsequent engulfment of mitochondria in double-membrane autophagic vesicles, indicating that clearance of damaged mitochondria is processed by mitophagy. CPF treatment resulted in PINK1 stabilization on the outer mitochondrial membrane and subsequent increases in recruitment of Parkin from the cytosol to the abnormal mitochondria. Furthermore, knockdown of PINK1 strongly inhibited LC3-II expression by blocking Parkin recruitment, indicating that mitophagy induced by CPF is due to stabilization of PINK1 in mitochondria. We observed that PINK1 stabilization was selectively regulated by JNK and ERK1/2 signaling activation but not p38 signaling. Pretreatment with specific inhibitors of JNK and ERK1/2 significantly decreased the stabilization of PINK1 and Parkin recruitment and blocked LC3-II expression in mitochondria of CPF-exposed cells. Specifically, JNK and ERK1/2 inhibition also dramatically blocked the interaction of PINK1 with Parkin. There results indicate that the JNK and ERK1/2 signaling accurately regulate the PINK1 and Parkin-dependent mitophagy in mitochondria of CPF-treated cells. As the result of this study, we propose that inhibition of DRP1 is a possible therapeutic approach for pesticide-induced neurotoxicity and it would be a novel molecular regulatory mechanism of PINK1 stabilization-mediated mitophagy under CPF exposure.