Inhibition of autophagy enhances DNA damage-induced apoptosis by disrupting CHK1-dependent S phase arrest.
DNA damage has been shown to induce autophagy, but the role of autophagy in the DNA damage response and cell fate is not fully understood. BO-1012, a bifunctional alkylating derivative of 3a-aza-cyclopenta[a]indene, is a potent DNA interstrand cross-linking agent with anticancer activity. In this study, BO-1012 was found to reduce DNA synthesis, inhibit S phase progression, and induce phosphorylation of histone H2AX on serine 139 (γH2AX) exclusively in S phase cells. Both CHK1 and CHK2 were phosphorylated in response to BO-1012 treatment, but only depletion of CHK1, but not CHK2, impaired BO-1012-induced S phase arrest and facilitated the entry of γH2AX-positive cells into G2 phase. CHK1 depletion also significantly enhanced BO-1012-induced cell death and apoptosis. These results indicate that BO-1012-induced S phase arrest is a CHK1-dependent pro-survival response. BO-1012 also resulted in marked induction of acidic vesicular organelle (AVO) formation and microtubule-associated protein 1 light chain 3 (LC3) processing and redistribution, features characteristic of autophagy. Depletion of ATG7 or co-treatment of cells with BO-1012 and either 3-methyladenine or bafilomycin A1, two inhibitors of autophagy, not only reduced CHK1 phosphorylation and disrupted S phase arrest, but also increased cleavage of caspase-9 and PARP, and cell death. These results suggest that cells initiate S phase arrest and autophagy as pro-survival responses to BO-1012-induced DNA damage, and that suppression of autophagy enhances BO-1012-induced apoptosis via disruption of CHK1-dependent S phase arrest.
Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC.
Jong-Shian
JS
Liou
Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC.
Yi-Chen
YC
Wu
Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC.
Wen-Yen
WY
Yen
Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC.
Yu-Shuan
YS
Tang
Institute of Biomedical Sciences, Academia Sinica, Taipei 115, Taiwan, ROC.
Rajesh B
RB
Kakadiya
Institute of Biomedical Sciences, Academia Sinica, Taipei 115, Taiwan, ROC.
Tsann-Long
TL
Su
Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 115, Taiwan, ROC. Electronic address: lhyih@gate.sinica.edu.tw.
Ling-Huei
LH
Yih
D018906:Antineoplastic Agents, Alkylating / Q000819:agonists / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001343:Autophagy / Q000187:drug effects*; D000071193:Autophagy-Related Protein 7; D002219:Carbamates / Q000819:agonists / Q000494:pharmacology; D002277:Carcinoma / Q000188:drug therapy* / Q000201:enzymology / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D000071877:Checkpoint Kinase 1; D058546:Class III Phosphatidylinositol 3-Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D020868:Gene Silencing; D006367:HeLa Cells; D006575:Heterocyclic Compounds, 3-Ring / Q000819:agonists / Q000494:pharmacology; D006801:Humans; D007192:Indenes / Q000819:agonists / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D016196:S Phase / Q000187:drug effects; D044764:Ubiquitin-Activating Enzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism
D018906:Antineoplastic Agents, Alkylating; C000591165:BO-1012; D002219:Carbamates; D003432:Cross-Linking Reagents; D004791:Enzyme Inhibitors; D006575:Heterocyclic Compounds, 3-Ring; D007192:Indenes; D009363:Neoplasm Proteins; D011494:Protein Kinases; D058546:Class III Phosphatidylinositol 3-Kinases; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D025262:Vacuolar Proton-Translocating ATPases; C509349:ATG7 protein, human; D000071193:Autophagy-Related Protein 7; D044764:Ubiquitin-Activating Enzymes
Apoptosis; Autophagy; CHK1; DNA alkylating agent; DNA damage; S phase arrest