Proteomic analysis revealed association of aberrant ROS signaling with suberoylanilide hydroxamic acid-induced autophagy in Jurkat T-leukemia cells.
Suberoylanilide hydroxamic acid (SAHA) is a newly emerging histone deacetylase inhibitor (HDACi) and has been approved in phase II clinical trials for treating patients with cutaneous T-cell lymphoma. Autophagy is a conserved self-digestion process that degrades cytoplasmic materials and recycles long-lived proteins and organelles within cells. In this study, we demonstrate that SAHA stimulates autophagy in Jurkat T-leukemia cells, which was evidenced by the appearance of autophagic vacuoles, formation of acidic vesicular organelles, recruitment of LC3-II to the autophagosomes and conversion of LC3-I to LC3-II . Moreover, SAHA treatment upregulated expression of Beclin 1 and Atg7 and promoted formation of the Atg12-Atg5 conjugate. Furthermore, inhibition of autophagy by chloroquine (CQ) enhanced SAHA-induced apoptosis. To determine the underlying mechanism of SAHA-induced autophagy, two complementary proteomic approaches (2-DE and SILAC), coupled with ESI-Q-TOF MS/MS analysis are utilized to profile differentially expressed proteins between control and SAHA-treated Jurkat T-leukemia cells. In total, 72 proteins were identified with significant alterations. Cluster analysis of the changed proteins reveal several groups of enzymes associated with energy metabolism, anti-oxidative stress and cellular redox control, which suggested an abnormal reactive oxygen species (ROS) production in SAHA-treated Jurkat T-leukemia cells. These observations were further confirmed by ROS chemiluminescence assay. Mechanistic studies revealed that SAHA-triggered autophagy was mediated by ROS production, which could be attenuated by N-acetyl cysteine (NAC), a ROS inhibitor. Finally, we illustrated that Akt-mTOR signaling, a major suppressive cascade of autophagy, was inactivated by SAHA treatment. Taken together, our study identifies autophagy as a reaction to counter increased ROS and is thus involved as a cellular prosurvival mechanism in response to SAHA treatment.
Li
Jingyi
J
The State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, P.R. China.
Liu
Rui
R
Lei
Yunlong
Y
Wang
Kui
K
Lau
Quek Choon
QC
Xie
Na
N
Zhou
Shengtao
S
Nie
Chunlai
C
Chen
Lijuan
L
Wei
Yuquan
Y
Huang
Canhua
C
D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects* / Q000235:genetics; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology*; D015151:Immunoblotting; D019169:Jurkat Cells; D015458:Leukemia, T-Cell / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008954:Models, Biological; D009363:Neoplasm Proteins / Q000378:metabolism; D010588:Phagosomes / Q000187:drug effects / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D014617:Vacuoles / Q000187:drug effects / Q000378:metabolism; D000077337:Vorinostat
D006877:Hydroxamic Acids; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D017382:Reactive Oxygen Species; D049452:Green Fluorescent Proteins; D000077337:Vorinostat; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases