title: G9a Inhibition Induces Autophagic Cell Death via AMPK/mTOR Pathway in Bladder Transitional Cell Carcinoma.
abstract: G9a has been reported to highly express in bladder transitional cell carcinoma (TCC) and G9a inhibition significantly attenuates cell proliferation, but the underlying mechanism is not fully understood. The present study aimed at examining the potential role of autophagy in the anti-proliferation effect of G9a inhibition on TCC T24 and UMUC-3 cell lines in vitro. We found that both pharmaceutical and genetical G9a inhibition significantly attenuated cell proliferation by MTT assay, Brdu incorporation assay and colony formation assay. G9a inhibition induced autophagy like morphology as determined by transmission electron microscope and LC-3 fluorescence assay. In addition, autophagy flux was induced by G9a inhibition in TCC cells, as determined by p62 turnover assay and LC-3 turnover assay. The autophagy induced positively contributed to the inhibition of cell proliferation because the growth attenuation capacity of G9a inhibition was reversed by autophagy inhibitors 3-MA. Mechanically, AMPK/mTOR pathway was identified to be involved in the regulation of G9a inhibition induced autophagy. Intensively activating mTOR by Rheb overexpression attenuated autophagy and autophagic cell death induced by G9a inhibition. In addition, pre-inhibiting AMPK by Compound C attenuated autophagy together with the anti-proliferation effect induced by G9a inhibition while pre-activating AMPK by AICAR enhanced them. In conclusion, our results indicate that G9a inhibition induces autophagy through activating AMPK/mTOR pathway and the autophagy induced positively contributes to the inhibition of cell proliferation in TCC cells. These findings shed some light on the functional role of G9a in cell metabolism and suggest that G9a might be a therapeutic target in bladder TCC in the future.
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Feng
authors.initials: F
authors.lastname: Li
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Zeng
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Zhenfeng
authors.initials: Z
authors.lastname: Guan
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Zhenkun
authors.initials: Z
authors.lastname: Ma
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Shi
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Chong
authors.initials: C
authors.lastname: Du
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Jia
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Shan
authors.initials: S
authors.lastname: Xu
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Xinyang
authors.initials: X
authors.lastname: Wang
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Luke
authors.initials: L
authors.lastname: Chang
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Dalin
authors.initials: D
authors.lastname: He
authors.affiliation: Department of Urology, the First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Guo
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects; D002295:Carcinoma, Transitional Cell / Q000201:enzymology* / Q000473:pathology; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006649:Histocompatibility Antigens / Q000378:metabolism; D011495:Histone-Lysine N-Methyltransferase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D015398:Signal Transduction / Q000187:drug effects; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D001749:Urinary Bladder Neoplasms / Q000201:enzymology* / Q000473:pathology; D014617:Vacuoles / Q000187:drug effects / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D006649:Histocompatibility Antigens; C445405:EHMT2 protein, human; D011495:Histone-Lysine N-Methyltransferase; D058570:TOR Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases