title: Induction of H2AX phosphorylation in tumor cells by gossypol acetic acid is mediated by phosphatidylinositol 3-kinase (PI3K) family.
abstract: BACKGROUND
  H2AX is phosphorylated (γH2AX) by members of the phosphatidylinositol 3-kinase (PI3K) family, including Ataxia telangiectasia-mutated (ATM), ATM- and Rad3-related (ATR) and DNA-PK in response to DNA damage. Our study shows that gossypol acetic acid (GAA) alone can induce γH2AX in Human mucoepidermoid carcinoma cell line (MEC-1) in vitro. Thus, we further examined the possible mechanisms of GAA to induce γH2AX in tumor cells.
  
  
  METHODS
  The PI3K inhibitors caffeine and wortmannin were used in an effort to identify the kinase(s) responsible for GAA -induced γH2AX in MEC-1 cells. DNA dependent protein kinase (DNA-PK) - proficient and -deficient cells, human glioma cell lines M059K and M059J, were also used to evaluate the kinases responsible for GAA induced H2AX phosphorylation. γH2AX expression was detected by immunofluorescent microscopy. Flow cytometry assay was used to assay γH2AX and cell cycle.
  
  
  RESULTS
  GAA induced H2AX phosphorylation in a cell cycle-dependent manner and a significant G0/G1 phase arrest in MEC-1 cells was shown. Caffeine and wortmannin significantly inhibited GAA-induced H2AX phosphorylation in MEC-1 cells. GAA induced H2AX phosphorylation in M059K, but not in M059J. Taken together, these data suggested that GAA treatment alone could induce H2AX phosphorylation in a cell cycle dependent manner in MEC-1 and M059K, but not in M059J cells. A significant G0/G1 phase arrest was shown in MEC-1.
  
  
  CONCLUSIONS
  The member of PI3K family, DNA-PK, ATM and ATR are involved in the H2AX phosphorylation of MEC-1 cells.
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Zhong
authors.initials: Z
authors.lastname: Guo
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Jin
authors.initials: J
authors.lastname: Zhao
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Lei
authors.initials: L
authors.lastname: Song
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Jian-Xiu
authors.initials: JX
authors.lastname: Ma
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Chen-Jing
authors.initials: CJ
authors.lastname: Wang
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Shu-Yan
authors.initials: SY
authors.lastname: Pei
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Chao
authors.initials: C
authors.lastname: Jiang
authors.affiliation: Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.
authors.forename: Shang-Biao
authors.initials: SB
authors.lastname: Li
keywords: ATM; ATR; DNA-PK; Gossypol acetic acid; H2AX phosphorylation