Oxidative stress induces H2AX phosphorylation in human spermatozoa.
H2AX phosphorylation occurs following the induction of DNA double strand breaks (DSBs), thus collaborating with many other proteins to mediate important biological functions in somatic cells. In human spermatozoa, the present study showed that H(2)O(2) induced H2AX phosphorylation in a time- and dose-dependent manner. Moreover, such effect could be abolished by the phosphatidylinositol 3-kinase inhibitor wortmannin. Meanwhile, the neutral comet assay also revealed DSBs production in correlation with H2AX phosphorylation assessed by flow cytometry. Besides H2AX phosphorylation, two other collaborating proteins, Rad50 and 53BP1, were also generated in spermatozoa after H(2)O(2) exposure. However, unlike in somatic FL cells, there were no distinctive focuses, but rather a whole nuclei staining pattern of these three proteins in spermatozoa. Additionally, gammaH2AX (the phosphorylated form of H2AX) staining in spermatozoa persisted despite the fact of a decrease in the number of gammaH2AX foci in FL cells after H(2)O(2) removal. Collectively, these results demonstrate that oxidative stress can induce H2AX phosphorylation in human spermatozoa through DSB induction, and that gammaH2AX may be used as a sensitive, novel marker for such DSBs. Moreover, the surveillance system involving gammaH2AX, Rad50, and 53BP1 in human spermatozoa cannot function effectively in DNA repair, but this system may possess other biological functions in response to DSBs.
Li
Zhongxiang
Z
Center for Reproductive Medicine, Women's Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang 310006, China.
Yang
Jun
J
Huang
Hefeng
H
D017766:Acid Anhydride Hydrolases; D015415:Biomarkers; D004249:DNA Damage; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins; D006657:Histones / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008297:Male; D018384:Oxidative Stress / Q000502:physiology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; D010766:Phosphorylation; D013094:Spermatozoa / Q000378:metabolism*; D000071857:Tumor Suppressor p53-Binding Protein 1
D015415:Biomarkers; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D047908:Intracellular Signaling Peptides and Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; C498301:TP53BP1 protein, human; D000071857:Tumor Suppressor p53-Binding Protein 1; D006861:Hydrogen Peroxide; D017766:Acid Anhydride Hydrolases; C112158:RAD50 protein, human; D045643:DNA Repair Enzymes