Tyrosine phosphorylation in DNA damage and cell death in hypoxic injury to LLC-PK1 cells.
Hypoxia is classically considered to result in a necrotic form of cell injury. We have recently demonstrated a role of endonuclease activation, considered a feature of apoptosis, in DNA damage and cell death in chemical hypoxic injury to renal tubular epithelial cells (LLC-PK1 cells). Tyrosine phosphorylation has been implicated to be involved in cell signaling pathway leading to cell growth, proliferation, and apoptotic death. However, a role of tyrosine phosphorylation as a signal transduction pathway involved in DNA damage and cell death has not been previously examined in hypoxic injury in any tissue. In the present study, we have demonstrated that chemical hypoxia with a combination of antimycin A, a mitochondrial respiration inhibitor, and substrate deprivation resulted in rapid increase in protein tyrosine kinases activity and protein tyrosine phosphorylation prior to any evidence of cell death in LLC-PK1 cells. The inhibitors of protein tyrosine kinases, genistein, lavendustin A, tyrphostin, and herbimycin A provided a marked protection against chemical hypoxia-induced DNA damage (as measured by alkaline unwinding assay) and cell death (as measured by trypan blue exclusion assay). In a separate study, we confirmed the ability of the inhibitors, lavendustin A and herbimycin A to prevent chemical hypoxia-induced increase in protein tyrosine kinases activity and protein tyrosine phosphorylation. In addition, the inhibitors used did not affect ATP depletion induced by antimycin A, suggesting that the inhibitors do not alter cellular uptake of antimycin A. Taken together, our data provide a strong evidence that tyrosine phosphorylation plays as important role in DNA damage and cell death in chemical hypoxic injury to renal tubular epithelial cells.
Hagar
H
H
Division of Nephrology, University of Arkansas for Medical Sciences, Little Rock, USA.
Ueda
N
N
Shah
S V
SV
D000818:Animals; D016227:Benzoquinones; D016923:Cell Death; D004249:DNA Damage; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000860:Hypoxia / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*; D018374:LLC-PK1 Cells / Q000502:physiology; D047029:Lactams, Macrocyclic; D010636:Phenols / Q000494:pharmacology; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D013552:Swine; D014443:Tyrosine / Q000378:metabolism*
D016227:Benzoquinones; D004791:Enzyme Inhibitors; D047029:Lactams, Macrocyclic; D010636:Phenols; D011809:Quinones; D017828:Rifabutin; C062898:lavendustin A; D014443:Tyrosine; C020754:herbimycin; D011505:Protein-Tyrosine Kinases