It’s been reported that treatment with vanadate induces DNA single-strand breaks in human diploid fibroblasts, and mixed publicity of fibroblasts to other genotoxic agents produces DSBs . Thus, DNA injury intensified by vanadate publicity, if any, could activate the signaling pathways main to ATM activation. Besides, it’s been reported that ATM senses not merely DSBs but additionally non-DNA damaging events this kind of as heat shock, glucose starvation and insulin treatment . Taken with each other with all the insulinomimetic effects of vanadate , the signals not derived from DNA harm may well contribute more to your activation of ATM pathway in cells exposed to vanadate. Considering the fact that phosphorylation of p53 at Ser15 was nevertheless present in ATM knockdown cells, mechanisms besides ATM activation might possibly also be involved. Probable activation of other serine/threonine kinases and/or inhibition of protein serine/threonine phosphatases by vanadate exposure remain to be examined.
Following exposure to vanadate, production of reactive oxygen species together with superoxide anion, hydrogen peroxide and hydroxyl radical has been observed in A549 cells as well as other cell varieties . Unexpectedly, remedy with antioxidants such as catalase and N-acetylcysteine failed selleck chemicals small molecule library screening to suppress NaVO3-induced Ser15 phosphorylation in the current examine. Furthermore, therapy with diphenyleneiodonium chloride , an inhibitor of NADPH oxidase, did not suppress NaVO3-induced Ser15 phosphorylation . Thus, phosphorylation of p53 at Ser15 in A549 cells exposed to NaVO3 might possibly be brought about by mechanisms unrelated to reactive oxygen species, as has become proven for Na3VO4- induced cyclooxygenase-2 expression in A549 cells . Consequently, the possible direct interactions involving vanadate and DNA or cellular elements should really be deemed.
Vanadate compounds are already identified to inhibit potently protein tyrosine phosphatases . For this reason, even further scientific studies are expected to clarify regardless of whether activation of tyrosine phosphorylation buy SYR-322 cascade by vanadate publicity also plays a part in the phosphorylation of p53 at Ser15, notably by ATM pathway. To examine the biological significance of vanadate-induced phosphorylation of p53 at Ser15, a plasmid containing S15A mutant p53 was introduced into A549 cells. Inconsistent together with the transactivation of p53 exercise in Cl 41 cells treated with NaVO3 , the binding of p53 to its consensus DNA sequence within the nuclear extracts from A549 cells enhanced in response to NaVO3 publicity. Having said that, the difference in p53DNA binding activity among cells transfected with wild-type and S15A mutant p53 was identified to get reasonably minor .
Additionally, NaVO3-induced accumulation of cytoplasmic nucleosomes was nevertheless observed in cells transfected with S15A mutant p53. Treatment with wortmannin also failed to suppress drastically p53DNA binding exercise and DNA fragmentation induced by NaVO3 publicity .