Inhibitors of tyrosine and Ser/Thr phosphatases modulate the heat shock response.
Following heat shock the expression of heat shock genes is regulated by the heat shock transcription factor, HSF, known to bind to arrays of the heat shock element, NGAAN, upstream of the heat shock genes. Phosphorylation of HSF is necessary for its activation. We report that the treatment of Chinese hamster HA-1 cells with 250 nM of okadaic acid (OA), a ser/thr phosphatase inhibitor, leads to an increase in activated HSF after heat shock. This is followed by the activation of the transcription of heat shock genes as assayed by the increase in the synthesis of beta-galactosidase in an HA-1 cell line containing the heat shock promoter ligated to the beta-galactosidase gene. To investigate the specificity of OA, we used other phosphatase inhibitors. We found that treatment of HA-1 cells with 500 microM of sodium vanadate, an inhibitor of tyr/phosphatases, resulted in a three to fivefold reduction in HSF activation and binding to the heat shock element following heat shock. Such reduction in HSF activation virtually abolished beta-galactosidase induction. Reduced HSP synthesis was further confirmed by SDS-PAGE and Western blot analysis using anti-HSP-70 and 28 antibodies. Sodium vanadate treatment of heat shocked cells greatly reduced levels of thermotolerance. These results show that ser/thr and specifically tyr/phosphatase inhibitors modulate the signal transduction pathway of HSF activation.
Mivechi
N F
NF
Department of Radiation Oncology, Stanford University, California 94305.
Murai
T
T
Hahn
G M
GM
D000818:Animals; D016466:CHO Cells; D002467:Cell Nucleus / Q000187:drug effects; D006224:Cricetinae; D004988:Ethers, Cyclic / Q000494:pharmacology*; D006360:Heat-Shock Proteins / Q000096:biosynthesis*; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors*; D014158:Transcription, Genetic / Q000187:drug effects; D014638:Vanadates / Q000494:pharmacology*
D004988:Ethers, Cyclic; D006360:Heat-Shock Proteins; D019319:Okadaic Acid; D014638:Vanadates; D010749:Phosphoprotein Phosphatases; D017027:Protein Tyrosine Phosphatases