Calcineurin inhibitors suppress the high-temperature stress sensitivity of the yeast ubiquitin ligase Rsp5 mutant: a new method of screening for calcineurin inhibitors.

Abstract

The ubiquitin/proteasome system plays significant and important roles in the regulation of metabolism of various proteins. The dysfunction of this system is involved in several diseases, for example, cancer, neurogenic diseases and chronic inflammation. Therefore, the compounds, which regulate the ubiquitin/proteasome system, might be candidates for the development use as clinical drugs. The Saccharomyces cerevisiae mutant (rsp5(A401E)) has a single amino acid change, Ala401Glu, in the RSP5 gene, which encodes an essential E3 ubiquitin ligase, is hypersensitive to high-temperature stress. Here, we found that the immunosuppressants FK506 and cyclosporin A, both known as calcineurin inhibitors, complemented the high-temperature stress-induced growth defect of rsp5(A401E) strain. The defect of calcineurin pathway by disrupting the CNB1 and CRZ1 gene also partially complemented the high-temperature stress sensitivity of rsp5(A401E) cells. Thus, these results suggest that inhibition of the calcineurin pathway confers the tolerance to high-temperature stress on rsp5(A401E) cells. Furthermore, some diterpenoid compounds, which restore the growth of rsp5(A401E) cells, showed the activities of calcineurin inhibition and protein phosphatase 2C activation. These results indicate that calcineurin inhibitors suppress the high-temperature stress sensitivity of rsp5(A401E) cells and that analysis of their physiological function is effective for the screening of calcineurin inhibitors in yeast cells.