CALCINEURIN, THE CA2+ CALMODULIN-DEPENDENT PROTEIN PHOSPHATASE, IS ESSENTIAL IN YEAST MUTANTS WITH CELL INTEGRITY DEFECTS AND IN MUTANTS THAT LACK A FUNCTIONAL VACUOLAR H+-ATPASE/

Calcineurin is a conserved Ca2+/calmodulin-dependent protein phosphata se that plays a critical role in Ca2+-mediated signaling in many cells . Yeast cells lacking functional calcineurin (cna1 cna2 or cnb1 mutant s) display growth defects under specific environmental conditions, for example, in the presence of high concentrations of Na+, Li+, Mn2+, or OH- but are indistinguishable from wild-type cells under standard cul ture conditions. To characterize regulatory pathways that may overlap with calcineurin, we performed a synthetic lethal screen to identify m utants that require calcineurin on standard growth media. The characte rization of one such mutant, cnd1-8, is presented. The CND1 gene was c loned, and sequence analysis predicts that it encodes a novel protein 1,876 amino acids in length with multiple membrane-spanning domains. C ND1 is identical to the gene identified previously as FKS1, ETG1, and CWH53. cnd1 mutants are sensitive to FK506 and cyclosporin A and exhib it slow growth that is improved by the addition of osmotic stabilizing agents. This osmotic agent-remedial growth defect and microscopic evi dence of spontaneous cell lysis in cnd1 cultures suggest that cell int egrity is compromised in these mutants. Mutations in the genes for yea st protein kinase C (pkc1) and a MAP kinase (mpk1/slt2) disrupt a Ca2-dependent signaling pathway required to maintain a normal cell wall a nd cell integrity. We show that pkc1 and mpk1/slt2 growth defects are more severe in the absence of calcineurin function and less severe in the presence of a constitutively active form of calcineurin. These obs ervations suggest that calcineurin and protein kinase C perform indepe ndent but physiologically related functions in yeast cells. We show th at several mutants that lack a functional vacuolar H+-ATPase (vma) req uire calcineurin for vegetative growth. We discuss possible roles for calcineurin in regulating intracellular ion homeostasis and in maintai ning cell integrity.