Schizosaccharomyces pombe ehs1p is involved in maintaining cell wall integrity and in calcium uptake

Abstract

The Schizosaccharomyces pombe ehs1-1 mutant was isolated on the basis of its hypersensitivity to Echinocandin and Calcofluor White, which inhibit cell wall synthesis. The mutant shows a thermosensitive growth phenotype that is suppressed in the presence of an osmotic stabiliser. The mutant also showed other cell wall-associated phenotypes, such as enhanced sensitivity to enzymatic cell wall degradation and an imbalance in polysaccharide synthesis. The ehs1 ⁺ gene encodes a predicted integral membrane protein that is 30% identical to Saccharomyces cerevisiae Mid1p, a protein that has been proposed to form part of a calcium channel. As expected for such a function, we found that ehs1 ⁺ is involved in intracellular Ca²⁺ accumulation. High external Ca²⁺ concentrations suppressed all phenotypes associated with the ehs1 null mutation, suggesting that the cell integrity defects of ehs1 mutants result from inadequate levels of calcium in the cell. We observed a genetic relationship between ehs1 ⁺ and the protein kinase C homologue pck2 ⁺. pck2 ⁺ suppressed all phenotypes of ehs1-1 mutant cells. Overproduction of pck2p is deleterious to wild-type cells, increasing 1,3-β-D-glucan synthase activity and promoting accumulation of extremely high levels of Ca²⁺. The lethality associated with pck2p, the increase in 1,3-β-D-glucan synthase production and the strong Ca²⁺ accumulation are all dependent on the presence of ehs1p. Our results suggest that in fission yeast ehs1p forms part of a calcium channel that is involved in the cell wall integrity pathway that includes the kinase pck2p.