Conditional expression of harpin < inf > Pss < /inf > causes yeast cell death that shares features of cell death pathway with harpin < inf > Pss < /inf > -mediated plant hypersensitive response (HR)

Abstract

Conditional expression of harpinPss causes yeast cell death that shares features of cell death pathway with harpinPss-mediated plant hypersensitive response (HR). Pseudomonas syringae pv. syringae 61 hrpZ gene encodes harpinPss, a 34.7 kD extracellular protein that elicits a hypersensitive response (HR) in plants. Conditional expression of either full-length or truncated hrpZ sequences under the GAL1 promoter caused cell death in Saccharomyces cerevisiae Y187. Plating of pYEUT-hrpZ transformants on a medium containing galactose resulted in complete inhibition of colony formation, whereas their growth on a glucose-based medium was unaffected. Western blot analysis confirmed the expression of harpinPss in yeast cells transformed with pYEUT-hrpZ and grown in galactose-containing medium. A time-dependent decline in the percentage of trypan blue-excluding cells in cultures of pYEUT-hrpZ transformants was observed when cultured on galactose-containing medium. Similarly, the number of viable cells reduced to about 50% within 6 h. There were similarities in the harpinPss-mediated cell death in plants and yeast cell death (YCD). Galactose-induced cell death in pYEUT-hrpZ transformants of S. cerevisiae Y187 was suppressed by a protein kinase inhibitor K252a (10 μM). The viability of pYEUT-hrpZ transformants was prolonged in the presence of 100 U ml-1 catalase suggesting a role for the oxidative burst in YCD that was further supported by the flow cytometric patterns of propidium iodide uptake by yeast cells. Overall, it appears that yeast provides a useful model system to understand the molecular mechanism of harpinPss-mediated cell death. © 2001 Academic Press.