Abstract
The pkcA gene, which encodes a protein kinase C (PKC) in the filamentous fungus Aspergillus nidulans, is essential for its viability. However, little is known about its functions. To address this issue, we constructed and characterized temperature-sensitive mutants of pkcA. The conidia of these mutants swelled slightly and exhibited apoptotic phenotypes at 42°C. The apoptotic phenotypes were suppressed by an osmotic stabilizer. Under these conditions, the conidia swelled extensively and did not form germ tubes. Moreover, polarized distribution of F-actin was not observed. We then utilized deletion mutants of bckA, an ortholog of Saccharomyces cerevisiae bck1 that encodes a mitogen-activated protein (MAP) kinase kinase kinase and functions downstream of PKC in the cell wall integrity pathway. These mutants exhibited apoptotic phenotypes at 42°C, but they did not show defects in polarity establishment under osmotically stabilized conditions. These results suggest that PkcA plays multiple roles during germination under conditions of heat stress. The first of these roles is the suppression of apoptosis induction, while the other involves polarity establishment. The former depends on the MAP kinase cascade, whereas the latter does not. In addition, repolarization, which was observed after depolarization in the wild-type strain and the bckA deletion mutant under conditions of heat stress, was not observed in the pkcA-ts mutant. This suggests that PkcA also plays role in polarity establishment during hyphal growth independent of the MAP kinase cascade under these conditions.
Highlights
Protein kinase C (PKC) is a serine/threonine kinase that is conserved among eukaryotes and is of central importance in various signal-transduction processes
Our results suggest that PkcA is involved in the suppression of apoptosis induction and in polarity establishment under conditions of heat stress
Since Pro1023 of Pkc1p is conserved in the amino acid sequence of PkcA of A. nidulans (Pro959 of PkcA), we constructed pkcA-tsLA-1 strain (pkcA-ts)-2, 23, and 25 strains that produce mutated PkcA in which Pro959 was replaced by Leu; we characterized these strains (Table 1)
Summary
Protein kinase C (PKC) is a serine/threonine kinase that is conserved among eukaryotes and is of central importance in various signal-transduction processes. The budding yeast Saccharomyces cerevisiae has only 1 PKC-encoding gene, i.e., pkc. The budding yeast Saccharomyces cerevisiae has only 1 PKC-encoding gene, i.e., pkc1 Deletion of this gene is lethal under normal conditions; this lethality is suppressed by osmotic stabilization. When cells are exposed to cell wall stress, Pkc1p is activated and phosphorylates Bck1p, resulting in the activation of a mitogen-activated protein (MAP) kinase cascade. This MAP kinase cascade includes Bck1p, Mkk1p and Mkk2p, and Slt2p, as a MAPKKK, MAPKKs, and a MAPK, respectively. Slt2p phosphorylates and activates a transcription factor, Rlm1p, which regulates the expression of many genes whose products are involved in cell wall biosynthesis [4]. Pkc1p is suggested to have other functions in polarity establishment, cell-cycle control, regulation of oligosaccharyl transferase, and control of chitin synthase distribution [2,3]
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