Moser M J, Geiser J R, Davis T N
Department of Biochemistry, University of Washington, Seattle 98195, USA.
Mol Cell Biol. 1996 Sep;16(9):4824-31. doi: 10.1128/MCB.16.9.4824.
The cmd1-6 allele contains three mutations that block Ca2+ binding to calmodulin from Saccharomyces cerevisiae. We find that strains containing cmd1-6 lose viability during cell cycle arrest induced by the mating pheromone alpha-factor. The 50% lethal dose (LD50) of alpha-factor for the calmodulin mutant is almost fivefold below the LD50 for a wild-type strain. The calmodulin mutants are not more sensitive to alpha-factor, as measured by activation of a pheromone-responsive reporter gene. Two observations indicate that activation of the Ca2+-calmodulin-dependent protein phosphatase calcineurin contributes to survival of pheromone-induced arrest. First, deletion of the gene encoding the calcineurin regulatory B subunit, CNB1, from a wild-type strain decreases the LD50 of alpha-factor but has no further effect on a cmd1-6 strain. Second, a dominant constitutive calcineurin mutant partially restores the ability of the cmd1-6 strain to survive exposure to alpha-factor. Activation of the Ca2+-calmodulin-dependent protein kinase (CaMK) also contributes to survival, thus revealing a new function for this enzyme. Deletion of the CMK1 and CMK2 genes, which encode CaMK, decreases the LD50 of pheromone compared with that for a wild-type strain but again has no effect in a cmd1-6 strain. Furthermore, the LD50 of alpha-factor for a mutant in which the calcineurin and CaMK genes have been deleted is the same as that for the calmodulin mutant. Finally, the CaMK and calcineurin pathways appear to be independent since the ability of constitutive calcineurin to rescue a cmd1-6 strain is not blocked by deletion of the CaMK genes.
cmd1-6等位基因包含三个突变，这些突变会阻止Ca2+与酿酒酵母的钙调蛋白结合。我们发现，含有cmd1-6的菌株在由交配信息素α因子诱导的细胞周期停滞期间失去活力。钙调蛋白突变体对α因子的50%致死剂量（LD50）几乎比野生型菌株的LD50低五倍。通过信息素响应报告基因的激活来衡量，钙调蛋白突变体对α因子并不更敏感。两项观察结果表明，Ca2+ - 钙调蛋白依赖性蛋白磷酸酶钙调神经磷酸酶的激活有助于信息素诱导的停滞状态下的存活。首先，从野生型菌株中删除编码钙调神经磷酸酶调节B亚基CNB1的基因会降低α因子 的LD50，但对cmd1-6菌株没有进一步影响。其次，显性组成型钙调神经磷酸酶突变体部分恢复了cmd1-6菌株在暴露于α因子时的存活能力。Ca2+ - 钙调蛋白依赖性蛋白激酶（CaMK）的激活也有助于存活，从而揭示了这种酶的新功能。删除编码CaMK的CMK1和CMK2基因，与野生型菌株相比，会降低信息素的LD50，但对cmd1-6菌株同样没有影响。此外，钙调神经磷酸酶和CaMK基因已被删除的突变体对α因子的LD50与钙调蛋白突变体的相同。最后，CaMK和钙调神经磷酸酶途径似乎是独立的，因为组成型钙调神经磷酸酶拯救cmd1-6菌株的能力不会被CaMK基因的删除所阻断。
