Regulation of yeast protein kinase C activity by interaction with the small GTPase Rho1p through its amino‐terminal HR1 domain
Regulation of yeast protein kinase C activity by interaction with the small GTPase Rho1p through its amino‐terminal HR1 domain
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DOI：
10.1046/j.1365-2958.2002.02925.x
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发表时间：
2002-05
影响因子：
3.6
通讯作者：
H. Schmitz;A. Lorberg;J. Heinisch
中科院分区：
文献类型：
--
作者：
H. Schmitz;A. Lorberg;J. Heinisch
来自酿酒酵母的蛋白激酶C（Pkc 1 p）构成蛋白激酶C超家族的原型成员，因为它共享分散在高等真核生物同工酶中的所有保守区域。酵母酶中的一些保守结构域的功能意义尚未研究。我们检查了在酶的氨基末端区域携带部分缺失的菌株，其与蛋白激酶C相关激酶的HR 1同源。该菌株对咖啡因、Calcofluor白色和刚果红的存在敏感，已知所有药物都会影响信号转导途径中有缺陷的突变体，从而确保细胞完整性，其中Pkc 1 p是中心组分。HR 1A中的一个单一点突变的分离，其对上述药物具有敏感性，证实了该区域对于体内蛋白激酶C活性的适当调节的重要性。双杂交分析提供了证据，证明小GTdR Rho 1 p与HR 1A区域的相互作用，除了报道的这种蛋白质与Pkc 1 p的C1区域的相互作用。MAP激酶磷酸化测定表明，这种Rho 1 p-Pkc 1 p/HR 1A相互作用不会导致激酶级联的激活。在这项工作中报告的HR 1A和C1结构域的突变体的基因内致死性意味着Rho 1 p-Pkc 1 p在酵母中的相互作用的重要作用。
Protein kinase C from Saccharomyces cerevisiae (Pkc1p) constitutes a prototypic member of the protein kinase C superfamily, as it shares all the conserved regions scattered among the isoenzymes of higher eukaryotes. The functional significance of some of the conserved domains in the yeast enzyme has not yet been investigated. We examined strains carrying a partial deletion in the amino‐terminal region of the enzyme, which is homologous to the HR1 of the protein kinase C‐related kinases. This strain was sensitive to the presence of caffeine, Calcofluor white and Congo red, all drugs known to affect mutants defective in the signal transduction pathway ensuring cellular integrity in which Pkc1p is a central component. Isolation of a single point mutation in HR1A, which shares the sensitivity to the drugs mentioned, confirmed the importance of this region for proper regulation of protein kinase C activity in vivo. Two‐hybrid analysis provided evidence for an interaction of the small GTPase Rho1p with the HR1A region, in addition to the reported interaction of this protein with the C1 region of Pkc1p. MAP kinase phosphorylation assays indicate that this Rho1p–Pkc1p/HR1A interaction does not result in an activation of the kinase cascade. The intragenic lethality of mutants affected in both HR1A and the C1 domain reported in this work implies an essential role for Rho1p–Pkc1p interaction in yeast.