Laboratory of Molecular Microbiology, Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan; Laboratory of Molecular Function of Food, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
Laboratory of Molecular Microbiology, Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan.
Biochim Biophys Acta Gen Subj. 2021 May;1865(5):129853. doi: 10.1016/j.bbagen.2021.129853. Epub 2021 Jan 26.
A serine/threonine kinase Pkc1 is the sole protein kinase C in the budding yeast Saccharomyces cerevisiae, and plays an important role in the regulation of polarized growth and stress responses such as those due to heat shock. Exposure of cells to high temperature transiently arrests polarized growth and leads to depolarization of the actin cytoskeleton, followed by actin repolarization during adaptation to heat shock stress. Actin repolarization is ensured by the activation of Pkc1 signaling; however, the molecular mechanisms underlying this phenomenon remain poorly understood.
Using an overexpression construct of a constitutively active mutant of Pkc1 (Pkc1), we explored the Pkc1 target molecules involved in actin repolarization.
PKC1 overexpression as well as heat shock stress increased the phosphorylation levels of Rho GTPase-activating protein (RhoGAP) Rgd1. Rgd1 was found to contribute to Pkc1-signaling-related actin repolarization during adaptation to heat shock stress in a GAP activity-independent manner, with Ser148 in Rgd1 playing a crucial role. Furthermore, Rgd1 was involved in the maintenance of phosphorylation status of the mitogen-activated protein (MAP) kinase Mpk1, a downstream effector of Pkc1, under heat shock stress.
Rgd1 is a target of Pkc1 signaling under conditions of heat shock stress, and required for the normal process of actin repolarization during adaptation to heat shock stress.
Our results provide insights into the molecular mechanism underlying Pkc1-mediated modulation of actin repolarization under heat shock stress.
丝氨酸/苏氨酸激酶 Pkc1 是芽殖酵母酿酒酵母中唯一的蛋白激酶 C，在调节极性生长和应激反应（如热休克）中发挥重要作用。细胞暴露于高温会暂时停止极性生长，并导致肌动蛋白细胞骨架去极化，随后在适应热休克应激时重新极化。肌动蛋白的重新极化是通过 Pkc1 信号的激活来保证的；然而，这一现象的分子机制仍知之甚少。
我们使用 Pkc1 的组成性激活突变体（Pkc1）的过表达构建体，探索了参与肌动蛋白重新极化的 Pkc1 靶分子。
Pkc1 的过表达以及热休克应激均增加了 Rho GTP 酶激活蛋白（RhoGAP）Rgd1 的磷酸化水平。发现 Rgd1 以非 GAP 活性依赖的方式有助于适应热休克应激过程中的 Pkc1 信号相关肌动蛋白重新极化，其中 Rgd1 中的 Ser148 起着关键作用。此外，Rgd1 参与了 Pkc1 的下游效应物丝裂原激活蛋白（MAP）激酶 Mpk1 的磷酸化状态的维持，在热休克应激下。
Rgd1 是热休克应激条件下 Pkc1 信号的靶标，是适应热休克应激过程中肌动蛋白重新极化的正常过程所必需的。
我们的结果为 Pkc1 介导的热休克应激下肌动蛋白重新极化的分子机制提供了新的见解。
