Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, California 94720, USA.
Department of Mechanical Engineering, University of California at Berkeley, Berkeley, California 94720, USA.
Genes Dev. 2018 Dec 1;32(23-24):1576-1590. doi: 10.1101/gad.318709.118. Epub 2018 Nov 26.
target of rapamycin (TOR) complex 2 (TORC2) is an essential regulator of plasma membrane lipid and protein homeostasis. How TORC2 activity is modulated in response to changes in the status of the cell envelope is unclear. Here we document that TORC2 subunit Avo2 is a direct target of Slt2, the mitogen-activated protein kinase (MAPK) of the cell wall integrity pathway. Activation of Slt2 by overexpression of a constitutively active allele of an upstream Slt2 activator (Pkc1) or by auxin-induced degradation of a negative Slt2 regulator (Sln1) caused hyperphosphorylation of Avo2 at its MAPK phosphoacceptor sites in a Slt2-dependent manner and diminished TORC2-mediated phosphorylation of its major downstream effector, protein kinase Ypk1. Deletion of Avo2 or expression of a phosphomimetic Avo2 allele rendered cells sensitive to two stresses (myriocin treatment and elevated exogenous acetic acid) that the cell requires Ypk1 activation by TORC2 to survive. Thus, Avo2 is necessary for optimal TORC2 activity, and Slt2-mediated phosphorylation of Avo2 down-regulates TORC2 signaling. Compared with wild-type Avo2, phosphomimetic Avo2 shows significant displacement from the plasma membrane, suggesting that Slt2 inhibits TORC2 by promoting Avo2 dissociation. Our findings are the first demonstration that TORC2 function is regulated by MAPK-mediated phosphorylation.
雷帕霉素靶蛋白（TOR）复合物 2（TORC2）是质膜脂质和蛋白质动态平衡的重要调节剂。TORC2 活性如何响应细胞外膜状态的变化而调节尚不清楚。在这里，我们记录了 TORC2 亚基 Avo2 是细胞壁完整性途径中丝裂原活化蛋白激酶（MAPK）Slt2 的直接靶标。通过过表达组成型激活的上游 Slt2 激活剂（Pkc1）的等位基因或通过生长素诱导的负性 Slt2 调节剂（Sln1）的降解来激活 Slt2，导致 Avo2 在其 MAPK 磷酸接受位点以 Slt2 依赖的方式发生过度磷酸化，并减少 TORC2 介导的其主要下游效应物蛋白激酶 Ypk1 的磷酸化。Avo2 的缺失或磷酸模拟 Avo2 等位基因的表达使细胞对两种应激（麦角甾醇处理和升高的外源性乙酸）敏感，而细胞需要 Ypk1 通过 TORC2 激活才能存活。因此，Avo2 是 TORC2 活性的必要条件，Slt2 介导的 Avo2 磷酸化下调 TORC2 信号。与野生型 Avo2 相比，磷酸模拟 Avo2 从质膜显著移位，表明 Slt2 通过促进 Avo2 解离来抑制 TORC2。我们的发现首次证明了 TORC2 功能受 MAPK 介导的磷酸化调节。
