Atypical protein kinase C (aPKC) isozymes are unique in the PKC superfamily in that they are not regulated by the lipid second messenger diacylglycerol, which has led to speculation about whether a different second messenger acutely controls their function. Here, using a genetically encoded reporter that we designed, aPKC-specific C kinase activity reporter (aCKAR), we found that the lipid mediator sphingosine 1-phosphate (S1P) promoted the cellular activity of aPKC. Intracellular S1P directly bound to the purified kinase domain of aPKC and relieved autoinhibitory constraints, thereby activating the kinase. In silico studies identified potential binding sites on the kinase domain, one of which was validated biochemically. In HeLa cells, S1P-dependent activation of aPKC suppressed apoptosis. Together, our findings identify a previously undescribed molecular mechanism of aPKC regulation, a molecular target for S1P in cell survival regulation, and a tool to further explore the biochemical and biological functions of aPKC.

非典型蛋白激酶 C (aPKC) 同工酶在 PKC 超家族中是独一无二的，因为它们不受脂质第二信使二酰甘油的调节，这引发了人们对是否有不同的第二信使敏锐地控制其功能的猜测。在这里，使用我们设计的基因编码报告基因，aPKC 特异性 C 激酶活性报告基因 (aCKAR)，我们发现脂质介质 1-磷酸鞘氨醇 (S1P) 促进 aPKC 的细胞活性。细胞内的S1P直接与aPKC的纯化激酶结构域结合并解除自身抑制限制，从而激活激酶。计算机研究确定了激酶结构域上的潜在结合位点，其中之一经过生化验证。在 HeLa 细胞中，S1P 依赖性 aPKC 激活抑制细胞凋亡。总之，我们的研究结果确定了先前未描述的 aPKC 调节分子机制、S1P 在细胞存活调节中的分子靶标，以及进一步探索 aPKC 生化和生物学功能的工具。