Dept. of Genetics and Biotechnology, Saint Petersburg State University, St. Petersburg, Russia.
Vavilov Institute of General Genetics, St. Petersburg Branch, Russian Academy of Sciences, St. Petersburg, Russia.
PLoS One. 2018 Mar 1;13(3):e0193726. doi: 10.1371/journal.pone.0193726. eCollection 2018.
The Sch9 kinase of Saccharomyces cerevisiae is one of the major TOR pathway effectors and regulates diverse processes in the cell. Sch9 belongs to the AGC kinase family. In human, amplification of AGC kinase genes is connected with cancer. However, not much is known about the effects of Sch9 overproduction in yeast cells. To fill this gap, we developed a model system to monitor subcellular location and aggregation state of overproduced Sch9 or its regions fused to a fluorescent protein. With this system, we showed that Sch9-YFP forms detergent-resistant aggregates, and multiple protein regions are responsible for this. This finding corroborated the fact that Sch9-YFP is visualized as various fluorescent foci. In addition, we found that Sch9 overproduction caused cell elongation, and this effect was determined by its C-terminal region containing kinase domains. The constructs we present can be exploited to create superior yeast-based model systems to study processes behind kinase overproduction in cancers.
酿酒酵母中的 Sch9 激酶是 TOR 通路的主要效应物之一，调节细胞中的多种过程。Sch9 属于 AGC 激酶家族。在人类中，AGC 激酶基因的扩增与癌症有关。然而，对于酵母细胞中 Sch9 过表达的影响知之甚少。为了填补这一空白，我们开发了一个模型系统来监测过表达的 Sch9 或其与荧光蛋白融合的区域的亚细胞定位和聚集状态。使用该系统，我们表明 Sch9-YFP 形成耐去污剂的聚集体，并且多个蛋白区域对此负责。这一发现证实了 Sch9-YFP 被可视化成各种荧光焦点的事实。此外，我们发现 Sch9 的过表达导致细胞伸长，而这种效应是由其包含激酶结构域的 C 末端区域决定的。我们提出的这些构建体可用于创建更优越的基于酵母的模型系统，以研究癌症中激酶过表达背后的过程。
