Kume Kazunori, Koyano Takayuki, Takata Junpei, Wakabayashi Ko, Mizunuma Masaki, Miyakawa Tokichi, Hirata Dai
a Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter , Hiroshima University , 1-3-1 kagamiyama, Higashi-Hiroshima , 739-8530 Japan.
Biosci Biotechnol Biochem. 2015;79(5):790-4. doi: 10.1080/09168451.2014.1003132. Epub 2015 Jan 23.
Calcineurin, which is a Ca(2+)/calmodulin-dependent protein phosphatase, is a key mediator in calcium signaling in diverse biological processes and of clinical importance as the target of the immunosuppressant FK506. To identify a mutant(s) in which calcineurin is activated, inhibiting cellular growth as a result, we screened for a mutant(s) whose temperature sensitivity would be suppressed by FK506 from the budding yeast non-essential gene deletion library. We found that the temperature sensitivity of cells in which the conserved Verprolin VRP1 gene had been deleted, which gene is required for actin organization and endocytosis, was suppressed by either FK506 or by cnb1 deletion. Indeed, the calcineurin activity increased significantly in the ∆vrp1 cells. Finally, we demonstrated that the ∆vrp1 strain to be useful as an indicator in a positive screening for bioactive compounds inhibiting calcineurin.
钙调神经磷酸酶是一种Ca(2+)/钙调蛋白依赖性蛋白磷酸酶，是多种生物过程中钙信号传导的关键介质，并且作为免疫抑制剂FK506的靶点具有临床重要性。为了鉴定钙调神经磷酸酶被激活从而抑制细胞生长的突变体，我们从芽殖酵母非必需基因缺失文库中筛选对FK506温度敏感性有抑制作用的突变体。我们发现，缺失保守的维普洛林VRP1基因（该基因是肌动蛋白组织和内吞作用所必需的）的细胞的温度敏感性可被FK506或cnb1缺失所抑制。实际上，∆vrp1细胞中的钙调神经磷酸酶活性显著增加。最后，我们证明∆vrp1菌株可作为一种指示物，用于阳性筛选抑制钙调神经磷酸酶的生物活性化合物。
