Division of Molecular Pharmacology and Pharmacogenomics, Department of Biochemistry and Molecular Biology, Kobe University Graduate School of Medicine, Kobe, Japan.
PLoS One. 2011;6(8):e23422. doi: 10.1371/journal.pone.0023422. Epub 2011 Aug 5.
We have been studying calcineurin signal transduction pathway in fission yeast Schizosaccharomyces pombe (S. pombe) by developing a genetic screen for mutants that show hypersensitivity to the immunosuppressive calcineurin inhibitor FK506 (tacrolimus). In the present study, to identify nonessential genes that are functionally related to the calcineurin signaling pathway, we performed a genome-wide screen of 3004 haploid deletion strains and confirmed 72 deletion strains to be FK506 sensitive. These 72 genes are classified into nine functional groups to include membrane trafficking (16 genes), signal transduction (10 genes), ubiquitination (8 genes), chromatin remodeling (6 genes), cytokinesis (4 genes), ribosomal protein (3 genes), RNA binding protein (3 genes), and a variety of other known functions (17 genes) or still unknown functions (5 genes) in the biological system. In our previous screening of FK506-sensitive mutants we isolated several membrane-trafficking mutants showing defective cell wall integrity. Here, we further examined the vacuolar fusion, the v-SNARE synaptobrevin Syb1 localization, and the sensitivity to the β-glucan synthase inhibitor micafungin in these 72 FK506-sensitive strains. Results showed that 25 deletion strains exhibited abnormal vacuole fusion, 19 deletion strains exhibited Syb1 mislocalization, and 14 deletion strains exhibited both abnormal vacuole fusion and Syb1 mislocalization, while 42 deletion strains showed both normal vacuole fusion and Syb1 localization. Likewise, 16 deletion strains showed sensitivity to micafungin. Altogether, our present study indicates that calcineurin mediates a plethora of physiological processes in fission yeast, and that calcineurin is extensively involved in cross-talk between signaling pathways.
我们一直在通过开发一种针对突变体的遗传筛选方法来研究裂殖酵母 Schizosaccharomyces pombe（S. pombe）中的钙调神经磷酸酶信号转导途径，该突变体对免疫抑制性钙调神经磷酸酶抑制剂 FK506（他克莫司）表现出超敏反应。在本研究中，为了鉴定与钙调神经磷酸酶信号通路功能相关的非必需基因，我们对 3004 个单倍体缺失株进行了全基因组筛选，并确认 72 个缺失株对 FK506 敏感。这 72 个基因分为九个功能组，包括膜转运（16 个基因）、信号转导（10 个基因）、泛素化（8 个基因）、染色质重塑（6 个基因）、胞质分裂（4 个基因）、核糖体蛋白（3 个基因）、RNA 结合蛋白（3 个基因）以及生物系统中各种其他已知功能（17 个基因）或仍未知功能（5 个基因）。在我们之前对 FK506 敏感突变体的筛选中，我们分离了一些膜转运突变体，这些突变体显示出细胞壁完整性缺陷。在这里，我们进一步检查了这些 72 个 FK506 敏感株的液泡融合、v-SNARE 突触融合蛋白 Syb1 的定位以及对β-葡聚糖合酶抑制剂米卡芬净的敏感性。结果表明，25 个缺失株表现出异常的液泡融合，19 个缺失株表现出 Syb1 定位错误，14 个缺失株同时表现出液泡融合和 Syb1 定位错误，而 42 个缺失株同时表现出正常的液泡融合和 Syb1 定位。同样，16 个缺失株对米卡芬净敏感。总之，本研究表明钙调神经磷酸酶在裂殖酵母中介导了众多生理过程，并且钙调神经磷酸酶广泛参与信号通路之间的串扰。
