Devasahayam Gina, Ritz Danilo, Helliwell Stephen B, Burke Daniel J, Sturgill Thomas W
Departments of Pharmacology, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA.
Proc Natl Acad Sci U S A. 2006 Nov 21;103(47):17840-5. doi: 10.1073/pnas.0604303103. Epub 2006 Nov 9.
The rapamycin.FKBP12 complex inhibits target of rapamycin (TOR) kinase in TORC1. We screened the yeast nonessential gene deletion collection to identify mutants that conferred rapamycin resistance, and we identified PMR1, encoding the Golgi Ca2+/Mn2+ -ATPase. Deleting PMR1 in two genetic backgrounds confers rapamycin resistance. Epistasis analyses show that Pmr1 functions upstream from Npr1 and Gln-3 in opposition to Lst8, a regulator of TOR. Npr1 kinase is largely cytoplasmic, and a portion localizes to the Golgi where amino acid permeases are modified and sorted. Nuclear translocation of Gln-3 and Gln-3 reporter activity in pmr1 cells are impaired, but expression of functional Gap1 in the plasma membrane of a pmr1 strain in response to nitrogen limitation is enhanced. These two phenotypes suggest up-regulation of Npr1 function in the absence of Pmr1. Together, our results establish that Pmr1-dependent Ca2+ and/or Mn2+ ion homeostasis is necessary for TOR signaling.
雷帕霉素与FKBP12复合物可抑制TORC1中的雷帕霉素靶蛋白(TOR)激酶。我们筛选了酵母非必需基因缺失文库,以鉴定赋予雷帕霉素抗性的突变体,并鉴定出了编码高尔基体Ca2+/Mn2+ -ATP酶的PMR1。在两种遗传背景下删除PMR1均可赋予雷帕霉素抗性。上位性分析表明,Pmr1在与TOR的调节因子Lst8相反的方向上,在Npr1和Gln-3的上游发挥作用。Npr1激酶主要位于细胞质中,有一部分定位于高尔基体,在高尔基体中氨基酸通透酶会被修饰和分选。pmr1细胞中Gln-3的核转位及Gln-3报告基因活性受损,但在氮限制条件下,pmr1菌株质膜中功能性Gap1的表达会增强。这两种表型表明在缺乏Pmr1的情况下Npr1功能上调。总之,我们的结果表明,Pmr1依赖的Ca2+和/或Mn2+离子稳态对于TOR信号传导是必需的。