Bean Bjorn D M, Davey Michael, Snider Jamie, Jessulat Matthew, Deineko Viktor, Tinney Matthew, Stagljar Igor, Babu Mohan, Conibear Elizabeth
Centre for Molecular Medicine and Therapeutics, Child and Family Research Institute, University of British Columbia, Vancouver, BC V5Z 4H4, Canada.
Donnelly Centre, University of Toronto, Toronto, ON M5S3E1, Canada Department of Biochemistry and Department of Molecular Genetics, University of Toronto, Toronto, ON M5S1A8, Canada.
Mol Biol Cell. 2015 Mar 15;26(6):1119-28. doi: 10.1091/mbc.E14-08-1281. Epub 2015 Jan 21.
The retromer complex facilitates the sorting of integral membrane proteins from the endosome to the late Golgi. In mammalian cells, the efficient recruitment of retromer to endosomes requires the lipid phosphatidylinositol 3-phosphate (PI3P) as well as Rab5 and Rab7 GTPases. However, in yeast, the role of Rabs in recruiting retromer to endosomes is less clear. We identified novel physical interactions between retromer and the Saccharomyces cerevisiae VPS9-domain Rab5-family guanine nucleotide exchange factors (GEFs) Muk1 and Vps9. Furthermore, we identified a new yeast VPS9 domain-containing protein, VARP-like 1 (Vrl1), which is related to the human VARP protein. All three VPS9 domain-containing proteins show localization to endosomes, and the presence of any one of them is necessary for the endosomal recruitment of retromer. We find that expression of an active VPS9-domain protein is required for correct localization of the phosphatidylinositol 3-kinase Vps34 and the production of endosomal PI3P. These results suggest that VPS9 GEFs promote retromer recruitment by establishing PI3P-enriched domains at the endosomal membrane. The interaction of retromer with distinct VPS9 GEFs could thus link GEF-dependent regulatory inputs to the temporal or spatial coordination of retromer assembly or function.
回收体复合物促进整合膜蛋白从内体向晚期高尔基体的分选。在哺乳动物细胞中，回收体有效募集到内体需要脂质磷脂酰肌醇3-磷酸（PI3P）以及Rab5和Rab7 GTP酶。然而，在酵母中，Rabs在内体募集回收体中的作用尚不清楚。我们鉴定了回收体与酿酒酵母VPS9结构域Rab5家族鸟嘌呤核苷酸交换因子（GEFs）Muk1和Vps9之间新的物理相互作用。此外，我们鉴定了一种新的含酵母VPS9结构域的蛋白，VARP样蛋白1（Vrl1），它与人VARP蛋白相关。所有这三种含VPS9结构域的蛋白都定位于内体，并且它们中的任何一种的存在对于回收体在内体的募集都是必需的。我们发现活性VPS9结构域蛋白的表达对于磷脂酰肌醇3-激酶Vps34的正确定位和内体PI3P的产生是必需的。这些结果表明，VPS9 GEFs通过在内体膜上建立富含PI3P的结构域来促进回收体的募集。回收体与不同的VPS9 GEFs的相互作用因此可以将GEF依赖的调节输入与回收体组装或功能的时间或空间协调联系起来。
