Division of Molecular Pharmacology and Pharmacogenomics, Department of Biochemistry and Molecular Biology, Kobe University Graduate School of Medicine, Kobe, Japan.
PLoS One. 2012;7(7):e41946. doi: 10.1371/journal.pone.0041946. Epub 2012 Jul 25.
We previously identified Cis4, a zinc transporter belonging to the cation diffusion facilitator protein family, and we demonstrated that Cis4 is implicated in Golgi membrane trafficking in fission yeast. Here, we identified three glycosylphosphatidylinositol (GPI)-anchored proteins, namely Ecm33, Aah3, and Gaz2, as multicopy suppressors of the MgCl(2)-sensitive phenotype of cis4-1 mutant. The phenotypes of ecm33, aah3 and gaz2 deletion cells were distinct from each other, and Cis4 overexpression suppressed Δecm33 phenotypes but did not suppress Δaah3 defects. Notably, green fluorescent protein-tagged Ecm33, which was observed at the cell surface in wild-type cells, mostly localized as intracellular dots that are presumed to be the Golgi and endosomes in membrane-trafficking mutants, including Δapm1, ypt3-i5, and chc1-1 mutants. Interestingly, all these membrane-trafficking mutants showed hypersensitivity to BE49385A, an inhibitor of Its8 that is involved in GPI-anchored protein synthesis. Taken together, these results suggest that GPI-anchored proteins are transported through a clathrin-mediated post-Golgi membrane trafficking pathway and that zinc transporter Cis4 may play roles in membrane trafficking of GPI-anchored proteins in fission yeast.
我们之前鉴定了 Cis4，一种锌转运蛋白，属于阳离子扩散促进蛋白家族，我们证明 Cis4 参与了裂殖酵母高尔基体膜运输。在这里，我们鉴定了三种糖基磷脂酰肌醇（GPI）锚定蛋白，即 Ecm33、Aah3 和 Gaz2，它们是 cis4-1 突变体中 MgCl2 敏感表型的多拷贝抑制子。ecm33、aah3 和 gaz2 缺失细胞的表型彼此不同，Cis4 过表达抑制了Δecm33 表型，但不能抑制Δaah3 缺陷。值得注意的是，在野生型细胞中观察到的绿色荧光蛋白标记的 Ecm33，主要定位于膜运输突变体（包括Δapm1、ypt3-i5 和 chc1-1 突变体）中的细胞内点，这些点被认为是高尔基体和内体。有趣的是，所有这些膜运输突变体对 BE49385A 均表现出超敏反应，BE49385A 是 Its8 的抑制剂，Its8 参与 GPI 锚定蛋白的合成。总之，这些结果表明 GPI 锚定蛋白通过网格蛋白介导的高尔基体后膜运输途径进行运输，锌转运蛋白 Cis4 可能在裂殖酵母 GPI 锚定蛋白的膜运输中发挥作用。
