Department of Biological Science and Technology, Tokyo University of Science, Niijyuku, Katsushika-ku, Tokyo, Japan.
School of Health Science, Tokyo University of Technology, Nishikamada, Ota-ku, Tokyo, Japan.
PLoS One. 2019 Jan 25;14(1):e0210223. doi: 10.1371/journal.pone.0210223. eCollection 2019.
The small GTPases, Rab5 and Rab7, are key regulators at multiple stages of the endocytic/endolysosomal pathway, including fusion and maturation of endosomes. In yeast, Vps21p (Rab5 homolog) recruits a GEF for Rab7 and activates the downstream Ypt7p (Rab7 homolog) on endosomal membrane. Although the model of this sequential activation from Vps21p to Ypt7p in the endocytic pathway has been established, activation mechanism of Ypt7p in the Vps21p-independent pathway has not been completely clarified. Here we show that Ypt7p is activated and mediates vacuolar fusion in cells lacking all yeast Rab5 genes, VPS21, YPT52, and YPT53. We also demonstrate that deletion of both VPS21 and YPT7 genes cause severe defect in the AP-3 pathway as well as the CPY pathway although the AP-3 pathway is mostly intact in each vps21Δ or ypt7Δ mutant. Interestingly, in vps21Δ ypt7Δ mutant cargos trafficked via the VPS or endocytic pathway accumulate beside nucleus whereas cargo trafficked via the AP-3 pathway disperse in the cytosol. These findings suggest that Ypt7p is activated and plays a Rab5-independent role in the AP-3-mediated pathway.
小分子 GTP 酶 Rab5 和 Rab7 是内吞/内溶酶体途径多个阶段的关键调节因子，包括内体融合和成熟。在酵母中，Vps21p（Rab5 同源物）招募 Rab7 的 GEF 并激活内体膜上的下游 Ypt7p（Rab7 同源物）。虽然内吞途径中 Vps21p 到 Ypt7p 的这种顺序激活模型已经建立，但 Vps21p 非依赖性途径中 Ypt7p 的激活机制尚未完全阐明。在这里，我们表明在缺乏所有酵母 Rab5 基因、VPS21、YPT52 和 YPT53 的细胞中，Ypt7p 被激活并介导液泡融合。我们还证明，尽管在每个 vps21Δ 或 ypt7Δ 突变体中，AP-3 途径大部分完整，但 VPS21 和 YPT7 基因的缺失都会导致 AP-3 途径和 CPY 途径严重缺陷。有趣的是，在 vps21Δ ypt7Δ 突变体中，通过 VPS 或内吞途径运输的货物在核旁积累，而通过 AP-3 途径运输的货物则分散在细胞质中。这些发现表明 Ypt7p 在 AP-3 介导的途径中被激活并发挥 Rab5 非依赖性作用。
