Accessory ESCRT-III proteins selectively regulate Rab11-exosome biogenesis in Drosophila secondary cells
Accessory ESCRT-III proteins selectively regulate Rab11-exosome biogenesis in Drosophila secondary cells
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DOI：
10.1101/2020.06.18.158725
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发表时间：
2020-06
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影响因子：
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通讯作者：
Pauline P Marie;Shih‐Jung Fan;C. Mendes;M. Wainwright;A. Harris;D. Goberdhan;Clive Wilson
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文献类型：
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作者：
Pauline P Marie;Shih‐Jung Fan;C. Mendes;M. Wainwright;A. Harris;D. Goberdhan;Clive Wilson
外泌体是具有有效信号传导活性的分泌型纳米囊泡，其最初形成为多囊内体中的管腔内囊泡（ILV），随后与质膜融合。这些ILV在晚期内体和再循环内体中产生，后者由小GTdR ab 11标记，并产生具有不同货物和功能的外泌体。四种转运所需的内体分选复合物（ESCRT）组装体（0-III）内的核心蛋白在晚期内体外泌体生物发生和ILV介导的通过内溶酶体系统的泛素化货物的破坏中起关键的顺序作用。它们还控制其他细胞过程，如胞质分裂和其他囊泡出芽。相比之下，一些辅助ESCRT的功能没有很好地定义。在这里，我们评估ESCRT依赖Rab 11外泌体，使用RNA敲低果蝇次级细胞（SC）的男性附属腺，其中有异常扩大Rab 11阳性车厢。出乎意料的是，不仅在Rab 11-外泌体形成所需的所有四种ESCRT复合物中的核心蛋白，而且还有辅助ESCRT-III蛋白CHMP 1、CHMP 5和IST 1。与大多数核心ESCRT敲除不同，抑制这些辅助蛋白的表达不会影响细胞形态的其他方面，并且不会导致泛素化货物的积累。我们的结论是，辅助ESCRT-III组件在Rab 11-外泌体生成中具有特定的和潜在的泛素独立作用，这可能为阻断这些囊泡在癌症中的促肿瘤活性提供了靶点。
Exosomes are secreted nanovesicles with potent signalling activity that are initially formed as intraluminal vesicles (ILVs) in multivesicular endosomes, which subsequently fuse with the plasma membrane. These ILVs are made in both late endosomes and recycling endosomes, the latter marked by the small GTPase Rab11 and generating exosomes with different cargos and functions. Core proteins within four Endosomal Sorting Complex Required for Transport (ESCRT) assemblies (0-III) play key sequential roles in late endosomal exosome biogenesis and ILV-mediated destruction of ubiquitinylated cargos through the endolysosomal system. They also control additional cellular processes, such as cytokinesis and other vesicle budding. By contrast, the functions of several accessory ESCRTs are not well defined. Here we assess the ESCRT-dependency of Rab11-exosomes, using RNA knockdown in Drosophila secondary cells (SCs) of the male accessory gland, which have unusually enlarged Rab11-positive compartments. Unexpectedly, not only are core proteins in all four ESCRT complexes required for Rab11-exosome formation, but also accessory ESCRT-III proteins, CHMP1, CHMP5 and IST1. Suppressing expression of these accessory proteins does not affect other aspects of cell morphology, unlike most core ESCRT knockdowns, and does not lead to accumulation of ubiquitinylated cargos. We conclude that accessory ESCRT-III components have a specific and potentially ubiquitin-independent role in Rab11-exosome generation, which might provide a target for blocking the pro-tumorigenic activities of these vesicles in cancer.