Comprehensive knockout analysis of the Rab family GTPases in epithelial cells
Comprehensive knockout analysis of the Rab family GTPases in epithelial cells
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DOI：
10.1083/jcb.201810134
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发表时间：
2019-06-01
影响因子：
7.8
通讯作者：
Fukuda, Mitsunori
中科院分区：
文献类型：
--
作者：
Homma, Yuta;Kinoshita, Riko;Fukuda, Mitsunori
在细胞膜内运输的调节因子中，小gtpase的Rab家族包含的蛋白质数量最多（哺乳动物中约为60个），但许多Rabs的确切功能以及Rabs的功能冗余和多样性在很大程度上仍然未知。在这里，我们为整个Rab家族生成了一个全面的敲除（KO） MDCK细胞集合。我们同时敲除密切相关的类似物（Rab亚家族敲除）以规避功能补偿，发现Rab1A/B和Rab5A/B/C对细胞存活和/或生长至关重要。此外，我们证明Rab6-KO细胞缺乏基底膜，可能是因为无法分泌细胞外基质成分。进一步分析表明Rab6对可溶性物质分泌的一般需求。在Rab6-KO细胞中，跨膜货物到质膜的运输也明显延迟，但表型相对较轻。我们的rabo - ko系列具有相同的背景，将成为分析各种膜运输事件的宝贵资源。
The Rab family of small GTPases comprises the largest number of proteins (similar to 60 in mammals) among the regulators of intracellular membrane trafficking, but the precise function of many Rabs and the functional redundancy and diversity of Rabs remain largely unknown. Here, we generated a comprehensive collection of knockout (KO) MDCK cells for the entire Rab family. We knocked out closely related paralogs simultaneously (Rab subfamily knockout) to circumvent functional compensation and found that Rab1A/B and Rab5A/B/C are critical for cell survival and/or growth. In addition, we demonstrated that Rab6-KO cells lack the basement membrane, likely because of the inability to secrete extracellular matrix components. Further analysis revealed the general requirement of Rab6 for secretion of soluble cargos. Transport of transmembrane cargos to the plasma membrane was also significantly delayed in Rab6-KO cells, but the phenotype was relatively mild. Our Rab-KO collection, which shares the same background, would be a valuable resource for analyzing a variety of membrane trafficking events.