Department of Biochemistry and Molecular Biology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.
Faculty of Health Sciences, University of Macau, Avenida da Universidade, Taipa, Macau SAR, China.
PLoS Genet. 2020 May 8;16(5):e1008763. doi: 10.1371/journal.pgen.1008763. eCollection 2020 May.
The ACK family tyrosine kinase SID-3 is involved in the endocytic uptake of double-stranded RNA. Here we identified SID-3 as a previously unappreciated recycling regulator in the Caenorhabditis elegans intestine. The RAB-10 effector EHBP-1 is required for the endosomal localization of SID-3. Accordingly, animals with loss of SID-3 phenocopied the recycling defects observed in ehbp-1 and rab-10 single mutants. Moreover, we detected sequential protein interactions between EHBP-1, SID-3, NCK-1, and DYN-1. In the absence of SID-3, DYN-1 failed to localize at tubular recycling endosomes, and membrane tubules breaking away from endosomes were mostly absent, suggesting that SID-3 acts synergistically with the downstream DYN-1 to promote endosomal tubule fission. In agreement with these observations, overexpression of DYN-1 significantly increased recycling transport in SID-3-deficient cells. Finally, we noticed that loss of RAB-10 or EHBP-1 compromised feeding RNAi efficiency in multiple tissues, implicating basolateral recycling in the transport of RNA silencing signals. Taken together, our study demonstrated that in C. elegans intestinal epithelia, SID-3 acts downstream of EHBP-1 to direct fission of recycling endosomal tubules in concert with NCK-1 and DYN-1.
ACK 家族酪氨酸激酶 SID-3 参与双链 RNA 的内吞摄取。在这里，我们鉴定出 SID-3 是秀丽隐杆线虫肠道中以前未被重视的再循环调节剂。 RAB-10 效应物 EHBP-1 是 SID-3 内体定位所必需的。因此，SID-3 缺失的动物表现出 ehbp-1 和 rab-10 单突变体中观察到的再循环缺陷。此外，我们检测到 EHBP-1、SID-3、NCK-1 和 DYN-1 之间的顺序蛋白相互作用。在 SID-3 缺失的情况下，DYN-1 未能定位在管状再循环内体上，并且从内体上断裂的膜小管大多不存在，这表明 SID-3 与下游的 DYN-1 协同作用以促进内体小管的分裂。与这些观察结果一致，DYN-1 的过表达显著增加了 SID-3 缺陷细胞中的再循环运输。最后，我们注意到 RAB-10 或 EHBP-1 的缺失会损害多种组织中 RNAi 的喂食效率，这表明基底外侧再循环参与了 RNA 沉默信号的运输。总之，我们的研究表明，在秀丽隐杆线虫肠道上皮细胞中，SID-3 在 EHBP-1 下游发挥作用，与 NCK-1 和 DYN-1 一起指导再循环内体小管的分裂。
