TBC1D9B functions as a GTPase-activating protein for Rab11a in polarized MDCK cells.
TBC1D9B functions as a GTPase-activating protein for Rab11a in polarized MDCK cells.
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TBC1D9B充当极化MDCK细胞中Rab11a的GTPase激活蛋白。
DOI：
10.1091/mbc.e13-10-0604
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发表时间：
2014-11-15
影响因子：
3.3
通讯作者：
Yin XM
中科院分区：
文献类型：
--
作者：
Gallo LI;Liao Y;Ruiz WG;Clayton DR;Li M;Liu YJ;Jiang Y;Fukuda M;Apodaca G;Yin XM
Rab 11 a是囊泡运输过程的关键调节剂，但关于调节其GTP-GDP循环的GEF和GAP的信息有限。TBC 1D 9 B被鉴定为MDCK细胞中的Rab 11 a GAP，在那里它调节Rab 11 a依赖性基底外侧至顶端的胞吞途径。Rab 11 a是囊泡运输过程的关键调节剂，但关于调节其GTP-GDP循环的鸟嘌呤核苷酸交换因子和GTP酶激活蛋白（GAP）的信息有限。我们观察到，在Mg 2+（2.5 mM）的存在下，TBC 1D 9 B通过其Tre 2-Bub 2-Cdc 16（TBC）结构域与Rab 11 a，Rab 11b和Rab 4a以核苷酸依赖性方式相互作用。然而，只有Rab 11 a是TBC 1D 9 B刺激的GTP水解的底物。在限制性Mg 2+浓度（<0.5mM）下，Rab 8a是该GAP的额外底物。在极化Madin-Darby犬肾细胞中，内源性TBC 1D 9 B与Rab 11 a阳性再循环内体共定位，但与EEA 1阳性早期内体、转铁蛋白阳性再循环内体或晚期内体共定位较少。TBC 1D 9 B的过表达，而不是一个失活的突变体，降低了基底外侧到顶端的伊加转胞吞的速率-Rab 11 a依赖的途径-和shRNA介导的TBC 1D 9 B的消耗增加了这一过程的速率。相反，TBC 1D 9 B对两个Rab 11 a独立的通路--转铁蛋白受体的基底外侧再循环或表皮生长因子受体的降解没有影响。最后，TBC 1D 9 B的表达降低了细胞中活性Rab 11 a的量，同时破坏了Rab 11 a与其效应子Sec 15 A之间的相互作用。我们的结论是，TBC 1D 9 B是Rab 11 a GAP，调节极化MDCK细胞的基底侧至顶端的转胞吞。
Rab11a is a key modulator of vesicular trafficking processes, but there is limited information about the GEFs and GAPs that regulate its GTP-GDP cycle. TBC1D9B is identified as a Rab11a GAP in MDCK cells, where it regulates the Rab11a-dependent basolateral-to-apical transcytotic pathway. Rab11a is a key modulator of vesicular trafficking processes, but there is limited information about the guanine nucleotide-exchange factors and GTPase-activating proteins (GAPs) that regulate its GTP-GDP cycle. We observed that in the presence of Mg2+ (2.5 mM), TBC1D9B interacted via its Tre2-Bub2-Cdc16 (TBC) domain with Rab11a, Rab11b, and Rab4a in a nucleotide-dependent manner. However, only Rab11a was a substrate for TBC1D9B-stimulated GTP hydrolysis. At limiting Mg2+ concentrations (<0.5 mM), Rab8a was an additional substrate for this GAP. In polarized Madin–Darby canine kidney cells, endogenous TBC1D9B colocalized with Rab11a-positive recycling endosomes but less so with EEA1-positive early endosomes, transferrin-positive recycling endosomes, or late endosomes. Overexpression of TBC1D9B, but not an inactive mutant, decreased the rate of basolateral-to-apical IgA transcytosis—a Rab11a-dependent pathway—and shRNA-mediated depletion of TBC1D9B increased the rate of this process. In contrast, TBC1D9B had no effect on two Rab11a-independent pathways—basolateral recycling of the transferrin receptor or degradation of the epidermal growth factor receptor. Finally, expression of TBC1D9B decreased the amount of active Rab11a in the cell and concomitantly disrupted the interaction between Rab11a and its effector, Sec15A. We conclude that TBC1D9B is a Rab11a GAP that regulates basolateral-to-apical transcytosis in polarized MDCK cells.