Role of the Guanine Nucleotide Exchange Factor Ost in Negative Regulation of Receptor Endocytosis by the Small GTPase Rac1*
Role of the Guanine Nucleotide Exchange Factor Ost in Negative Regulation of Receptor Endocytosis by the Small GTPase Rac1*
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DOI：
10.1074/jbc.m700950200
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发表时间：
2007-08
影响因子：
4.8
通讯作者：
Katsuaki Ieguchi;Shuji Ueda;T. Kataoka;T. Satoh
中科院分区：
文献类型：
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作者：
Katsuaki Ieguchi;Shuji Ueda;T. Kataoka;T. Satoh
GTP酶的Rho家族参与细胞内囊泡运输的调节。在这里，我们研究了由Rho家族蛋白Rac 1介导的网格蛋白介导的细胞表面受体内吞的负调控机制。与以前的报道相反，只有Rac 1的激活突变体，而不是其他Rho家族成员，包括RhoA和Cdc 42，抑制转铁蛋白受体的内化。另一方面，通过RNA干扰下调Rac 1表达导致受体内化增强，表明内源性Rac 1实际上起负调节剂的作用。我们确定了一个鸟嘌呤核苷酸交换因子剪接变异体指定Ost-III，其中包含一个独特的C-末端区域，包括Src同源3域，作为调节Rac 1参与抑制受体内吞作用。相比之下，其他剪接变体Ost-I和Ost-II对受体内吞作用几乎没有影响。我们还研究了synaptojanin 2的亚细胞定位，synaptojanin 2是一种假定的Rac 1效应子，参与受体内吞的负调控。每个Ost剪接变体诱导不同的亚细胞定位的synaptojanin 2，这取决于Rac 1激活。此外，我们分离出γ-氨基丁酸A型受体相关蛋白（GABARAP）作为与Ost-III的C-末端区域结合的蛋白质。当异位表达时，GABARAP与Ost-III共定位，并有效地抑制Ost-III依赖的Rac 1激活和抑制受体内吞作用。GABARAP的脂质修饰对于抑制Ost-III是必要的。这些结果进行了讨论，在亚细胞区域的Rac 1依赖的信号通路，负调节网格蛋白介导的内吞作用的具体规定。
The Rho family of GTPases has been implicated in the regulation of intracellular vesicle trafficking. Here, we investigated the mechanism underlying the negative regulation of clathrin-mediated endocytosis of cell surface receptors mediated by the Rho family protein Rac1. Contrary to previous reports, only the activated mutant of Rac1, but not other Rho family members including RhoA and Cdc42, suppressed internalization of the transferrin receptor. On the other hand, down-regulation of Rac1 expression by RNA interference resulted in enhanced receptor internalization, suggesting that endogenous Rac1 in fact functions as a negative regulator. We identified a guanine nucleotide exchange factor splice variant designated Ost-III, which contains a unique C-terminal region including an Src homology 3 domain, as a regulator of Rac1 involved in the inhibition of receptor endocytosis. In contrast, other splice variants Ost-I and Ost-II exerted virtually no effect on receptor endocytosis. We also examined subcellular localization of synaptojanin 2, a putative Rac1 effector implicated in negative regulation of receptor endocytosis. Each Ost splice variant induced distinct subcellular localization of synaptojanin 2, depending on Rac1 activation. Furthermore, we isolated γ-aminobutyric acid type A receptor-associated protein (GABARAP) as a protein that binds to the C-terminal region of Ost-III. When ectopically expressed, GABARAP was co-localized with Ost-III and potently suppressed the Ost-III-dependent Rac1 activation and the inhibition of receptor endocytosis. Lipid modification of GABARAP was necessary for the suppression of Ost-III. These results are discussed in terms of subcellular region-specific regulation of the Rac1-dependent signaling pathway that negatively regulates clathrin-mediated endocytosis.