Involvement of the endosomal autoantigen EEA1 in homotypic fusion of early endosomes
Involvement of the endosomal autoantigen EEA1 in homotypic fusion of early endosomes
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DOI：
10.1016/s0960-9822(07)00351-x
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发表时间：
1998-07-16
期刊：
影响因子：
9.2
通讯作者：
Clague, MJ
中科院分区：
文献类型：
--
作者：
Mills, IG;Jones, AT;Clague, MJ
在哺乳动物细胞中，早期内吞囊泡之间的融合已显示需要普遍存在的细胞内融合因子N乙基马来酰亚胺敏感因子（NSF）和α-SNAP，以及对早期内体特异的因子，小GTdR abB [1-3]。我们以前已经证明了对磷脂酰肌醇3-激酶的额外要求（PI 3-激酶）活性[4]，早期内体抗原1（EEA 1）的膜结合，早期内体的特异性标志物[5，6]，最近已被证明类似地依赖于PI 3-激酶活性[7]，因此我们假设它可能参与内体融合。我们提供的证据表明EEA 1在体外确定内体融合效率方面具有重要作用。当包括在体外测定中时，EEA 1的羧基末端结构域（残基1098-1411）和针对EEA 1的特异性抗体都抑制内体融合。此外，EEA 1的消耗，无论是从用盐洗涤的测定中使用的膜级分和使用EEA 1特异性抗体从胞质溶胶，导致抑制内体融合。EEA 1参与内体融合解释了内体融合测定对PI 3-激酶抑制剂的敏感性。
In mammalian cells, fusion between early endocytic vesicles has been shown to require the ubiquitous intracellular fusion factors N ethylmaleimide-sensitive factor (NSF) and a-SNAP, as well as a factor specific for early endosomes, the small GTPase RabB [1-3]. We have previously demonstrated an additional requirement for phosphatidylinositol 3-kinase (PI 3-kinase) activity [4], The membrane association of early endosomal antigen 1 (EEA1), a specific marker of early endosomes [5,6], has recently been shown to be similarly dependent on PI 3-kinase activity [7], and we therefore postulated that it might be involved in endosome fusion, Here, we present evidence that EEA1 has an important role in determining the efficiency of endosome fusion in vitro. Both the carboxy-terminal domain of EEA1 (residues 1098-1411) and specific antibodies against EEA1 inhibited endosome fusion when included in an in vitro assay. Furthermore, depletion of EEA1, both from the membrane fraction used in the assay by washing with salt and from the cytosol using an EEA1-specific antibody, resulted in inhibition of endosome fusion, The involvement of EEA1 in endosome fusion accounts for the sensitivity of the endosome fusion assay to inhibitors of PI 3-kinase.