EpsinR: an ENTH domain-containing protein that interacts with AP-1

被引:172
作者
Hirst, J
Motley, A
Harasaki, K
Chew, SYP
Robinson, MS [1 ]
机构
[1] Univ Cambridge, Inst Med Res, Dept Clin Biochem, Cambridge CB2 2XY, England
[2] MRC, Mol Biol Lab, Cambridge CB2 2QH, England
基金
英国惠康基金;
关键词
D O I
10.1091/mbc.E02-09-0552
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
We have used GST pulldowns from A431 cell cytosol to identify three new binding partners for the gamma-adaptin appendage: Snx9, ARF GAP1, and a novel ENTH domain-containing protein, epsinR. EpsinR is a highly conserved protein that colocalizes with AP-1 and is enriched in purified clathrin-coated vesicles. However, it does not require AP-1 to get onto membranes and remains membrane-associated in AP-1-deficient cells. Moreover, although epsinR binds AP-1 via its COOH-terminal domain, its NH2-terminal ENTH domain can be independently recruited onto membranes, both in vivo and in vitro. Brefeldin A causes epsinR to redistribute into the cytosol, and recruitment of the ENTH domain requires GTPgammaS, indicating that membrane association is ARF dependent. In protein-lipid overlay assays, the epsinR ENTH domain binds to PtdIns(4)P, suggesting a possible mechanism for ARF-dependent recruitment onto TGN membranes. When epsinR is depleted from cells by RNAi, cathepsin D is still correctly processed intracellularly to the mature form. This indicates that although epsinR is likely to be an important component of the AP-1 network, it is not necessary for the sorting of lysosomal enzymes.
引用
收藏
页码:625 / 641
页数:17
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