The cytoplasmic tail of L-selectin interacts with the adaptor-protein complex AP-1 subunit μ1A via a novel basic binding motif
The cytoplasmic tail of L-selectin interacts with the adaptor-protein complex AP-1 subunit μ1A via a novel basic binding motif
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L-选择素的胞质尾部通过新型基本结合基序与接头蛋白复合物 AP-1 亚基 μ1A 相互作用
DOI：
10.1074/jbc.m116.768598
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发表时间：
2017
期刊：
影响因子：
--
通讯作者：
P. Schu
中科院分区：
文献类型：
--
作者：
K. Dib;I. Tikhonova;A. Ivetic;P. Schu
L-选择素调节白细胞粘附并沿内皮沿着。与L-选择素的胞质尾区结合的蛋白质调节L-选择素功能。我们使用L-选择素胞质尾肽下拉分析结合高灵敏度液相色谱/质谱法，以确定新的L-选择素尾结合蛋白。将L-选择素尾部与佛波醇12-肉豆蔻酸酯13-乙酸酯刺激的Raw 264.7巨噬细胞的细胞提取物孵育，导致网格蛋白包被的囊泡AP-1复合物的μ 1 A结合。此外，全长GST-μ1A和GST-μ1A C-末端结构域，而不是GST-μ1A N-末端结构域，结合L-选择素尾肽，并且L-选择素的细胞内池与AP-1共定位在trans-Golgi网络中。我们鉴定了一种新的碱性蛋白基序，其由位于L-选择素尾近膜区的三个二元残基（356 RR 357、359 KK 360和362 KK 363）组成，以及位于L-选择素尾膜远端的一对天冬氨酸残基（369 DD 370）组成，参与μ1A结合。用PMA刺激Raw 264.7巨噬细胞增加了与抗L-选择素免疫沉淀物相关的μ1A的量。然而，全长GST-μ1A不与磷酸化L-选择素尾或磷酸化模拟S364 D L-选择素尾结合。因此，我们提出μ1A的磷酸化是与L-选择素尾部相互作用所必需的，并且L-选择素尾部磷酸化可能在体内调节这种相互作用。使用L-选择素尾与μ1A的分子对接来鉴定结合L-选择素尾的μ1A表面结构域，并解释L-选择素尾的磷酸化如何消除μ1A相互作用。我们的研究结果表明，L-选择素是由AP-1复合物组成性运输，导致形成一个trans-Golgi网络储备池和磷酸化的L-选择素尾部块AP-1依赖的逆行运输L-选择素。
L-selectin regulates leukocyte adhesion and rolling along the endothelium. Proteins binding to the cytoplasmic tail of L-selectin regulate L-selectin functions. We used L-selectin cytoplasmic tail peptide pulldown assays combined with high sensitivity liquid chromatography/mass spectrometry to identify novel L-selectin tail-binding proteins. Incubation of the L-selectin tail with cell extracts from phorbol 12-myristate 13-acetate-stimulated Raw 264.7 macrophages resulted in the binding of μ1A of the clathrin-coated vesicle AP-1 complex. Furthermore, full-length GST-μ1A and the GST-μ1A C-terminal domain, but not the GST-μ1A N-terminal domain, bind to L-selectin tail peptide, and the intracellular pool of L-selectin colocalizes with AP-1 at the trans-Golgi network. We identified a novel basic protein motif consisting of a cluster of three dibasic residues (356RR357, 359KK360, and 362KK363) in the membrane-proximal domain of the L-selectin tail as well as a doublet of aspartic acid residues (369DD370) in the membrane-distal end of the L-selectin tail involved in μ1A binding. Stimulation of Raw 264.7 macrophages with PMA augmented the amount of μ1A associated with anti-L-selectin immunoprecipitates. However, full-length GST-μ1A did not bind to the phospho-L-selectin tail or phospho-mimetic S364D L-selectin tail. Accordingly, we propose that phosphorylation of μ1A is required for interaction with the L-selectin tail and that L-selectin tail phosphorylation may regulate this interaction in vivo. Molecular docking of the L-selectin tail to μ1A was used to identify the μ1A surface domain binding the L-selectin tail and to explain how phosphorylation of the L-selectin tail abrogates μ1A interaction. Our findings indicate that L-selectin is transported constitutively by the AP-1 complex, leading to the formation of a trans-Golgi network reserve pool and that phosphorylation of the L-selectin tail blocks AP-1-dependent retrograde transport of L-selectin.
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DOI：
10.1073/pnas.0406102101
发表时间：
2004-09-28
影响因子：
11.1
作者：
Heldwein, EE;Macia, E;Harrison, SC
通讯作者：
Harrison, SC
影响因子：
4.4
作者：
James D. Griffin;O. Spertini;T. Ernst;M. Belvin;H. Levine;Y. Kanakura;T. Tedder
通讯作者：
James D. Griffin;O. Spertini;T. Ernst;M. Belvin;H. Levine;Y. Kanakura;T. Tedder
影响因子：
5.6
作者：
Deng, Wei;Cho, Sungyun;Li, Renhao
通讯作者：
Li, Renhao
影响因子：
32.4
作者：
ARBONES, ML;ORD, DC;TEDDER, TF
通讯作者：
TEDDER, TF