cAMP-dependent protein kinase phosphorylation of EVL, a mena/VASP relative, regulates its interaction with actin and SH3 domains
cAMP-dependent protein kinase phosphorylation of EVL, a mena/VASP relative, regulates its interaction with actin and SH3 domains
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DOI：
10.1074/jbc.m006274200
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发表时间：
2000-11-17
影响因子：
4.8
通讯作者：
Gertler, FB
中科院分区：
文献类型：
--
作者：
Lambrechts, A;Kwiatkowski, AV;Gertler, FB
Ena/Vasp家族的蛋白参与了需要动态肌动蛋白重塑的过程，如轴突引导和血小板激活。在这项工作中，我们探索了一些可能部分通过EVL(Ena/Vasp样蛋白)调节肌动蛋白动态的途径。EVL和EVL-I两种亚型在胸腺和脾的造血细胞中高表达。在CDS激活的T细胞中，EVL存在于富含F-肌动蛋白的斑块和激活侧T细胞上形成的微刺的末端。与其他家族成员一样，EVL在成纤维细胞中表达时，定位于局灶性粘连和板脂前缘。EVL是cAMP依赖的蛋白激酶的底物，这种磷酸化调节EVL与其配体之间的几个相互作用。与Vasp不同，EVL在生理条件下使肌动蛋白聚合成核，而EVL和Vasp的磷酸化降低了它们的成核活性。EVL直接结合到Abl、Lyn和NSRC SH3结构域；FE65 WW结构域；以及Profilin，可能是通过其富含脯氨酸的核心。EVL的cAMP依赖的蛋白激酶磷酸化取消了Ab1和NSRCSH3结构域的结合，但不能与Profilin或其他SH3结构域结合。我们发现两个Profilin二聚体在EVL的多聚脯氨酸序列上有很强的协同结合，此外，Profilin与SH3结构域竞争结合到部分重叠的结合部位。这些数据表明，EVL的功能可以通过与多个配体的相互作用和环核苷酸依赖蛋白激酶的磷酸化以一种复杂的方式进行调节。
Proteins of the Ena/VASP family are implicated in processes that require dynamic actin remodeling such as axon guidance and platelet activation. In this work, we explored some of the pathways that likely regulate actin dynamics in part via EVL (Ena/VASP-like protein). Two isoforms, EVL and EVL-I, were highly expressed in hematopoietic cells of thymus and spleen. In CDS-activated T-cells, EVL was found in F-actin-rich patches and at the distal tips of the microspikes that formed on the activated side of the T-cells. Like the other family members, EVL localized to focal adhesions and the leading edge of lamellipodia when expressed in fibroblasts. EVL was a substrate for the cAMP-dependent protein kinase, and this phosphorylation regulated several of the interactions between EVL and its ligands. Unlike VASP, EVL nucleated actin polymerization under physiological conditions, whereas phosphorylation of both EVL and VASP decreased their nucleating activity. EVL bound directly to the Abl, Lyn, and nSrc SH3 domains; the FE65 WW domain; and profilin, likely via its proline-rich core. Binding of Abl and nSrc SH3 domains, but not profilin or other SH3 domains, was abolished by cAMP-dependent protein kinase phosphorylation of EVL. We show strong cooperative binding of two profilin dimers on the polyproline sequence of EVL, Additionally, profilin competed with the SH3 domains for binding to partially overlapping binding sites. These data suggest that the function of EVL could be modulated in a complex manner by its interactions with multiple ligands and through phosphorylation by cyclic nucleotide dependent kinases.