Dab2 regulates clathrin assembly and cell spreading
Dab2 regulates clathrin assembly and cell spreading
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DOI：
10.1042/bj20081288
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发表时间：
2009-03-15
影响因子：
4.1
通讯作者：
Ehrlich, Marcelo
中科院分区：
文献类型：
--
作者：
Chetrit, David;Ziv, Naomi;Ehrlich, Marcelo
招募网格蛋白的膜和它的组件成包被的坑的结果，从它的相互作用与内吞衔接子和其他调节蛋白的背景下，一个特定的脂质微环境。Dab 2（disabled 2）是一种有丝分裂磷蛋白，是网格蛋白介导的内吞作用的单体接头。在本研究中，我们采用GFP（绿色荧光蛋白）融合构建的不同亚型和突变体的大鼠Dab 2和其特征的大小，分布和动态的网格蛋白组件的影响。在COS 7细胞中Dab 2的p82同种型的表达水平的增强诱导了质膜处的扩大的网格蛋白组装体。p82-网格蛋白组装体是动态结构，其中p82和网格蛋白在膜结合亚群和胞质亚群之间进行活性交换，所述组装体浓缩额外的内吞蛋白，例如AP 2（衔接蛋白2）和epsin。p82诱导扩大网格蛋白组件的能力取决于其功能性PTB结构域（磷酸酪氨酸结合结构域）的存在，油与网格蛋白和磷脂的结合，以及在PTB结构域之前的新鉴定的和进化上保守的聚赖氨酸延伸。Dab 2需要这些相同的分子特征来增强COS 7细胞在纤连蛋白上的铺展。在HeLa细胞和HBL细胞（人乳腺上皮细胞）中证实了Dab 2的p82同种型增强细胞铺展的能力。表达GFP-p82并铺在纤连蛋白上的COS 7细胞将β 1整联蛋白浓缩成网格蛋白-p82组装体。此外，在细胞铺展过程中，p82-网格蛋白组件集中在最初的细胞-基质接触的位点，并且不存在于强烈的膜皱褶区域。我们提出了Dab 2和网格蛋白在整合素介导的细胞扩散中的作用。
The recruitment of clathrin to the membrane and its assembly into coated pits results from its interaction with endocytic adaptors and other regulatory proteins in the context of a specific lipid microenvironment. Dab2 (disabled 2) is a mitotic phosphoprotein and a monomeric adaptor for clathrin-mediated endocytosis. In the present study, we employed GFP (green fluorescent protein) fusion constructs of different isoforms and mutants of rat Dab2 and characterized their effect on the size, distribution and dynamics of clathrin assemblies. Enhanced levels of expression of the p82 isoform of Dab2 in COS7 cells induced enlarged clathrin assemblies at the plasma membrane. p82-clathrin assemblies, which concentrate additional endocytic proteins, Such as AP2 (adaptor protein 2) and epsin, are dynamic structures in which both p82 and clathrin exchange actively between the membrane-bound and cytosolic sub-populations. The ability of p82 to induce enlarged clathrin assemblies is dependent on the presence of it functional PTB domain (phosphotyrosine-binding domain), oil binding to clathrin and phospholipids, and on a newly identified and evolutionarily conserved poly-lysine stretch which precedes the PTB domain. These same molecular features are required for Dab2 to enhance the spreading of COS7 cells on fibronectin. The ability of the p82 isoform of Dab2 to enhance cell spreading was confirmed ill both HeLa cells and HBL cells (human breast epithelial cells). COS7 cells expressing GFP-p82 and plated on to fibronectin concentrate the beta 1 integrin into clathrin-p82 assemblies. Furthermore, during cell spreading, p82-clathrin assemblies concentrate at the site of the initial cell-matrix contact and are absent from regions of intense membrane ruffling. We propose a role for Dab2 and clathrin in integrin-mediated cell spreading.