Free Brick1 is a trimeric precursor in the assembly of a functional wave complex.
Free Brick1 is a trimeric precursor in the assembly of a functional wave complex.
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DOI：
10.1371/journal.pone.0002462
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发表时间：
2008-06-18
期刊：
影响因子：
3.7
通讯作者：
Gautreau A
中科院分区：
文献类型：
--
作者：
Derivery E;Fink J;Martin D;Houdusse A;Piel M;Stradal TE;Louvard D;Gautreau A
Wave复合体激活Arp2/3复合体，诱导片状脂膜和膜皱纹中的肌动蛋白聚合。Wave复合体由5个亚基组成，其中最小的Brick1/Hspc300(Brk1)是最不具特征的。我们以前报道过，与其他亚基不同，Brk1也是以自由形式存在的。在这里，我们报告这种自由形式的Brk1是由同源三聚体组成的。利用一种新的方法，将纯化的游离Brk1电穿孔到HeLa细胞中，我们能够跟踪其在细胞中的生化命运，并表明游离Brk1被整合到Wave复合体中。重要的是，一旦蛋白质合成受到抑制，游离的Brk1掺入Wave复合体就会被阻止，而掺入的Brk1被发现优先与新合成的亚基结合。Brk1耗竭的HeLa细胞和NAP1、Wave2或ARPC2耗竭的细胞一样被发现起泡，这表明Brk1耗竭的细胞的这种起泡表型是由于Wave复合体功能的损害，而不是游离Brk1的特定功能。Brk1耗竭细胞的气泡是在正常情况下产生片状脂肪和膜皱纹的部位发出的。在Brk1缺失的细胞中，游离Brk1的电穿孔足以恢复Wave复合体的组装并挽救起泡表型。综上所述，这些结果证实了Brk1的自由形式是组装功能波复合体的必要前体。
The Wave complex activates the Arp2/3 complex, inducing actin polymerization in lamellipodia and membrane ruffles. The Wave complex is composed of five subunits, the smallest of which, Brick1/Hspc300 (Brk1), is the least characterized. We previously reported that, unlike the other subunits, Brk1 also exists as a free form. Here we report that this free form of Brk1 is composed of homotrimers. Using a novel assay in which purified free Brk1 is electroporated into HeLa cells, we were able to follow its biochemical fate in cells and to show that free Brk1 becomes incorporated into the Wave complex. Importantly, incorporation of free Brk1 into the Wave complex was blocked upon inhibition of protein synthesis and incorporated Brk1 was found to associate preferentially with neosynthesized subunits. Brk1 depleted HeLa cells were found to bleb, as were Nap1, Wave2 or ARPC2 depleted cells, suggesting that this blebbing phenotype of Brk1 depleted cells is due to an impairment of the Wave complex function rather than a specific function of free Brk1. Blebs of Brk1 depleted cells were emitted at sites where lamellipodia and membrane ruffles were normally emitted. In Brk1 depleted cells, the electroporation of free Brk1 was sufficient to restore Wave complex assembly and to rescue the blebbing phenotype. Together these results establish that the free form of Brk1 is an essential precursor in the assembly of a functional Wave complex.
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影响因子：
7.8
作者：
Raucher, D;Sheetz, M P
通讯作者：
Sheetz, M P
DOI：
10.1073/pnas.0502390102
发表时间：
2005-09-27
影响因子：
11.1
作者：
Kammerer, RA;Kostrewa, D;Steinmetz, MO
通讯作者：
Steinmetz, MO
影响因子：
5.7
作者：
Iwaya, Keiichi;Oikawa, Kosuke;Mukai, Kiyoshi
通讯作者：
Mukai, Kiyoshi
影响因子：
7.5
作者：
Iwaya, Keiichi;Norio, Kohno;Mukai, Kiyoshi
通讯作者：
Mukai, Kiyoshi
影响因子：
64.8
作者：
Charras, GT;Yarrow, JC;Mitchison, TJ
通讯作者：
Mitchison, TJ