Capping protein regulatory cycle driven by CARMIL and V-1 may promote actin network assembly at protruding edges
Capping protein regulatory cycle driven by CARMIL and V-1 may promote actin network assembly at protruding edges
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DOI：
10.1073/pnas.1313738111
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发表时间：
2014-05-13
影响因子：
11.1
通讯作者：
Hammer, John A.
中科院分区：
文献类型：
--
作者：
Fujiwara, Ikuko;Remmert, Kirsten;Hammer, John A.
虽然加帽蛋白（CP）终止肌动蛋白丝的伸长，它促进Arp 2/3依赖的肌动蛋白网络组装和加速肌动蛋白为基础的运动在体外和体内。在体外，加帽蛋白Arp 2/3肌球蛋白I接头（CARMIL）拮抗CP通过降低其对倒刺末端的亲和力和通过使CP加帽的细丝脱帽，而蛋白V-1/肌营养因子将CP隔离在非活性复合物中。先前的工作表明，CARMIL可以很容易地从CP：V-1复合物中回收CP，从而将无活性的CP转化为对倒刺末端具有中等亲和力的形式。在这里，我们进一步阐明了这种交换反应的机制，我们证明了CP：CARMIL复合物通过复合物交换产生的，通过快速与倒刺末端结合和解离来减缓倒刺末端伸长的速率。重要的是，这里确定的V-1和CP的细胞浓度表明，大多数CP在体内稳态时被V-1螯合。最后，我们表明，CARMIL被招募到质膜，只有在细胞边缘进行积极的突起。假设CARMIL仅在该位置活跃，我们的数据表明，大量自由扩散的非活性CP（CP：V-1）通过CARMIL驱动的复合物交换，在质膜的突出边缘形成弱帽复合物（CP：CARMIL）。因此，在体内，CARMIL应该促进Arp 2/3依赖的肌动蛋白网络组装的前沿，促进倒钩端帽那里。
Although capping protein (CP) terminates actin filament elongation, it promotes Arp2/3-dependent actin network assembly and accelerates actin-based motility both in vitro and in vivo. In vitro, capping protein Arp2/3 myosin I linker (CARMIL) antagonizes CP by reducing its affinity for the barbed end and by uncapping CP-capped filaments, whereas the protein V-1/myotrophin sequesters CP in an inactive complex. Previous work showed that CARMIL can readily retrieve CP from the CP:V-1 complex, thereby converting inactive CP into a version with moderate affinity for the barbed end. Here we further clarify the mechanism of this exchange reaction, and we demonstrate that the CP:CARMIL complex created by complex exchange slows the rate of barbed-end elongation by rapidly associating with, and dissociating from, the barbed end. Importantly, the cellular concentrations of V-1 and CP determined here argue that most CP is sequestered by V-1 at steady state in vivo. Finally, we show that CARMIL is recruited to the plasma membrane and only at cell edges undergoing active protrusion. Assuming that CARMIL is active only at this location, our data argue that a large pool of freely diffusing, inactive CP (CP:V-1) feeds, via CARMIL-driven complex exchange, the formation of weak-capping complexes (CP:CARMIL) at the plasma membrane of protruding edges. In vivo, therefore, CARMIL should promote Arp2/3-dependent actin network assembly at the leading edge by promoting barbed-end capping there.