Arabidopsis VILLIN1 and VILLIN3 Have Overlapping and Distinct Activities in Actin Bundle Formation and Turnover
Arabidopsis VILLIN1 and VILLIN3 Have Overlapping and Distinct Activities in Actin Bundle Formation and Turnover
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DOI：
10.1105/tpc.110.076240
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发表时间：
2010-08-01
期刊：
影响因子：
11.6
通讯作者：
Staiger, Christopher J.
中科院分区：
文献类型：
--
作者：
Khurana, Parul;Henty, Jessica L.;Staiger, Christopher J.
肌动蛋白丝束是高级细胞骨架结构，对于维持细胞结构和细胞扩张至关重要。它们是由单个肌动蛋白丝通过捆绑蛋白质（如fimbrins、LIM和villins）的作用产生的。然而，动态束形成和营业额的分子机制在很大程度上是未知的。Villins属于villin/gelsolin/fragmin超家族，并且在拟南芥中包含至少五种同功变异体。绒毛蛋白同种变体的不同组合在各种组织和细胞中共表达。目前尚不清楚这些同种变体是否共同发挥作用并发挥冗余作用，或者它们是否具有独特的活性。VILLIN 1（VLN 1）是一种简单的粘附捆绑蛋白，对Ca 2+不敏感。基于系统发育分析和保守的Ca 2+结合位点，我们预测VLN 3是一个Ca 2+调节的绒毛能够切断肌动蛋白丝，有助于束营业额。两种异构体的集束活性直接观察到延时成像和全内反射荧光（TIRF）显微镜在体外，和机制模拟的“捕捉和拉链”的行动在体内观察到。使用时间推移TIRF显微镜，我们观察和量化的切断个别肌动蛋白丝VLN 3在生理钙浓度。此外，VLN 3可以切断肌动蛋白丝束在VLN 1的存在下，当钙离子升高到微摩尔水平。总的来说，这些结果表明，两个绒毛蛋白同种变体具有重叠和不同的活动。
Actin filament bundles are higher-order cytoskeletal structures that are crucial for the maintenance of cellular architecture and cell expansion. They are generated from individual actin filaments by the actions of bundling proteins like fimbrins, LIMs, and villins. However, the molecular mechanisms of dynamic bundle formation and turnover are largely unknown. Villins belong to the villin/gelsolin/fragmin superfamily and comprise at least five isovariants in Arabidopsis thaliana. Different combinations of villin isovariants are coexpressed in various tissues and cells. It is not clear whether these isovariants function together and act redundantly or whether they have unique activities. VILLIN1 (VLN1) is a simple filament-bundling protein and is Ca2+ insensitive. Based on phylogenetic analyses and conservation of Ca2+ binding sites, we predict that VLN3 is a Ca2+-regulated villin capable of severing actin filaments and contributing to bundle turnover. The bundling activity of both isovariants was observed directly with time-lapse imaging and total internal reflection fluorescence (TIRF) microscopy in vitro, and the mechanism mimics the "catch and zipper" action observed in vivo. Using time-lapse TIRF microscopy, we observed and quantified the severing of individual actin filaments by VLN3 at physiological calcium concentrations. Moreover, VLN3 can sever actin filament bundles in the presence of VLN1 when calcium is elevated to micromolar levels. Collectively, these results demonstrate that two villin isovariants have overlapping and distinct activities.