AtFim1 is an actin filament crosslinking protein from Arabidopsis thaliana
AtFim1 is an actin filament crosslinking protein from Arabidopsis thaliana
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DOI：
10.1046/j.1365-313x.2000.00907.x
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发表时间：
2000-12-01
期刊：
影响因子：
7.2
通讯作者：
McCurdy, DW
中科院分区：
文献类型：
--
作者：
Kovar, DR;Staiger, CJ;McCurdy, DW
ATFIM 1是拟南芥中广泛表达的基因，其编码一种推定的肌动蛋白亲和交联蛋白AtFim 1，属于肌动蛋白结合蛋白的fimplastin/plastin类。在这份报告中，我们已经使用细菌表达的AtFim 1和肌动蛋白分离玉米花粉证明AtFim 1作为一个肌动蛋白的免疫交联蛋白的功能。AtFim 1以钙非依赖性方式结合花粉肌动蛋白丝（F-actin），平均解离常数（Kd）为0.55 +/- 0.21 μ M，饱和时的化学计量比为1：4（mol AtFim 1：mol肌动蛋白单体）。AtFim 1也交联花粉F-肌动蛋白的钙依赖性机制，在植物肌动蛋白交联的人T-plastin，一个已知的钙敏感的肌动蛋白交联蛋白。当显微注射到高浓度的活紫露草雄蕊毛细胞，AtFim 1引起细胞质流和transvacuolar链动力学在2-4分钟内停止。使用“核置换试验”作为衡量的完整性肌动蛋白细胞骨架活雄蕊毛细胞，我们证明了AtFim 1保护肌动蛋白丝在这些细胞中Z。玉米profilin（ZmPRO 5）诱导的解聚，在剂量依赖性的方式。AtFim 1的明显能力，以保护肌动蛋白丝在体内从profilin介导的解聚证实了在体外沉降试验。我们的研究结果表明，AtFim 1是一种钙离子非依赖性的，肌动蛋白介导的交联蛋白，在活的植物细胞中与肌动蛋白细胞骨架相互作用。
ATFIM1 is a widely expressed gene in Arabidopsis thaliana that encodes a putative actin filament-crosslinking protein, AtFim1, belonging to the fimbrin/plastin class of actin-binding proteins. In this report we have used bacterially expressed AtFim1 and actin isolated from Zea mays pollen to demonstrate that AtFim1 functions as an actin filament-crosslinking protein. AtFim1 binds pollen actin filaments (F-actin) in a calcium-independent manner, with an average dissociation constant (K-d) of 0.55 +/- 0.21 muM and with a stoichiometry at saturation of 1:4 (mol AtFim1:mol actin monomer). AtFim1 also crosslinks pollen F-actin by a calcium-independent mechanism, in contrast to crosslinking of plant actin by human T-plastin, a known calcium-sensitive actin-crosslinking protein. When microinjected at high concentration into living Tradescantia virginiana stamen hair cells, AtFim1 caused cessation of both cytoplasmic streaming and transvacuolar strand dynamics within 2-4 min. Using the 'nuclear displacement assay' as a measure of the integrity of the actin cytoskeleton in living stamen hair cells, we demonstrated that AtFim1 protects actin filaments in these cells from Z. mays profilin (ZmPRO5)-induced depolymerization, in a dose-dependent manner. The apparent ability of AtFim1 to protect actin filaments in vivo from profilin-mediated depolymerization was confirmed by in vitro sedimentation assays. Our results indicate that AtFim1 is a calcium-independent, actin filament-crosslinking protein that interacts with the actin cytoskeleton in living plant cells.