Characterization of the minimum domain required for targeting budding yeast myosin II to the site of cell division.
Characterization of the minimum domain required for targeting budding yeast myosin II to the site of cell division.
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靶向细胞分裂部位靶向萌芽酵母肌球蛋白II所需的最小结构域的表征。
DOI：
10.1186/1741-7007-4-19
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发表时间：
2006-06-26
期刊：
影响因子：
5.4
通讯作者：
Li, Rong
中科院分区：
文献类型：
--
作者：
Lister, Ida Mb;Tolliday, Nicola J.;Li, Rong
除植物外，所有真核生物都使用肌动球蛋白环在有丝分裂和减数分裂过程中的细胞分裂部位（卵裂沟）产生收缩力。位于主要成分之一肌球蛋白 II 的 C 端或尾部区域的结构和丝形成能力对于胞质分裂过程中将分子定位到收缩环 (CR) 非常重要。然而，人们对肌球蛋白 II 如何被募集到细胞分裂位点以及这种募集与肌球蛋白丝组装有何关系仍知之甚少。参与胞质分裂的成分（包括 CR 的成分）在物种之间具有显着的保守性，因此可以在胞质分裂的研究中使用易于遗传操作的生物体，例如芽殖酵母（酿酒酵母）。芽殖酵母有一种肌球蛋白 II 蛋白，称为 Myo1。与大多数其他 II 类肌球蛋白不同，Myo1 的尾部有一个不规则的卷曲线圈。在本报告中，我们使用分子遗传学、生物化学和活细胞成像来表征芽殖酵母 Myo1 的最小定位域 (MLD)。我们证明 MLD 是 Myo1 尾部中心的一个小区域，它对于将 Myo1 定位到酵母芽颈（细胞分裂的预定位点）既是必要的也是充分的。从细菌或酵母中纯化的 MLD 的流体动力学测量表明，它很可能以三聚体形式存在。我们还研究了 MLD 内一个低盘绕线圈形成概率的小区域（我们称之为铰链区域）的重要性。铰链区域的去除可防止 CR 收缩。使用光漂白后的荧光恢复 (FRAP)，我们表明 GFP 标记的 MLD 比 GFP 标记的全长分子动态稍强，但比缺少铰链区的 GFP 标记的 Myo1 构建体动态较差。我们的结果定义了芽殖酵母肌球蛋白 II 定位的内在决定因素，并表明它是暂定 3 个单体的低聚物。我们认为这是最小的寡聚单元（而不是传统的肌球蛋白 II 二聚体），它允许以类似于全长分子的方式特异性组装到胞质分裂位点。柔性铰链区域也有助于 CR 结构的完整性和收缩性。
All eukaryotes with the exception of plants use an actomyosin ring to generate a constriction force at the site of cell division (cleavage furrow) during mitosis and meiosis. The structure and filament forming abilities located in the C-terminal or tail region of one of the main components, myosin II, are important for localising the molecule to the contractile ring (CR) during cytokinesis. However, it remains poorly understood how myosin II is recruited to the site of cell division and how this recruitment relates to myosin filament assembly. Significant conservation between species of the components involved in cytokinesis, including those of the CR, allows the use of easily genetically manipulated organisms, such as budding yeast (Saccharomyces cerevisiae), in the study of cytokinesis. Budding yeast has a single myosin II protein, named Myo1. Unlike most other class II myosins, the tail of Myo1 has an irregular coiled coil. In this report we use molecular genetics, biochemistry and live cell imaging to characterize the minimum localisation domain (MLD) of budding yeast Myo1. We show that the MLD is a small region in the centre of the tail of Myo1 and that it is both necessary and sufficient for localisation of Myo1 to the yeast bud neck, the pre-determined site of cell division. Hydrodynamic measurements of the MLD, purified from bacteria or yeast, show that it is likely to exist as a trimer. We also examine the importance of a small region of low coiled coil forming probability within the MLD, which we call the hinge region. Removal of the hinge region prevents contraction of the CR. Using fluorescence recovery after photobleaching (FRAP), we show that GFP-tagged MLD is slightly more dynamic than the GFP-tagged full length molecule but less dynamic than the GFP-tagged Myo1 construct lacking the hinge region. Our results define the intrinsic determinant for the localization of budding yeast myosin II and show it to be an oligomer of tentatively 3 monomers. We suggest that this is the minimum oligomeric unit (rather than the traditional myosin II dimer) that would allow specific assembly to the site of cytokinesis in a manner similar to the full length molecule. The flexible hinge region also contributes to CR structural integrity and contractility.
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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：
3.3
作者：
Mulvihill, DP;Barretto, C;Hyams, JS
通讯作者：
Hyams, JS
影响因子：
3.3
作者：
Sabry, JH;Moores, SL;Spudich, JA
通讯作者：
Spudich, JA
DOI：
10.1083/jcb.200402097
发表时间：
2004-06-07
期刊：
The Journal of cell biology
影响因子：
--
作者：
Motegi F;Mishra M;Balasubramanian MK;Mabuchi I
通讯作者：
Mabuchi I
影响因子：
3.3
作者：
Bezanilla, M;Pollard, TD
通讯作者：
Pollard, TD