Noncore components of the fission yeast γ-tubulin complex
Noncore components of the fission yeast γ-tubulin complex
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DOI：
10.1091/mbc.e05-11-1009
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发表时间：
2006-12-01
影响因子：
3.3
通讯作者：
Sawin, Kenneth E.
中科院分区：
文献类型：
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作者：
Anders, Andreas;Lourenco, Paula C. C.;Sawin, Kenneth E.
对于真核生物γ-微管蛋白复合物（γ -Tuc）各个组分在体内的功能了解相对较少。我们在筛选影响微管组织的裂殖酵母突变体时，鉴定出了三个基因，gfh1 +、mod21 +和mod22 +。gfh1 +是一种先前已被描述的γ -TuC蛋白，与人的γ -TuC亚基GCP4有微弱的相似性，而mod21 +是新发现的，与人的γ -TuC亚基GCP5有微弱的相似性。我们表明mod21p是一种真正的γ -TuC蛋白，并且像gfh1Δ突变体一样，mod21Δ突变体是可存活的。我们发现gfh1Δ和mod21Δ突变体在体内从所有类型的微管组织中心（MTOC）进行的微管成核在性质上是正常的，但从间期MTOC进行的成核在数量上有所减少，并且mod22 +中的突变会加剧这种情况。同时缺失gfh1p、mod21p和alp16p（第三种非必需的γ -TuC蛋白）不会导致累加性缺陷，这表明这三种蛋白都对单一功能有贡献。免疫共沉淀实验表明，gfh1p和alp16p相互依赖以与一个小的“核心”γ -TuC结合，而mod21p的结合更偏向外围，并且gfh1p和mod21p可能独立于小的γ -TuC形成一个亚复合物。有趣的是，蔗糖梯度分析表明裂殖酵母中γ -TuC的主要形式可能是一个小复合物。我们提出gfh1p、mod21p和alp16作为裂殖酵母γ -TuC的兼性“非核心”组分，增强其微管成核能力。
Relatively little is known about the in vivo function of individual components of the eukaryotic gamma-tubulin complex (gamma-Tuc). We identified three genes, gfh1+, mod21+, and mod22+, in a screen for fission yeast mutants affecting microtubule organization. gfh1+ is a previously characterized gamma-TuC protein weakly similar to human gamma-TuC subunit GCP4, whereas mod21+ is novel and shows weak similarity to human gamma-TuC subunit GCP5. We show that mod21p is a bona fide y-TuC protein and that, like gfh1 Delta mutants, mod21 Delta mutants are viable. We find that gfh1 Delta and mod21A mutants have qualitatively normal microtubule nucleation from all types of microtubule-organizing centers (MTOCs) in vivo but quantitatively reduced nucleation from interphase MTOCs, and this is exacerbated by mutations in mod22+. Simultaneous deletion of gfh1p, mod21p, and alp16p, a third nonessential gamma-TuC protein, does not lead to additive defects, suggesting that all three proteins contribute to a single function. Coimmunoprecipitation experiments suggest that gfh1p and alp16p are codependent for association with a small "core" gamma-TuC, whereas mod21p is more peripherally associated, and that gfh1p and mod21p may form a subcomplex independently of the small gamma-TuC. Interestingly, sucrose gradient analysis suggests that the major form of the gamma-TuC in fission yeast may be a small complex. We propose that gfh1p, mod21p, and alp16 act as facultative "noncore" components of the fission yeast gamma-TuC and enhance its microtubule-nucleating ability.