Cytoplasmic nucleation and atypical branching nucleation generate endoplasmic microtubules in Physcomitrella patens.
Cytoplasmic nucleation and atypical branching nucleation generate endoplasmic microtubules in Physcomitrella patens.
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小立碗藓中的细胞质成核和非典型分支成核产生内质微管。
DOI：
10.1105/tpc.114.134817
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发表时间：
2015
期刊：
影响因子：
--
通讯作者：
Goshima,Gohta
中科院分区：
文献类型：
--
作者：
Nakaoka,Yuki;Kimura,Akatsuki;Tani,Tomomi;Goshima,Gohta
植物中微管（MT）生成的机制主要是使用皮质 MT 阵列进行研究，其中固定角度分支成核和剑蛋白依赖性 MT 切断占主导地位。然而，人们对内质中 MT 的产生知之甚少。在此，我们探讨了小立碗藓原质细胞内质MT产生的机制。我们开发了一种利用流动池和倾斜照明荧光显微镜的检测方法，可以对单个 MT 动态进行可视化和量化。 MT 断裂很少发生，并且剑蛋白的破坏不会严重影响 MT 的产生。观察到分支成核，但它表现出明显变化的分支角度，并且偶尔伴随有核MT的运输。当解聚的 MT 重新生长时，看似随机位置的细胞质成核最常见且占主导地位。在大多数成核位点检测到 MT 成核剂 γ-微管蛋白，在这些位点处以随机方向生成单个 MT。当 γ-微管蛋白被敲低时，在再生测定中 MT 的生成显着延迟。然而，在稳定状态下成核以正常频率发生，表明存在不依赖于γ-微管蛋白的备用机制。因此，这种细胞类型中的内质MT以不太有序的方式产生，显示出植物中更广泛的成核机制。
The mechanism underlying microtubule (MT) generation in plants has been primarily studied using the cortical MT array, in which fixed-angled branching nucleation and katanin-dependent MT severing predominate. However, little is known about MT generation in the endoplasm. Here, we explored the mechanism of endoplasmic MT generation in protonemal cells ofPhyscomitrella patens. We developed an assay that utilizes flow cell and oblique illumination fluorescence microscopy, which allowed visualization and quantification of individual MT dynamics. MT severing was infrequently observed, and disruption of katanin did not severely affect MT generation. Branching nucleation was observed, but it showed markedly variable branch angles and was occasionally accompanied by the transport of nucleated MTs. Cytoplasmic nucleation at seemingly random locations was most frequently observed and predominated when depolymerized MTs were regrown. The MT nucleator γ-tubulin was detected at the majority of the nucleation sites, at which a single MT was generated in random directions. When γ-tubulin was knocked down, MT generation was significantly delayed in the regrowth assay. However, nucleation occurred at a normal frequency in steady state, suggesting the presence of a γ-tubulin-independent backup mechanism. Thus, endoplasmic MTs in this cell type are generated in a less ordered manner, showing a broader spectrum of nucleation mechanisms in plants.
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