Subcellular localization and possible function of actin, tropomyosin and actin-related protein 3 (Arp3) in the fission yeast Schizosaccharomyces pombe
Subcellular localization and possible function of actin, tropomyosin and actin-related protein 3 (Arp3) in the fission yeast Schizosaccharomyces pombe
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DOI：
10.1016/s0171-9335(98)80007-1
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发表时间：
1998-08-01
影响因子：
6.6
通讯作者：
Mabuchi, I
中科院分区：
文献类型：
--
作者：
Arai, R;Nakano, K;Mabuchi, I
我们研究了亚细胞定位和相互作用的肌动蛋白和两个肌动蛋白cytokeratin相关蛋白，Cdc 8原肌球蛋白和肌动蛋白相关蛋白3，Arp 3，在裂殖酵母裂殖酵母，使用特异性抗体和基因破坏。肌动蛋白定位于内侧微丝环在该地区的隔膜胞质分裂和皮质补丁免疫电镜。F-actin电缆检测整个细胞周期的荧光染色与Bodipy-phallacidin。在其形成过程中，电缆通常连接到补片和内侧环。原肌球蛋白定位于内侧环和线缆。它也分布在细胞中的补丁，虽然与F-肌动蛋白的共定位是不常见的。在cdc 8（ts）突变体细胞中，虽然检测到F-肌动蛋白斑块，但未观察到F-肌动蛋白索，细胞极性保持不变。这些观察结果表明，F-肌动蛋白电缆可能参与形成的内侧环，原肌球蛋白起着重要的作用，在组织的环和电缆，但不参与F-肌动蛋白补丁的形成或维持细胞polarity.Binding的Arp 3肌动蛋白的免疫沉淀，以及DNase I柱层析。Arp 3似乎与细胞提取物中的几种蛋白质形成复合物，如先前对其他生物体的报道。与先前的报告相反（McCollum等人，EMBO J. 15，6438-6446，1996），发现Arp 3集中在从后期早期到后期胞质分裂的中间区域。Arp 3基因破坏后，F-肌动蛋白补丁在整个细胞中离域，细胞没有经历极化生长，表明Arp 3影响肌动蛋白补丁在细胞中的适当定位，从而控制细胞的极化生长。
We investigated subcellular localizations and interactions of actin and two actin cytoskeleton-related proteins, Cdc8 tropomyosin and actin-related protein 3, Arp3, in the fission yeast Schizosaccharomyces pombe, using specific antibodies and by gene disruption. Actin was localized to the medial microfilamentous ring in the region of the septum during cytokinesis and to cortical patches by immunoelectron microscopy. F-actin cables were detected throughout the cell cycle by fluorescent staining with Bodipy-phallacidin. Cables were often linked to the patches and to the medial ring during its formation. Tropomyosin was localized to the medial ring and the cables. It was also distributed in the cell as patches, although co-localization with F-actin was not frequent. In cdc8(ts) mutant cells, F-actin cables were not observed although the F-actin patches were detected and cell polarity was maintained. These observations suggest that the F-actin cables may be involved in the formation of the medial ring, and that tropomyosin plays an important role in organizing both the ring and the cable, but is not involved in the F-actin patch formation or maintenance of cell polarity.Binding of Arp3 to actin was revealed by immunoprecipitation as well as by DNase I column chromatography. Arp3 seemed to form a complex with several proteins in the cell extracts, as previously reported for other organisms. Contrary to a previous report (McCollum et al., EMBO J. 15, 6438-6446, 1996), Arp3 was found to be concentrated in the medial region from early anaphase to late cytokinesis. Following arp3 gene disruption, F-actin patches were delocalized throughout the cell and cells did not undergo polarized growth, suggesting that Arp3 influences the proper localization of the actin patches in the cell and thereby controls the polarized growth of the cell.