Immunolocalization of cytoplasmic dynein to lysosomes in cultured cells.
Immunolocalization of cytoplasmic dynein to lysosomes in cultured cells.
复制标题
培养细胞中细胞质动力蛋白免疫定位至溶酶体。
作者：
Sharron X. H. Lin;C. A. Collins
从牛脑和大鼠睾丸中提纯的胞浆动力蛋白已被制备成多克隆抗血清。这些抗体与74 kDa动力蛋白中间链反应最强，但也识别410 kDa重链，以及先前观察到的与大鼠组织胞浆动力蛋白结合的150 kDa和45 kDa多肽。使用成纤维细胞系通过间接免疫荧光显微镜进行定位研究。动力蛋白特异性染色呈泡状，分布于整个细胞，但更多集中在细胞核附近。使用膜细胞器荧光标记的双标记研究表明动力蛋白与溶酶体共定位。动力蛋白阳性溶酶体的分布被微管活性药物处理细胞和细胞质酸化所破坏。将溶酶体的分布与外周微管进行比较，结果显示高度一致。这些结果与细胞质动力蛋白参与膜细胞器逆行运动的假设是一致的。在有丝分裂细胞中，动力蛋白染色沿着有丝分裂器官的微管也很明显，尽管泡状染色仍然很明显。动力蛋白在囊泡和纺锤体微管上的存在表明，动力蛋白在这些隔室之间的分布可能受到不同结合蛋白的调节。
Polyclonal antisera have been raised against cytoplasmic dynein purified from calf brain and rat testis. These antibodies reacted most strongly with the 74 kDa dynein intermediate chain, but also recognized the 410 kDa heavy chain, and the 150 and 45 kDa polypeptides previously observed to copurify with cytoplasmic dynein from rat tissues. Localization studies were performed by indirect immunofluorescence microscopy using a fibroblast cell line. Dynein-specific staining appeared vesicular, distributed throughout the cell, but more concentrated near the nucleus. Double-labeling studies using fluorescent markers for membranous organelles indicated a co-localization of dynein with lysosomes. The distribution of the dynein-positive lysosomes was disrupted by treatment of the cells with microtubule-active drugs, and by acidification of the cytoplasm. Comparison of the distribution of lysosomes with peripheral microtubules indicated a high degree of coincidence. These results are consistent with the hypothesis that cytoplasmic dynein is involved in retrograde-directed movement of membranous organelles. In mitotic cells, dynein staining was also apparent along the microtubules of the mitotic apparatus, though vesicular staining was still conspicuous. The presence of dynein on vesicles as well as on spindle microtubules indicates that dynein distribution between these compartments may be regulated by distinct binding proteins.
Hirokawa,N.：“脑动力蛋白 (MAP1C) 定位于顺行和逆行运输的膜细胞器。”
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