Immunocytochemical study of endocytotic structures accumulated in HeLa cells transformed with a temperature-sensitive mutant of dynamin
Immunocytochemical study of endocytotic structures accumulated in HeLa cells transformed with a temperature-sensitive mutant of dynamin
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DOI：
10.1177/002215549904700506
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发表时间：
1999-05-01
影响因子：
3.2
通讯作者：
Ohno, S
中科院分区：
文献类型：
--
作者：
Baba, T;Ueda, H;Ohno, S
动力蛋白是一种100 kd的GTPase，它是网格蛋白介导的内吞作用所必需的。最近的研究表明，动力蛋白与网格蛋白包被囊泡的形成密切相关。在这项研究中，我们研究了用温度敏感（ts）突变体转化的HeLa细胞中积累的内吞结构的超微结构，以阐明dyn（ts）细胞中的哪一步被阻断。转铁蛋白受体的内吞作用在表面连接的膜结构水平上受到限制。管状和囊泡膜内陷在细胞的外周区域积累，表明在包被囊泡形成的夹断步骤之前，内吞作用被阻断。据报道，“项圈”管位于shibire（ts1）果蝇的神经末梢和GTP γ s处理的突触体中，即使在不允许的温度下，也没有在dynts细胞中观察到。dyn（ts）细胞深内陷包被凹区中动力蛋白的分布与dyn（wt）细胞相似，但与另一种内吞缺陷突变细胞dyn（K44A）细胞不同。这些形态学数据表明，dynts阻断了网格蛋白包被囊泡形成的夹断步骤，这可能是由与dyn（K44A）细胞不同的机制引起的。
Dynamin is a 100-kD GTPase, which is required for clathrin-mediated endocytosis. Recent studies have revealed that dynamin is closely involved in clathrin-coated vesicle formation. In this study we investigated the ultrastructure of endocytotic structures accumulated in HeLa cells that were transformed with a temperature-sensitive (ts) mutant of dynamin to clarify which step was blocked in dyn(ts) cells. Endocytosis of transferrin receptors was restricted at the level of surface-connected membrane structures. Tubular and vesicular membrane invaginations were accumulated in the cells' peripheral regions, suggesting that the endocytosis was blocked just before the pinching-off steps in coated vesicle formation. The "collared" tubes, which were reported to be localized in nerve terminals in shibire(ts1) flies and GTP gamma S-treated synaptosomes, were not observed in the dynts cells even at nonpermissive temperature. The distribution pattern of dynamin in deeply invaginated coated pits in dyn(ts) cells was similar to that in dyn(wt) cells but not to that in dyn(K44A) cells, which are other endocytosis-defective mutant cells. These morphological data suggest that dynts blocked the pinching-off steps in clathrin-coated vesicle formation, which may be caused by a different mechanism from that of dyn(K44A) cells.