Proton pumps populate the contractile vacuoles of Dictyostelium amoebae.
Proton pumps populate the contractile vacuoles of Dictyostelium amoebae.
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DOI：
10.1083/jcb.121.6.1311
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发表时间：
1993-06
期刊：
影响因子：
--
通讯作者：
Clarke M
中科院分区：
文献类型：
--
作者：
Heuser J;Zhu Q;Clarke M
真核微生物盘基网柄藻的变形虫被发现含有相互连接的小管和池阵列，其膜上布满直径为15 nm的“钉”。“将这些钉的超微结构和冷冻断裂行为与其他细胞和组织中发现的类似结构进行比较，表明它们是空泡型质子泵的头部区域。当破碎的阿米巴暴露于抗哺乳动物液泡H（+）-ATP酶B亚基的抗血清时，观察到钉装饰和聚集，从而支持了这一鉴定。PEG丰富的池在速冻阿米巴的外观取决于它们的渗透环境：在高渗条件下，池是平坦的，有许多狭窄的管状延伸，而在低渗条件下，池的范围从球根状到球形。然而，在所有情况下，它们的内容都像纯水一样深深地蚀刻。这些性质表明，相互连接的小管和池构成了网骨藻的收缩液泡系统。早期的研究已经证明，网骨藻中的收缩空泡膜极其富含钙调蛋白（Zhu，Q.，和M. Clarke，1992，J. Cell Biol.118：347-358）。光镜免疫荧光证实，对空泡质子泵的抗体与抗钙调素抗体共定位在这些细胞器。延时视频记录的生活阿米巴成像的干涉反射显微镜，或荧光显微镜染色后，收缩泡膜与电位敏感的苯乙烯染料，揭示的程度和动态的相互关系的池和管状元件在Dictyosteelium的收缩泡系统。整个膜的质子泵的高密度表明，质子梯度的产生可能是一个重要的因素，在收缩空泡的液体积累的机制。
Amoebae of the eukaryotic microorganism Dictyostelium discoideum were found to contain an interconnected array of tubules and cisternae whose membranes were studded with 15-nm-diameter "pegs." Comparison of the ultrastructure and freeze-fracture behavior of these pegs with similar structures found in other cells and tissues indicated that they were the head domains of vacuolar-type proton pumps. Supporting this identification, the pegs were observed to decorate and clump when broken amoebae were exposed to an antiserum against the B subunit of mammalian vacuolar H(+)-ATPase. The appearance of the peg-rich cisternae in quick-frozen amoebae depended on their osmotic environment: under hyperosmotic conditions, the cisternae were flat with many narrow tubular extensions, while under hypo-osmotic conditions the cisternae ranged from bulbous to spherical. In all cases, however, their contents deep etched like pure water. These properties indicated that the interconnected tubules and cisternae comprise the contractile vacuole system of Dictyostelium. Earlier studies had demonstrated that contractile vacuole membranes in Dictyostelium are extremely rich in calmodulin (Zhu, Q., and M. Clarke, 1992, J. Cell Biol. 118: 347-358). Light microscopic immunofluorescence confirmed that antibodies against the vacuolar proton pump colocalized with anti-calmodulin antibodies on these organelles. Time-lapse video recording of living amoebae imaged by interference-reflection microscopy, or by fluorescence microscopy after staining contractile vacuole membranes with potential-sensitive styryl dyes, revealed the extent and dynamic interrelationship of the cisternal and tubular elements in Dictyostelium's contractile vacuole system. The high density of proton pumps throughout its membranes suggests that the generation of a proton gradient is likely to be an important factor in the mechanism of fluid accumulation by contractile vacuoles.