MOLECULAR CYTOCHEMISTRY - INCORPORATION OF FLUORESCENTLY LABELED ACTIN INTO LIVING CELLS
MOLECULAR CYTOCHEMISTRY - INCORPORATION OF FLUORESCENTLY LABELED ACTIN INTO LIVING CELLS
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DOI：
10.1073/pnas.75.2.857
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发表时间：
1978-01-01
影响因子：
11.1
通讯作者：
WANG, YL
中科院分区：
文献类型：
--
作者：
TAYLOR, DL;WANG, YL
用5-碘乙酰胺荧光素标记的肌动蛋白，通过直接显微注射掺入到混沌卡罗琳和多头绒泡菌的肌动蛋白功能池中。标记的肌动蛋白的功能活性进行了分析，在3个层次的组织：与纯化的肌动蛋白，在运动提取物的细胞和活的运动细胞。标记的肌动蛋白表现出正常的聚合和激活肌球蛋白ATP酶的类似程度作为未标记的控制。标记的肌动蛋白和内源性肌动蛋白被纳入收缩小球在运动细胞提取物中的程度大致相同。将标记后的肌动蛋白显微注射到单个C. carolinensis细胞中，荧光肌动蛋白扩散到内质和外质中，而不形成明显的纤维。显微注射标记肌动蛋白后，多头绒泡菌的外质中出现了荧光束。这种实验方法与荧光光谱技术相结合，可能成为研究活细胞中组分的胞内分布和结构变化的有力工具。
Actin labeled with 5-iodoacetamidofluorescein was incorporated into the functional pool of actin in Chaos carolinensis and Physarum polycephalum by direct microinjection. The functional activity of the labeled actin was analyzed at 3 levels of organization as: with the purified actin, in motile extracts of cells and in living motile cells. The labeled actin exhibited normal polymerization and activated myosin ATPase to a similar extent as unlabeled controls. Labeled actin and endogenous actin were incorporated into contracted pellets to approximately the same extent in motile cell extracts. After labeled actin was microinjected into single C. carolinensis cells, the fluorescent actin spread into both the endoplasm and ectoplasm without forming distinct fibrils. Fluorescent bundles developed in the ectoplasm of P. polycephalum following microinjection of labeled actin. This experimental method in conjunction with fluorescence spectroscopic techniques could become a powerful tool for studying the intracellular distribution and structural changes of components in living cells.