Ishijima S, Kubo-Irie M, Mohri H, Hamaguchi Y
Biological Laboratory, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, Japan.
J Cell Sci. 1996 Dec;109 ( Pt 12):2833-42. doi: 10.1242/jcs.109.12.2833.
Active sliding between doublet microtubules of sea urchin sperm axonemes that were demembranated with Triton X-100 in the presence or absence of calcium was induced with ATP and elastase at various concentrations of Ca2+ to examine the effects of Ca2+ on the direction of the power stroke of the dynein arms. Dark-field light microscopy of microtubule sliding revealed that the sliding from the axonemes demembranated with Triton and millimolar calcium and disintegrated with ATP and elastase showed various patterns of sliding disintegration, including loops of doublet microtubules formed near the head or the basal body. These loops were often thicker than the remaining axonemal bundle. In contrast, only thinner loops were found from the axonemes demembranated with Triton in the absence of calcium and disintegrated with ATP and elastase at high Ca2+ concentrations. Electron microscopic examination of the direction of microtubule sliding showed that the doublet microtubules in the axonemes demembranated in the presence of millimolar calcium moved toward the base of the axonemes by the dynein arms on the adjacent doublet microtubule as well as by their own dynein arms. Doublet microtubules in the axonemes demembranated in the absence of calcium moved toward the base of the axonemes only by their own dynein arms. Similar observations have been obtained from the axonemes from which the outer dynein arms were selectively extracted. From these observations, we can conclude that the dynein arms generate force in both directions and this feature of the dynein arms arises from at least the inner dynein arms.
在有或无钙存在的情况下，用Triton X-100对海胆精子轴丝进行去膜处理，然后在不同浓度的Ca2+条件下，用ATP和弹性蛋白酶诱导其双联体微管之间发生主动滑动，以研究Ca2+对动力蛋白臂动力冲程方向的影响。微管滑动的暗视野光学显微镜观察显示，用Triton和毫摩尔浓度的钙去膜、并用ATP和弹性蛋白酶使其解体的轴丝的滑动呈现出各种解体模式，包括在头部或基体附近形成的双联体微管环。这些环通常比其余的轴丝束更粗。相比之下，在无钙条件下用Triton去膜、并在高Ca2+浓度下用ATP和弹性蛋白酶使其解体的轴丝中，只发现了较细的环。对微管滑动方向的电子显微镜检查表明，在毫摩尔浓度钙存在的情况下去膜的轴丝中的双联体微管，通过相邻双联体微管上的动力蛋白臂以及它们自身的动力蛋白臂朝着轴丝基部移动。在无钙条件下去膜的轴丝中的双联体微管仅通过它们自身的动力蛋白臂朝着轴丝基部移动。从选择性提取了外侧动力蛋白臂的轴丝中也获得了类似的观察结果。从这些观察中，我们可以得出结论，动力蛋白臂在两个方向上都能产生力，并且动力蛋白臂的这一特性至少源于内侧动力蛋白臂。
