CA++-CALMODULIN-DEPENDENT PHOSPHORYLATION OF MYOSIN, AND ITS ROLE IN BRUSH-BORDER CONTRACTION INVITRO
CA++-CALMODULIN-DEPENDENT PHOSPHORYLATION OF MYOSIN, AND ITS ROLE IN BRUSH-BORDER CONTRACTION INVITRO
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DOI：
10.1083/jcb.95.3.943
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发表时间：
1982-01-01
影响因子：
7.8
通讯作者：
MOOSEKER, MS
中科院分区：
文献类型：
--
作者：
KELLER, TCS;MOOSEKER, MS
我们重新研究了 Ca++ 和 ATP 对从肠上皮细胞中分离出的刷状缘的影响。在37℃时，Ca++(1/tM)和ATP引起刷状缘末端网的剧烈收缩，而不是如之前报道的微绒毛的收缩(MS Mooseker，1976，J.Cell Biol.71：417-433)。末端网状收缩在 37°C 的温度下持续 1-5 分钟，主动收缩刷状缘的粘附小带水平。收缩需要 ATP，由 Ca++(1/xM) 刺激，并且发生在膜完整和脱膜刷状缘中。微绒毛核心的 Ca++ 依赖性隔离需要 Ca++ 浓度略高于收缩所需的浓度 (10/~ M)。在刷状缘收缩的条件下，孤立的刷状缘中的许多蛋白质被磷酸化。然而，只有一种刷状缘蛋白，即刷状缘肌球蛋白的 20,000 道尔顿 (20-kdalton) 轻链 (BBMLC2o) 的磷酸化受到 Ca+ 的刺激。 37°C 时，BBMLC2o 磷酸化与刷状缘收缩直接相关。此外，BBMLC2o 磷酸化和刷状缘收缩均受到三氟拉嗪（一种抑制钙调蛋白活性的抗精神病吩噻嗪）的抑制。这些结果表明，Ca++ 通过刺激细胞骨架相关的、Ca+÷-和钙调蛋白依赖性刷状缘肌球蛋白轻链激酶来调节体外刷状缘收缩性。Ca++ 已被认为是非肌肉细胞中运动性和细胞质结构的调节剂。例如，游离Ca ++ 浓度的变化可以通过Ca ++ -钙调蛋白依赖性肌球蛋白轻链激酶控制肌球蛋白调节轻链的磷酸化来调节脊椎动物非肌肉和平滑肌细胞的运动性。平滑肌和非肌肉肌球蛋白的调节轻链的磷酸化会增加其肌动蛋白激活的 Mg++-ATP 酶活性，可能会增加力的产生并引起运动（有关综述，请参阅参考文献 1）。
We have reinvestigated the effects of Ca++ and ATP on brush borders isolated from intestinal epithelial cells. At 37 C, Ca++(1/tM) and ATP cause a dramatic contraction of brush border terminal webs, not a retraction of microvilli as previously reported (MS Mooseker, 1976, J. Cell Biol. 71: 417-433). Terminal web contraction, which occurs over the course of 1-5 min at 37 C, actively constricts brush borders at the level of their zonula adherens. Contraction requires ATP, is stimulated by Ca++(1/xM), and occurs in both membrane-intact and demembranated brush borders. Ca÷÷-dependent-solation of microvillus cores requires a concentration of Ca÷+ slightly greater (10/~ M) than that required for contraction. Under conditions in which brush borders contract, many proteins in the isolated brush borders become phosphorylated. However, the phosphorylation of only one of the brush border proteins, the 20,000 dalton (20-kdalton) light chain of brush border myosin (BBMLC2o), is stimulated by Ca÷+. At 37 C, BBMLC2o phosphorylation correlates directly with brush border contraction. Furthermore, both BBMLC2o phosphorylation and brush border contraction are inhibited by trifluoperazine, an anti-psychotic phenothiazine that inhibits calmodulin activity. These results indicate that Ca++ regulates brush border contractility in vitro by stimulating cytoskeleton-associated, Ca+÷-and calmodulin-dependent brush border myosin light chain kinase.Ca++ has been implicated as a regulator of both motility and cytoplasmic structure in nonmuscle ceils. For example, changes in free Ca++ concentration may regulate motility in vertebrate nonmuscle and smooth muscle cells by controlling phosphorylation of the regulatory light chain of myosin by a Ca++-calmodulin-dependent myosin light chain kinase. Phosphorylation of the regulatory light chain of smooth and nonmuscle myosin increases its actin-activated Mg++-ATPase activity, presumably increasing force production and causing motility (for review, see reference 1).