Correlation of myosin light chain phosphorylation with isometric contraction of fibroblasts.
Correlation of myosin light chain phosphorylation with isometric contraction of fibroblasts.
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发表时间：
1993-11
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通讯作者：
M. Kolodney;Elliot L. Elson
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作者：
M. Kolodney;Elliot L. Elson
体外研究表明，来自非肌肉细胞的肌球蛋白II的酶活性是由其调节轻链(LC20)的磷酸化控制的。我们通过测量活的鸡胚胎成纤维细胞(CEF)产生的收缩力量，研究了在活的鸡胚胎成纤维细胞(CEF)中磷酸化LC20的一个可能的功能后果。使用最近开发的方法，我们记录了一组细胞在有丝分裂刺激后产生的等长力的定量变化。胎牛血清、凝血酶和溶血磷脂酸刺激CEF快速等长收缩。用凝血酶刺激的细胞在5-10分钟内产生最大力。力量的发展在时间上与LC_(20)磷酸化的总比例增加3-5倍有关，与单磷酸化和二磷酸化状态的LC_(20)的比例在时间上相关。在凝血酶刺激后，无负荷缩短率也增加。虽然力和磷酸化水平在刺激后10min开始下降，但磷酸化水平下降的速度比力更快。这些结果表明，LC20磷酸化在调节成纤维细胞收缩中的作用类似于它在调节平滑肌收缩中已被公认的作用，对成纤维细胞(或其他培养细胞)所产生的力的定量测量可用于在体内从分子水平上研究非肌节肌球蛋白的调节。
In vitro studies have indicated that the enzymatic activity of myosin II from non-muscle cells is controlled by phosphorylation of its regulatory light chain (LC20). We have studied one likely functional consequence of phosphorylating LC20 in living chick embryo fibroblasts (CEF) by measuring contractile force developed by these cells. Using a recently developed method, we recorded quantitative changes in isometric force generated by a population of cells following mitogenic stimulation. Fetal bovine serum, thrombin, and lysophosphatidic acid stimulate rapid isometric contraction of CEF. Cells stimulated with thrombin develop maximal force within 5-10 min. Force development correlates temporally with a 3-5-fold increase in the overall fraction of LC20 phosphorylated and with the fractions of LC20 in both the monophosphorylated and diphosphorylated states. Unloaded shortening velocity also increases after thrombin stimulation. Although both force and phosphorylation begin to decline 10 min after stimulation, the level of phosphorylation declined more rapidly than the force. These results suggest that the role of LC20 phosphorylation in regulating fibroblast contractility is analogous to its well established role in regulating smooth muscle contraction and that quantitative measurements of the force developed by populations of fibroblasts (or other cultured cells) can be used to study the regulation of non-sarcomeric myosin at the molecular level in vivo.