Correlation of myosin light chain phosphorylation with isometric contraction of fibroblasts.
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.
Kolodney
M S
MS
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, Missouri 63110.
Elson
E L
EL
D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D003094:Collagen / Q000502:physiology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D007537:Isometric Contraction / Q000187:drug effects; D008934:Mitogens / Q000494:pharmacology; D009218:Myosins / Q000378:metabolism*; D010766:Phosphorylation
D008934:Mitogens; D003094:Collagen; D009218:Myosins