title: Evidence for regulation of lamellipodial and tail contraction of glycerinated chicken embryonic fibroblasts by myosin light chain kinase.
abstract: Permeabilized cell models of muscle and nonmuscle cells have proven useful for examining the regulation of actin, myosin, and other cytoskeletal proteins during cell contraction. Upon addition of Ca2+ and ATP, glycerinated chick embryonic skin fibroblasts retract their tails and lamellipodia. Ca2+-independent contractions are obtained by preincubation of cell models in Ca2+ ATP gamma S, followed by EGTA and ATP addition, or by addition of trypsin-treated myosin light chain kinase that no longer requires Ca2+ for reactivation. By pretreating cells before glycerination with colchicine, it is possible to study lamellipodial contraction independent of tail contraction. Similar responses to ATP gamma S pretreatment and unregulated myosin light chain kinase are observed in cells that only contain lamellipodia. SDS-PAGE electrophoresis of glycerinated fibroblasts incubated in ATP gamma 35S and Ca2+ shows that only two major proteins are thiophosphorylated, and that one of them, a band that comigrates with the 20K MW light chain of myosin, is thiophosphorylated in a Ca2+-dependent manner. Since the rate of tail contraction is several-fold faster after Ca2+ and ATP gamma S pretreatment or incubation in excess myosin light chain kinase, myosin light chain phosphorylation may be a rate-limiting step during contraction.
mesh_terms: D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002642:Chick Embryo; D003598:Cytoskeletal Proteins / Q000502:physiology; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D005990:Glycerol; D009119:Muscle Contraction / Q000187:drug effects; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*
chemical_list: D003598:Cytoskeletal Proteins; D009219:Myosin-Light-Chain Kinase; D005990:Glycerol; D002118:Calcium