MLCK-independent phosphorylation of MLC20 and its regulation by MAP kinase pathway in human bladder smooth muscle cells.
Myosins are a superfamily of actin-based molecular motor proteins, which hydrolyze ATP and generate various forms of eukaryotic motility and muscle contraction. Myosin light chain 20 (MLC20) is small ring around the neck region of heavy chain of myosins. Phosphorylation of MLC20 is thought to play a key role in regulation of smooth muscle contraction. Calcium- and calmodulin-dependent myosin light chain kinase (MLCK) is considered the primary regulator of MLC20 phosphorylation. However, several observations in smooth muscle contraction cannot be explained by the mode of phosphorylation. By performing a series of experiments in vitro and in vivo, we report here MLCK-independent MLC20 phosphorylation. Gene expression study reveals that expression of MLCK in smooth muscles is inconsistent with MLC20 phosphorylation at Ser19. None of inactivating calmodulin/MLCK, depriving of calcium and silencing MLCK expression by siRNA blocks effectively the phosphorylation of MLC20 at Ser19. In addition, by overexpressing active human MAP (mitogen-activated protein)-ERK kinase kinase-1 (MEKK1) and blocking its downstream messengers, we have demonstrated a new regulatory system of MLC phosphorylation via MEKK1, which downregulates Ser19 phosphorylation of MLC20 through its downstream molecules, p38, JNK, and ERK in human bladder smooth muscle cells.
Deng
Maoxian
M
Department of Animal Biology, School of Animal Husbandry and Veterinary Medicine, Jiangsu Polytechnic College of Agriculture and Forestry, Jurong, Jiangsu, China. dengok@hotmail.com
Ding
Wei
W
Min
Xuewen
X
Xia
Ying
Y
D000199:Actins / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002465:Cell Movement; D002478:Cells, Cultured; D006801:Humans; D047468:Immunoprecipitation; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009119:Muscle Contraction; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D018994:Myosin Light Chains / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000235:genetics; D001743:Urinary Bladder / Q000166:cytology / Q000378:metabolism*
D000199:Actins; D018994:Myosin Light Chains; D034741:RNA, Small Interfering; D009219:Myosin-Light-Chain Kinase; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium