title: Distinct kinases are involved in contraction of cat esophageal and lower esophageal sphincter smooth muscles.
abstract: Contraction of smooth muscle depends on the balance of myosin light chain kinase (MLCK) and myosin light chain phosphatase (MLCP) activities. Because MLCK activation depends on the activation of calmodulin, which requires a high Ca(2+) concentration, phosphatase inhibition has been invoked to explain contraction at low cytosolic Ca(2+) levels. The link between activation of the Ca(2+)-independent protein kinase Cepsilon (PKCepsilon) and MLC phosphorylation observed in the esophagus (ESO) (Sohn UD, Cao W, Tang DC, Stull JT, Haeberle JR, Wang CLA, Harnett KM, Behar J, and Biancani P. Am J Physiol Gastrointest Liver Physiol 281: G467-G478, 2001), however, has not been elucidated. We used phosphatase and kinase inhibitors and antibodies to signaling enzymes in combination with intact and saponin-permeabilized isolated smooth muscle cells from ESO and lower esophageal sphincter (LES) to examine PKCepsilon-dependent, Ca(2+)-independent signaling in ESO. The phosphatase inhibitors okadaic acid and microcystin-LR, as well as an antibody to the catalytic subunit of type 1 protein serine/threonine phosphatase, elicited similar contractions in ESO and LES. MLCK inhibitors (ML-7, ML-9, and SM-1) and antibodies to MLCK inhibited contraction induced by phosphatase inhibition in LES but not in ESO. The PKC inhibitor chelerythrine and antibodies to PKCepsilon, but not antibodies to PKCbetaII, inhibited contraction of ESO but not of LES. In ESO, okadaic acid triggered translocation of PKCepsilon from cytosolic to particulate fraction and increased activity of integrin-linked kinase (ILK). Antibodies to the mitogen-activated protein (MAP) kinases ERK1/ERK2 and to ILK, and the MAP kinase kinase (MEK) inhibitor PD-98059, inhibited okadaic acid-induced ILK activity and contraction of ESO. We conclude that phosphatase inhibition potentiates the effects of MLCK in LES but not in ESO. Contraction of ESO is mediated by activation of PKCepsilon, MEK, ERK1/2, and ILK.
authors.affiliation: Department of Medicine, Seoul National University, Bundang Hospital, Seoungnam, Gyeronggi-Do 463-707, Korea.
authors.forename: Nayoung
authors.initials: N
authors.lastname: Kim
authors.forename: Weibiao
authors.initials: W
authors.lastname: Cao
authors.forename: In Sung
authors.initials: IS
authors.lastname: Song
authors.forename: Chung Yong
authors.initials: CY
authors.lastname: Kim
authors.forename: Karen M
authors.initials: KM
authors.lastname: Harnett
authors.forename: Ling
authors.initials: L
authors.lastname: Cheng
authors.forename: Michael P
authors.initials: MP
authors.lastname: Walsh
authors.forename: Piero
authors.initials: P
authors.lastname: Biancani
mesh_terms: D000818:Animals; D000906:Antibodies; D002415:Cats; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004943:Esophagogastric Junction / Q000201:enzymology*; D005260:Female; D008297:Male; D008387:Marine Toxins; D052998:Microcystins; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D009130:Muscle, Smooth / Q000166:cytology / Q000201:enzymology*; D032389:Myocytes, Smooth Muscle / Q000201:enzymology; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D019319:Okadaic Acid / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D051744:Protein Kinase C-epsilon; D054645:Protein Phosphatase 1; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D000099200:Scaffold Protein ILK
chemical_list: D000906:Antibodies; D004791:Enzyme Inhibitors; D008387:Marine Toxins; D052998:Microcystins; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases; D020928:Mitogen-Activated Protein Kinases; D009219:Myosin-Light-Chain Kinase; D019319:Okadaic Acid; D010456:Peptides, Cyclic; D010749:Phosphoprotein Phosphatases; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D054645:Protein Phosphatase 1; D017346:Protein Serine-Threonine Kinases; D000099200:Scaffold Protein ILK; C057862:cyanoginosin LR