Signaling processes for initiating smooth muscle contraction upon neural stimulation.
Relationships among biochemical signaling processes involved in Ca2+/calmodulin (CaM)-dependent phosphorylation of smooth muscle myosin regulatory light chain (RLC) by myosin light chain kinase (MLCK) were determined. A genetically-encoded biosensor MLCK for measuring Ca(2+)-dependent CaM binding and activation was expressed in smooth muscles of transgenic mice. We performed real-time evaluations of the relationships among [Ca2+](i), MLCK activation, and contraction in urinary bladder smooth muscle strips neurally stimulated for 3 s. Latencies for the onset of [Ca2+](i) and kinase activation were 55 +/- 8 and 65 +/- 6 ms, respectively. Both increased with RLC phosphorylation at 100 ms, whereas force latency was 109 +/- 3 ms. [Ca2+](i), kinase activation, and RLC phosphorylation responses were maximal by 1.2 s, whereas force increased more slowly to a maximal value at 3 s. A delayed temporal response between RLC phosphorylation and force is probably due to mechanical effects associated with elastic elements in the tissue. MLCK activation partially declined at 3 s of stimulation with no change in [Ca2+](i) and also declined more rapidly than [Ca2+](i) during relaxation. The apparent desensitization of MLCK to Ca2+ activation appears to be due to phosphorylation in its calmodulin binding segment. Phosphorylation of two myosin light chain phosphatase regulatory proteins (MYPT1 and CPI-17) or a protein implicated in strengthening membrane adhesion complexes for force transmission (paxillin) did not change during force development. Thus, neural stimulation leads to rapid increases in [Ca2+](i), MLCK activation, and RLC phosphorylation in phasic smooth muscle, showing a tightly coupled Ca2+ signaling complex as an elementary mechanism initiating contraction.
Ding
Hai-Lei
HL
Department of Physiology, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA.
Ryder
Jeffrey W
JW
Stull
James T
JT
Kamm
Kristine E
KE
D000818:Animals; D002118:Calcium / Q000502:physiology; D002147:Calmodulin / Q000502:physiology; D004558:Electric Stimulation; D007537:Isometric Contraction / Q000502:physiology; D007700:Kinetics; D051379:Mice; D008822:Mice, Transgenic; D009119:Muscle Contraction / Q000502:physiology*; D009130:Muscle, Smooth / Q000201:enzymology / Q000294:innervation* / Q000502:physiology*; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D009474:Neurons / Q000502:physiology; D015398:Signal Transduction; D001743:Urinary Bladder / Q000201:enzymology / Q000502:physiology*
D002147:Calmodulin; D009219:Myosin-Light-Chain Kinase; D002118:Calcium