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Signaling through myosin light chain kinase in smooth muscles
Ning Gao1, Jian Huang1, Weiqi He2
1Department of Physiology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390.
Abstract:
Ca(2+)/calmodulin-dependent myosin light chain kinase (MLCK) phosphorylates smooth muscle myosin regulatory light chain (RLC) to initiate contraction. We used a tamoxifen-activated, smooth muscle-specific inactivation of MLCK expression in adult mice to determine whether MLCK was differentially limiting in distinct smooth muscles. A 50% decrease in MLCK in urinary bladder smooth muscle had no effect on RLC phosphorylation or on contractile responses, whereas an 80% decrease resulted in only a 20% decrease in RLC phosphorylation and contractile responses to the muscarinic agonist carbachol. Phosphorylation of the myosin light chain phosphatase regulatory subunit MYPT1 at Thr-696 and Thr-853 and the inhibitor protein CPI-17 were also stimulated with carbachol. These results are consistent with the previous findings that activation of a small fraction of MLCK by limiting amounts of free Ca(2+)/calmodulin combined with myosin light chain phosphatase inhibition is sufficient for robust RLC phosphorylation and contractile responses in bladder smooth muscle. In contrast, a 50% decrease in MLCK in aortic smooth muscle resulted in 40% inhibition of RLC phosphorylation and aorta contractile responses, whereas a 90% decrease profoundly inhibited both responses. Thus, MLCK content is limiting for contraction in aortic smooth muscle. Phosphorylation of CPI-17 and MYPT1 at Thr-696 and Thr-853 were also stimulated with phenylephrine but significantly less than in bladder tissue. These results indicate differential contributions of MLCK to signaling. Limiting MLCK activity combined with modest Ca(2+) sensitization responses provide insights into how haploinsufficiency of MLCK may result in contractile dysfunction in vivo, leading to dissections of human thoracic aorta.
Insights
Myosin light chain kinase (MLCK) plays a different role in smooth muscle contraction. Bladder smooth muscle contraction is not limited by MLCK, but aortic smooth muscle contraction is.
Area of Science:
- Physiology
- Molecular Biology
- Pharmacology
Background:
- Smooth muscle contraction relies on myosin light chain kinase (MLCK) to phosphorylate myosin regulatory light chain (RLC).
- The role of MLCK in regulating contraction varies across different smooth muscle types.
- Understanding MLCK's contribution is crucial for explaining contractile dysfunction.
Purpose of the Study:
- To investigate the differential contribution of MLCK to smooth muscle contraction in distinct tissues.
- To determine if MLCK expression levels are limiting for contractile function in urinary bladder and aortic smooth muscle.
- To explore the impact of MLCK reduction on RLC phosphorylation and associated signaling pathways.
Main Methods:
- Utilized tamoxifen-inducible, smooth muscle-specific inactivation of MLCK in adult mice.
- Assessed RLC phosphorylation and contractile responses in urinary bladder and aortic smooth muscle following MLCK reduction.
- Measured phosphorylation of MYPT1 and CPI-17 in response to agonists.
Main Results:
- A 50% decrease in MLCK in bladder smooth muscle had no effect on RLC phosphorylation or contraction.
- An 80% decrease in bladder MLCK caused only a 20% reduction in RLC phosphorylation and carbachol-induced contraction.
- In aortic smooth muscle, a 50% MLCK decrease inhibited RLC phosphorylation and contraction by 40%, while a 90% decrease profoundly inhibited both.
- Agonist-stimulated phosphorylation of MYPT1 and CPI-17 was observed in both tissues but was less pronounced in the aorta.
Conclusions:
- MLCK content is not limiting for urinary bladder smooth muscle contraction, suggesting other mechanisms like myosin light chain phosphatase inhibition are sufficient.
- MLCK content is limiting for aortic smooth muscle contraction, highlighting its critical role in this tissue.
- Differential MLCK contributions and modest Ca(2+) sensitization responses offer insights into how MLCK haploinsufficiency can lead to contractile dysfunction and aortic dissections.
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