A novel regulatory way of smooth muscle contraction

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Myosin light-chain kinase (MLCK) of smooth muscle consists of an actin-binding domain at the N-terminal the catalytic domain in the central portion, and the myosin-binding domain at the C-terminal. The kinase activity is mediated by the catalytic domain that phosphorylates the myosin light-chain of 20 kDa (MLC20), activating smooth muscle myosin to interact with actin. Although the regulatory role of the kinase activity is well established, the role of non-kinase activity derived from actin-binding and myosin-binding domains remains unknown. We hypothesized that the actin-myosin interaction of smooth muscle could be activated by the non-kinase activity independently of MLC20 phosphorylation. We expressed cDNA of MLCK as a recombinant protein, and mutated it to lose the kinase activity, while keeping non-kinase activity unaffected. With the mutant, we detected an activating effect on the actin-myosin interaction. We also down-regulated MLCK expression by its RNAi in the cultured smooth muscle cells, to observe the lack of the activation. The rescue experiments with the MLCK-deficient cells demonstrated that MLC20 phosphorylation is not obligatory for the smooth muscle to contract.