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bioRxiv · 10.1101/2023.05.22.541840

p90RSK2, a new MLCK, rescues contractility in myosin light chain kinase null smooth muscle

Kalra, J. K.·Artamonov, M.·Wang, H.·Franke, A.·Markowska, Z.·Li, J.·Derewenda, Z. S.·Ayon, R. J.·Somlyo, A. V.
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Abstract

BackgroundPhosphorylation of smooth muscle (SM) myosin regulatory light chain (RLC20) is a critical switch leading to contraction or cell migration. The canonical view held that the only kinase catalyzing this reaction is the short isoform of myosin light chain kinase (MLCK1). Auxiliary kinases may be involved and play a vital role in blood pressure homeostasis. We have previously reported that p90 ribosomal S6 kinase (RSK2) functions as such a kinase, in parallel with the classical MLCK1, contributing [~]25% of the maximal myogenic force in resistance arteries and regulating blood pressure. Here, we take advantage of a MLCK1 null mouse to further test our hypothesis that RSK2 can function as an MLCK, playing a significant physiological role in SM contractility. MethodsFetal (E14.5-18.5) SM tissues were used as embryos die at birth. We investigated the necessity of MLCK for contractility, cell migration and fetal development and determined the ability of RSK2 kinase to compensate for the lack of MLCK and characterized its signaling pathway in SM. ResultsAgonists induced contraction and RLC20 phosphorylation in mylk1-/- SM, that was inhibited by RSK2 inhibitors. Embryos developed and cells migrated in the absence of MLCK. The pCa-tension relationships in WT vs mylk1-/- muscles demonstrated a Ca2+-dependency due to the Ca2+-dependent tyrosine kinase Pyk2, known to activate PDK1 that phosphorylates and fully activates RSK2. The magnitude of contractile responses was similar upon addition of GTP{gamma}S to activate the RhoA/ROCK pathway. The Ca2+-independent component was through activation of Erk1/2/PDK1/RSK2 leading to direct phosphorylation of RLC20, to increase contraction. RSK2, PDK1, Erk1/2 and MLCK formed a signaling complex on the actin filament, optimally positioning them for interaction with adjacent myosin heads. ConclusionsRSK2 signaling constitutes a new third signaling pathway, in addition to the established Ca2+/CAM/MLCK and RhoA/ROCK pathways to regulate SM contractility and cell migration.

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Kalra, J. K., Artamonov, M., Wang, H., Franke, A., Markowska, Z., Li, J., Derewenda, Z. S., Ayon, R. J., Somlyo, A. V.. 2023-05-22. p90RSK2, a new MLCK, rescues contractility in myosin light chain kinase null smooth muscle. https://doi.org/10.1101/2023.05.22.541840

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