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Inhibition of Contraction and Myosin Light Chain Phosphorylation in Guinea‐Pig Smooth Muscle by p21‐Activated Kinase 1
Author(s) -
Wirth A.,
Schroeter M.,
KockHauser C.,
Manser E.,
Chalovich J. M.,
Lanerolle P.,
Pfitzer G.
Publication year - 2003
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2002.033167
Subject(s) - myosin light chain kinase , taenia coli , phosphorylation , caldesmon , myosin , muscle contraction , contraction (grammar) , actin , myosin light chain phosphatase , smooth muscle tissue , pak1 , kinase , biology , microbiology and biotechnology , chemistry , biochemistry , biophysics , calmodulin , anatomy , endocrinology , smooth muscle , enzyme
The p21‐activated protein kinases (PAKs) have been implicated in cytoskeletal rearrangements and modulation of non‐muscle contractility. Little, however, is known about the role of the PAK family members in smooth muscle contraction. Therefore, we investigated the effect of the predominant isoform in vascular smooth muscle cells, PAK1, on contraction and phosphorylation of the regulatory light chains of myosin (r‐MLC) in Triton‐skinned guinea‐pig smooth muscle. We also investigated which of the three putative substrates at the contractile apparatus ‐ MLCK, caldesmon or r‐MLC ‐ is phosphorylated by PAK1 in smooth muscle tissue. Incubation of Triton‐skinned carotid artery and taenia coli from guinea‐pig with an active mutant of PAK1 in relaxing solution for 30–60 min resulted in inhibition of submaximal force by about 50 %. The mechanism of inhibition of force was studied in the Triton‐skinned taenia coli. In this preparation, inhibition of force was associated with a respective inhibition of r‐MLC phosphorylation. In the presence of the myosin phosphatase inhibitor, microcystin‐LR (10 μ m ), the rate of contraction and r‐MLC phosphorylation elicited at pCa 6.79 were both decreased. Because under these conditions the rate of r‐MLC phosphorylation is solely dependent on MLCK activity, this result suggests that the inhibitory effect of PAK1 on steady‐state force and r‐MLC phosphorylation is due to inhibition of MLCK. In line with this, we found that MLCK was significantly phosphorylated by PAK1 while there was very little 32 P incorporation into caldesmon. PAK1 phosphorylated isolated r‐MLC but not those in the skinned fibres or in purified smooth muscle myosin II. In conclusion, these results suggest that PAK1 attenuates contraction of skinned smooth muscle by phosphorylating and inhibiting MLCK.