Effects of calponin on isometric force and shortening velocity in permeabilized Taenia coli smooth muscle

Calponin, a thin filament-associated protein, inhibits actomyosin adenosinetriphosphatase in solution and has been suggested to modulate smooth muscle contractility. We used permeabilized guinea pig taenia coli smooth muscle to investigate whether calponin can modulate actin-myosin interaction in a more organized contractile system. Fibers were permeabilized with Triton X-100 and glycerol, which permit access of large macromolecules to the contractile apparatus. For contractures elicited by Ca2+ (6.6 mu M + 0.1 mu M calmodulin), the recombinant alpha-isoform of chicken gizzard calponin (CaP) decreased isometric force (F-o) and unloaded shortening velocity (V-us) in a dose-dependent manner; 1 mu M CaP had minimal effects on force (< 10%) but reduced V-us by similar to 50% and 10 mu M CaP reduced F-o to 27% of control and V-us to near zero levels. To eliminate any effects of the binding of calmodulin by CaP and consequent inhibition of myosin light chain kinase activity, we also studied fibers activated by thiophosphorylation of the myosin regulatory light chain. F-o was only moderately inhibited, remaining at similar to 75% of control in the presence of CaP (10 mu M), whereas V-us was reduced to 32% of control. A similar inhibition was obtained with a mutant (CaPcys175) that retains the ability to bind to actin. CaP phosphorylated by protein kinase C and CaPcys175 mutant labeled with 1,5-IAEDANS, which bind actin poorly, were not effective inhibitors. Our results indicate that 1) CaP more strongly inhibits V-us (-cross-bridge cycle rate) than F-o (similar to number of activated cross bridges) and 2) the effects of CaP are related to its binding to actin. Thus the function of CaP in regulation of smooth muscle contractility may be more strongly related to its function as a modulator of velocity, as related to the ''latch state,'' than as an ''on-off' switch.