Role of thin-filament regulatory proteins in relaxation of colonic smooth muscle contraction.

Coordinated regulation of smooth muscle contraction and relaxation is required for colonic motility. Contraction is associated with phosphorylation of myosin light chain (MLC(20)) and interaction of actin with myosin. Thin-filament regulation of actomyosin interaction is modulated by two actin-binding regulatory proteins: tropomyosin (TM) and caldesmon (CaD). TM and CaD are known to play crucial role in actomyosin interaction promoting contraction. Contraction is associated with phosphorylation of the small heat shock protein HSP27, concomitant with the phosphorylation of TM and CaD. Phosphorylation of HSP27 is attributed as being the prime modulator of thin-filament regulation of contraction. Preincubation of colonic smooth muscle cells (CSMC) with the relaxant neurotransmitter vasoactive intestinal peptide (VIP) showed inhibition in phosphorylation of HSP27 (ser78). Attenuation of HSP27 phosphorylation can result in modulation of thin-filament-mediated regulation of contraction leading to relaxation; thus the role of thin-filament regulatory proteins in a relaxation milieu was investigated. Preincubation of CSMC with VIP exhibited a decrease in phosphorylation of TM and CaD. Furthermore, CSMC preincubated with VIP showed a reduced association of TM with HSP27 and with phospho-HSP27 (ser78) whereas there was reduced dissociation of TM from CaD and from phospho-CaD. We thus propose that, in addition to alteration in phosphorylation of MLC(20), relaxation is associated with alterations in thin-filament-mediated regulation that results in termination of contraction.