Molecular-level evidence of force maintenance by smooth muscle myosin during LC20 dephosphorylation

Abstract

Smooth muscle (SM) is found in most hollow organs of the body. Phasic SM, as found in the gut, contracts to propel content, whereas tonic SM, as found in most blood vessels, maintains tension. This force maintenance is referred to as the latch state and occurs at low levels of myosin activation (myosin light chain [LC20] phosphorylation). Molecular mechanisms have been proposed to explain the latch state but have been studied only at the whole-muscle level because of technological limitations. In the current study, an assay chamber was devised to allow injection of myosin light chain phosphatase (MLCP) during laser trap and in vitro motility assays, without creating bulk flow, to reproduce latch state conditions at the molecular level. Using the laser trap in a single-beam mode, an actin filament was brought in contact with several myosin molecules on a pedestal. Myosin pulled on the actin filament until a plateau force was reached, at which point, MLCP was injected. Force maintenance was observed during LC20 dephosphorylation, the level of which was assessed in a parallel in vitro motility assay performed in the same conditions. Force was maintained longer for myosin purified from tonic SM than from phasic SM. These data support the longstanding dogma of strong bonds caused by dephosphorylated, noncycling cross-bridges. Furthermore, MLCP injection in an in vitro motility mixture assay performed with SM and skeletal muscle myosin suggests that the maintenance of these strong bonds is possible only if no energy is provided by surrounding actively cycling myosin molecules.

Alteration of the Expression and Functional Activities of Myosin II Isoforms in Enlarged Hyperplastic Prostates

“…In short, the SM-1, SM-A, and MLC 17b isoforms are associated with tonic-type contractions (such as in the aorta), characterized by a slow shortening speed (V max ) but high force maintenance. On the other hand, the SM-2, SM-B, and MLC 17a isoforms are associated with phasic force generation (such as in the bladder), characterized by low force maintenance but fast V max and rapid rates of force activation and relaxation [8,11,12]. The SM cells also express type II nonmuscle myosin (NMM II).…”