Diphosphorylation of the myosin regulatory light chain enhances the tension acting on stress fibers in fibroblasts

Abstract

Regulation of the contractile force is crucial for cell migration, cell proliferation, and maintenance of cell morphology. Phosphorylation of the myosin II regulatory light chain (MRLC) is involved in these processes. To show whether the diphosphorylation of MRLC increases the tension acting on stress fibers, changes in the stiffness of fibroblasts expressing wild-type MRLC and a mutant type, which cannot be diphosphorylated, on treatment with lysophosphatidic acid (LPA) were examined by a mechanical-scanning probe microscope (M-SPM). The LPA treatment increased cellular stiffness in the wild-type MRLC expressing cells, while it had no effect on the mutated cells. Immunostaining showed that LPA stimulation induced the diphosphorylation of MRLC. These results suggest that the diphosphorylation of MRLC enhances the tension acting on stress fibers.3

Transgene integration into the human AAVS1 locus enhances myosin II-dependent contractile force by reducing expression of myosin binding subunit 85

“…Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting were performed as previously described [8]. Information on the primary antibodies used in this study is presented in Supplemental Table 2.…”

“…Since phosphorylation of MRLC enhances cellular contractile force [8], reduction of MBS85 expression may increase cellular contractile force by increasing MRLC phosphorylation. Cellular contractile force is the sole contributor to the mechanical function of organs and tissues such as the heart and skeletal muscle.…”

The Number of Cyclic Stretch Regulates Cellular Elasticity in C2C12 Myoblasts

“…To survey molecular mechanisms as to how cyclic stretch affect the cellular elasticity response by a single step-like stretch, we focused on phosphorylation of MRLC, which are thought to contribute to cellular elasticity [15]. First, we examined changes in pp-MRLC with a single step-like stretch and/or with cyclic stretches.…”

“…First, we examined changes in pp-MRLC with a single step-like stretch and/or with cyclic stretches. Previous reports have shown that pp-MRLC increases intracellular contractile force and cellular elasticity [15], and diphosphorylation can be induced by a single step-like stretch [16]. To quantify changes in pp-MRLC, we stained cells with anti-pp-MRLC and measured the intensity ( Figure 5).…”

Mechanical response of single myoblasts to various stretching patterns visualized by scanning probe microscopy

“…These results indicate that so-called "tensional homeostasis" exists at the single cell level (Mizutani et al, 2004). Moreover, this mechanical response originates in the diphosphorylation of myosin regulatory light chain of myosin II combined with F-actin, which increases cellular tension acting on stress fibers (Mizutani et al, 2006). SPM is unique positioned to visualize not only the surface topography but also the spatial distribution of the local viscoelasticity of living cells in a culture medium (Radmacher et al, 1996;Haga et al, 2000).…”