Actin‐binding proteins regulate the work performed by myosin II motors on single actin filaments

Abstract

Regulation of actin/myosin II force generation by calcium [Kamm and Stull, Annu. Rev. Physiol. 51:299-313, 1989] and phosphorylation of myosin II light chains [Sellers and Adelstein, "The Enzymes," Vol. 18, Orlando, FL: Academic Pres, 1987, pp. 381-418] is well established. However, additional regulation of actin/myosin II force generation/contraction may result from actin-binding proteins [Stossel et al., Ann. Rev. Cell Biol. 1:353-402, 1985; Pollard and Cooper, Ann. Rev. Biochem. 55:987-1035, 1986] as they affect the gel state of the actin cytomatrix [reviewed in Taylor and Condeelis, Int. Rev. Cytol., 56:57-143, 1979]. Regulation of the gel state of actin may determine whether an isotonic or isometric contraction results from the interaction between myosin and actin. We have extended the single actin filament motility assay of Kron and Spudich [Proc. Natl. Acad. Sci. U.S.A. 83:6272-6276, 1986] by including filamin or alpha-actinin on the substrate with myosin II to examine how actin-crosslinking proteins regulate the movements of single actin filaments. Increasing amounts of actin-crosslinking proteins inhibit filament velocity and decrease the number of filaments moving. Reversal of crosslinking yields increased velocities and numbers of moving filaments. These results support the solation-contraction coupling hypothesis [see Taylor and Fechheimer, Phil. Trans. Soc. London B 299:185-197, 1982] which proposes that increased crosslinking of actin inhibits myosin-based contraction. This study also illustrates the potentially varied roles of different actin-crosslinking proteins and offers a novel method to examine actin-binding protein activity and their regulation of motility at the single molecule level.

Interaction of a Dictyostelium member of the plastin/fimbrin family with actin filaments and actin-myosin complexes.

“…Our results differ from those of a previous study (Janson et al, 1992) in which two F-actin cross-linking proteins, nonmuscle a-actinin and filamin, were bound, along with the myosin heads, to a solid support. Under these conditions, the actin filaments added were primarily arrested at the surface rather than cross-linked to each other.…”

Mammalian Myosin-18A, a Highly Divergent Myosin

“…sin-2 is mixed with active, phosphorylated smooth muscle myosin-2 (39) or when actin-binding proteins are mixed with actively cycling myosins (40). This is consistent with the optical trapping results showing transient interactions with actin in a nucleotide-independent manner that do not result in a net displacement of the actin filament.…”

Molecular consequences of the R453C hypertrophic cardiomyopathy mutation on human β-cardiac myosin motor function

“…Specifically, we used the in vitro motility assay in the presence of an external load to examine whether the R453C mutant's ensemble force-generating ability was altered. We introduced a frictional load in the in vitro motility assay using α-actinin, an actin-binding protein that leads to a molecular tug of war with myosin (18,19). Typically, the amount of α-actinin needed to stop filament movement is reported as an indication of the myosin isometric force.…”