Abstract
Despite their near sequence identity, actin isoforms cannot completely replace each other in vivo and show marked differences in their tissue-specific and subcellular localization. Little is known about isoform-specific differences in their interactions with myosin motors and other actin-binding proteins. Mammalian cytoplasmic β- and γ-actin interact with nonsarcomeric conventional myosins such as the members of the nonmuscle myosin-2 family and myosin-7A. These interactions support a wide range of cellular processes including cytokinesis, maintenance of cell polarity, cell adhesion, migration, and mechano-electrical transduction. To elucidate differences in the ability of isoactins to bind and stimulate the enzymatic activity of individual myosin isoforms, we characterized the interactions of human skeletal muscle α-actin, cytoplasmic β-actin, and cytoplasmic γ-actin with human myosin-7A and nonmuscle myosins-2A, -2B and -2C1. In the case of nonmuscle myosins-2A and -2B, the interaction with either cytoplasmic actin isoform results in 4-fold greater stimulation of myosin ATPase activity than was observed in the presence of α-skeletal muscle actin. Nonmuscle myosin-2C1 is most potently activated by β-actin and myosin-7A by γ-actin. Our results indicate that β- and γ-actin isoforms contribute to the modulation of nonmuscle myosin-2 and myosin-7A activity and thereby to the spatial and temporal regulation of cytoskeletal dynamics. FRET-based analyses show efficient copolymerization abilities for the actin isoforms in vitro. Experiments with hybrid actin filaments show that the extent of actomyosin coupling efficiency can be regulated by the isoform composition of actin filaments.
Highlights
Mammalian actin isoforms are highly conserved and ubiquitously found in eukaryotic cells
Our results show clear increases in actin-activation and functional competence for all nonsarcomeric myosins tested, when they were allowed to interact with cytoplasmic isoactins instead of a-actin
Experiments, myosin-7A showed a distinct preference for c-actin over b-actin. In their proper physiological context, myosin-7A, band c-actin play a key role in the development, function and maintenance of cochlear hair cell stereocilia [14,39]
Summary
Mammalian actin isoforms are highly conserved and ubiquitously found in eukaryotic cells. The amino acid sequence of a-actin differs from cytoplasmic actin isoforms in more than 20 residues that are spread over the entire molecule. Cytoplasmic actins are ubiquitous and play a pivotal role in cell motility, intracellular transport, cell shape maintenance or mitosis [3]. They undergo spatial and temporal segregation during the formation of stress fibers and actin-based cell protrusions [4,5]. It is mainly organized as a branched meshwork with cortical and lamellar localization in moving cells, but can colocalize with b-actin in lamellipodia or be recruited into stress fibres [3,12]. Multiple missense mutations of c-actin have been described (T89I, K118M, K118N, I122V, E241K, P264L, T278I, P332A, V370A), all of them are associated with autosomal dominant non-syndromic sensorineural progressive hearing loss [21,22,23,24,25,26,27]
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