Abstract
The alternatively spliced SM1 and SM2 smooth muscle myosin heavy chains differ at their respective carboxyl termini by 43 versus 9 unique amino acids. To determine whether these tailpieces affect filament assembly, SM1 and SM2 myosins, the rod region of these myosin isoforms, and a rod with no tailpiece (tailless), were expressed in Sf 9 cells. Paracrystals formed from SM1 and SM2 rod fragments showed different modes of molecular packing, indicating that the tailpieces can influence filament structure. The SM2 rod was less able to assemble into stable filaments than either SM1 or the tailless rods. Expressed full-length SM1 and SM2 myosins showed solubility differences comparable to the rods, establishing the validity of the latter as a model for filament assembly. Formation of homodimers of SM1 and SM2 rods was favored over the heterodimer in cells coinfected with both viruses, compared with mixtures of the two heavy chains renatured in vitro. These results demonstrate for the first time that the smooth muscle myosin tailpieces differentially affect filament assembly, and suggest that homogeneous thick filaments containing SM1 or SM2 myosin could serve distinct functions within smooth muscle cells.
Highlights
The vertebrate smooth muscle myosin heavy chain is encoded by a single gene, but alternative splicing at two sites can create four different variants of this ف200-kD polypeptide
We produced full-length smooth muscle myosin molecules and rod fragments containing the SM1 or SM2 tailpiece, as well as a rod fragment, named tailless (TL), whose sequence was terminated at the alternative splice site (Fig. 1)
The SM2 and TL structures which we observed closely resemble end-to-end aggregates of the bipolar segments formed by proteolytically derived rod from chicken gizzard myosin (Kendrick-Jones et al, 1971), which is likely to be lacking a tailpiece
Summary
The vertebrate smooth muscle myosin heavy chain is encoded by a single gene, but alternative splicing at two sites can create four different variants of this ف200-kD polypeptide. The longer SM1 isoform appears developmentally earlier than SM2 in most mammals (Mohammad and Sparrow, 1988; Eddinger and Wolf, 1993), and its expression is maintained during the proliferative phase of cell culture, whereas SM2 expression ceases (Rovner et al, 1986a). These distinct patterns of expression suggest unique roles for the two isoforms in smooth muscle cell function, but a clear difference in properties between them has yet to be established in vitro.
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