Regulation of expressed truncated smooth muscle myosins. Role of the essential light chain and tail length.
Regulatory light chain (RLC) phosphorylation controls smooth muscle myosin's ability to act as a molecular motor. If thick filament-regulated myosins share a common structural basis for the "on-off" switch, then the interface between the RLC and essential light chain (ELC) should be as important in smooth muscle myosin as it is for calcium-dependent regulation in molluscan myosin (Xie, X., Harrison, D. H., Schlichting, I., Sweet, R. M., Kalabokis, V. N., Szent-Györgyi, A. G., and Cohen, C. (1994) Nature 368, 306-312). To test this hypothesis, a baculovirus expression system was used to obtain a truncated smooth muscle myosin that contained its native RLC and either smooth ELC, skeletal ELC, or no ELC. Movement of actin in a motility assay by heavy meromyosin (HMM) containing either skeletal or smooth ELC occurred at the same rate and in a phosphorylation-dependent manner. In contrast, substitution of skeletal RLC for smooth RLC produced an inactive molecule. HMM without an ELC moved actin at 25% the rate of control HMM, but the movement was phosphorylation-dependent even without an RLC/ELC interface. Poor regulation was observed, however, when the tail was truncated from 72 to 27 nm even though this species was primarily double-headed. These results suggest that the molecular changes induced at the active site by phosphorylation can occur independent of the ELC, but critically depend upon a stable coiled-coil tail that determines how the RLCs interact at the head/rod junction.
Trybus
K M
KM
Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts 02254.
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D008969:Molecular Sequence Data; D009036:Moths; D009130:Muscle, Smooth / Q000378:metabolism*; D009218:Myosins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D010766:Phosphorylation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014422:Turkeys
D000199:Actins; D011994:Recombinant Proteins; D009218:Myosins