Yoko Yano Toyoshima, Stephen J. Kron,
Elizabeth M. McNally*, Kenneth R. Niebling,
Chikashi Toyoshima & James A. Spudich
Department of Cell Biology, Stanford University School of Medicine,
Stanford, California 94305, USA
*Department of Microbiology and Immunology, Albert Einstein College of Medicine,
Bronx, New York 10461, USA
The rotating crossbridge model for muscle contraction proposes that force
is produced by a change in angle of the crossbridge between the overlapping
thick and thin filaments. Myosin, the major component of the thick filament,
is comprised of two heavy chains and two pairs of light chains. Together
they form two globular heads, which give rise to the crossbridge in muscle,
and a coiled-coil rod, which forms the shaft of the thick filament. The
isolated head fragment, subfragment-1 (S1), contains the ATPase and actin-binding
activities of myosin (Fig.1). Although S1 seems to have the
requisite enzymatic activity, direct evidence that S1 is sufficient to drive
actin movement has been lacking. It has long been recognized that in
vitro assays are an important approach for identifying the elements in
muscle responsible for force generation. Hynes et al. showed that
beads coated with heavy meromyosin (HMM), a soluble proteolytic fragment of
myosin consisting of a part of the rod and the two heads, can move on
Nitella actin filaments. Using the myosin-coated surface assay of Kron
and Spudich, Harada et al. showed that single-headed myosin filaments
bound to glass support movement of actin at nearly the same speed as intact
myosin filaments. These studies show that the terminal portion of the rod and
the two-headed nature of myosin are not required for movement. To restrict
the region responsible for movement further, we have modified the myosin-
coated surface assay by replacing the glass surface with a nitrocellulose
film. Here we report that myosin filaments, soluble myosin, HMM or S1,
when bound to a nitrocellulose film, support actin sliding movement (Fig. 2).
That S1 is sufficient to cause sliding movement of actin filaments in vitro
gives strong support to models of contraction that place the site of active
movement in muscle within the myosin head.