Myosin, actin, paramyosin and tropomyosin can be purified from N2 and mutant strains and shown to possess native properties by different functional and structural tests in vitro. One interesting physiological fact obtained from this approach is that, in nematode muscle, Ca++-mediated regulation of contraction is associated with proteins on both the thick and thin filaments. The application of these methods to obtaining purified native myosin from various
unc-54 strains has permitted us to clarify several questions concerning the previous observation of two kinds of myosin heavy chains in the body-wall. The two kinds of myosin heavy chain form primarily, if not exclusively, myosin molecules containing two such polypeptides of either one or the other class. The
unc-54 affected class of myosin is not required for the formation of bipolar or very long filaments or for complexing with paramyosin cores. Altered
unc-54 myosin although presumably disrupting the myofilament lattice in vivo, is capable of normal filament assembly in vitro. Preliminary studies suggest that
unc-54 myosin accumulates at a different rate than during development than the other myosins. The two major myosins are antigenically distinct from one another. This fact is permitting us to purify antibodies to
unc-54 myosin and to localize this myosin in bodywalls during development. A tentative inference from our studies and those of others is that the formation of a myofilament lattice involves the interaction of many gene products and actions, any one of which may have only a subtle influence. Accordingly, we are in the process of building up a sizeable collection of sarcomere-defective mutants. A wide range of phenotypes including a number of temperature-sensitive mutants has been obtained. We believe that only a rather complete genetic dissection in conjunction with detailed biochemical and structural studies will permit us to provide clearcut answers to such seemingly simple questions as 'why are there two myosins?'.