Worm Breeder's Gazette 2(2): 27
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
We have pursued detailed studies of the structure of N2 body-wall muscle as a necessary condition for deciphering the abnormalities of various sarcomere-defective mutants, particularly many which have more subtle defects than unc-54 mutants, and also to understand structural changes in body-wall muscle cells during development. Using polarized light microscopy, we find that the body wall muscle cells in relaxed, anesthetized N2 animals are arranged right to left +/-11.9 to the anterior-posterior axis. The sarcomeres within these cells are arranged +/- 5.9 to the same axis. Under these conditions, the sarcomere width normal to the cell is 1.67 and the similarly measured A band width is 1.0 . By both polarized light and electron microscopy we find that the orientation of the thick filaments is parallel to the N2 anterior-posterior axis. Thus, from these observations, the length of the thick filaments is calculated to be 9. 7 , assuming that all the thick filaments are held in register. It would be difficult to accurately measure the length of such very long filaments by electron microscopy of thin sectioned material. In roller mutants such as the dominant SU1006 where the cell orientation is significantly different from N2 and variable, all the internal relationships and dimensions are preserved. Similarly in dumpy mutants such as E61, although cell lengths are shortened, all other features such as orientation and internal dimensions are similar to N2. Electron microscopy of fixed, dehydrated, embedded and stained transverse thin sections of N2 body-wall muscles previously obtained have been ambiguous as to whether or not the thick filaments are arranged in any regular lattice. On the basis of new sections and freeze-etching work, we suggest that indeed there is an hexagonal packing of the thick filaments in the A-bands but that this packing may become easily disordered due to contraction or to preparation. We have been able to identify a new set of structures closely associated with the dense bodies to which the thin filaments attach. These are similar to the intermediate or 100 angstrom filaments observed in other muscles and non-muscle cells, and they may represent a kind of cytoskeleton. Observation of negatively stained, isolated and crosslinked dense bodies confirms these associations. Bridges between isolated thick and thin filaments can also be observed by these techniques. Examples of the application of these methods to more subtle sarcomere-defective mutants such as R73, a partial revertant of E569 ( unc-54), and E286, a temperature-sensitive mutant (unc-45).In connection with biochemical work in the laboratory concerning the two body-wall myosins and changes in their relative concentrations during development, studies of sarcomere structure during development and of immunochemical localization of myosin have been initiated. The dimensions of A-bands and sarcomeres remain in variant from 24 hours to 60 hours after hatching. However, the number of sarcomeres per cell changes from about two to eight during this development. Specific antibody to myosin using a perioxidase method shows strong staining only in the sarcomeres of the body-wall and in the pharynx. Studies are in progress on the localization of unc-54 affected myosin. This method may be useful in determining the molecular composition of abnormal structures in various mutants.