Drosophila muscle regulation characterized by electron microscopy and three-dimensional reconstruction of thin filament mutants.
Wild-type and mutant thin filaments were isolated directly from "myosinless" Drosophila indirect flight muscles to study the structural basis of muscle regulation genetically. Negatively stained filaments showed tropomyosin with periodically arranged troponin complexes in electron micrographs. Three-dimensional helical reconstruction of wild-type filaments indicated that the positions of tropomyosin on actin in the presence and absence of Ca(2+) were indistinguishable from those in vertebrate striated muscle and consistent with a steric mechanism of regulation by troponin-tropomyosin in Drosophila muscles. Thus, the Drosophila model can be used to study steric regulation. Thin filaments from the Drosophila mutant heldup(2), which possesses a single amino acid conversion in troponin I, were similarly analyzed to assess the Drosophila model genetically. The positions of tropomyosin in the mutant filaments, in both the Ca(2+)-free and the Ca(2+)-induced states, were the same, and identical to that of wild-type filaments in the presence of Ca(2+). Thus, cross-bridge cycling would be expected to proceed uninhibited in these fibers, even in relaxing conditions, and this would account for the dramatic hypercontraction characteristic of these mutant muscles. The interaction of mutant troponin I with Drosophila troponin C is discussed, along with functional differences between troponin C from Drosophila and vertebrates.
Cammarato
Anthony
A
Department of Physiology and Biophysics, Boston University School of Medicine, Boston, Massachusetts 02118-2526, USA.
Hatch
Victoria
V
Saide
Judith
J
Craig
Roger
R
Sparrow
John C
JC
Tobacman
Larry S
LS
Lehman
William
W
D008841:Actin Cytoskeleton / Q000737:chemistry* / Q000502:physiology / Q000648:ultrastructure*; D000818:Animals; D004330:Drosophila; D005786:Gene Expression Regulation / Q000502:physiology*; D005818:Genetic Engineering; D007090:Image Interpretation, Computer-Assisted / Q000379:methods*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008854:Microscopy, Electron / Q000379:methods*; D018482:Muscle, Skeletal / Q000737:chemistry / Q000502:physiology* / Q000648:ultrastructure*; D011817:Rabbits; D013045:Species Specificity; D013329:Structure-Activity Relationship