Drosophila melanogaster troponin-T mutations engender three distinct syndromes of myofibrillar abnormalities.

Abstract In vertebrates troponin complexes interact co-operatively with tropomyosin dimers to modulate skeletal muscle contraction. In order further to investigate troponin assembly and function in vivo , we are developing molecular genetic approaches. Here we report characterization of the gene that encodes Drosophila tropinin-T and analyses of muscle defects engendered by several mutant alleles. We found that the Drosophila troponin-T locus specifies at least three proteins having sequences similar to vertebrate troponin-T. All are significantly larger than any avian or mammalian isoforms, however, due to a highly acidic carboxy-terminal extension. Comparisons of the chromosomal arrangements of vertebrate and Drosophila troponin-T genes revealed that the location of one intron-exon boundary is conserved. This observation and the similarity of vertebrate and Drosophila troponin-T primary sequences suggest that the respective proteins are homologous, and that troponin-T pre-dates the divergence of vertebrate and invertebrate organisms. In situ hybridization of the Drosophila troponin-T gene to polytene chromosomes demonstrated that it resides within subdivision 12A of the X chromsome, precisely where upheld and indented thorax flight muscle mutations have been mapped previously. We determined the nucleotide sequences of troponin-T genes in five extant mutants. All have deleterious alterations, directly establishing that upheld and indented thorax muscle abnormalities are due to defective troponin-T. Two of the alleles, upheld 2 and upheld 3 , apparently disrupt RNA splicing and eliminate most or all troponin-T from flight and jump muscles, while the remaining three alleles change the identities of single amino acids of troponin-T. Electron microscopy of mutant muscles revealed that the two null alleles eliminate thin filaments, except where they are bound by electron-dense material presumed to be Z-disc proteins. Two of the point mutations, upheld 101 and indented thorax 3 , do not perturb assembly of myofibrils, but cause their degeneration within days after muscles begin to be utilized. The final mutation, upheld whu , reduces the diameter of the myofibril lattice by approximately one-half. We propose hypotheses to explain how each troponin-T mutation engenders the observed myofibrillar defects.