Functional and ultrastructural effects of a missense mutation in the indirect flight muscle-specific actin gene of Drosophila melanogaster.
A single-site mutation of the flight-muscle-specific actin gene of Drosophila melanogaster causes a substitution of glutamic acid 93 by lysine in all the actin encoded in the indirect flight muscle (IFM). In these Act88FE93K mutants, myofibrillar bundles of thick and thin filaments are present but lack Z-discs and all sarcomeric repeats. Dense filament bundles, which are probably aberrant Z-discs, are seen in myofibrils of pupal flies, but early in adult life these move to the periphery of the fibrils and are not seen in skinned adult fibres. Consistent with this observation, alpha-actinin and other high molecular weight proteins, possibly associated with Z-discs, are not detected on SDS/polyacrylamide gels or Western blots of skinned adult IFM. The mutation lies at the beginning of a loop in the small domain of actin, near the myosin binding region. However, that the mutant actin binds myosin heads is shown by (1) rigor crossbridges in electron micrographs, (2) the appropriate rise in stiffness when ATP is withdrawn in mechanical experiments, and (3) equal protection against tryptic digestion provided by rigor binding between actin and myosin in both wild-type and mutant fibres. Reversal of rigor chevron angle along some thin filaments reflects reversal of thin-filament polarity due to lattice disorder. The absence of Z-discs, alpha-actinin and two high molecular weight proteins, and binding studies by others, suggest that the substitution at residue 93 affects the binding of the mutant actin to a protein, possibly alpha-actinin, which is necessary for Z-disc assembly or maintenance.
Department of Biology, University of York, Heslington, U.K.
J
J
Sparrow
M
M
Reedy
E
E
Ball
V
V
Kyrtatas
J
J
Molloy
J
J
Durston
E
E
Hennessey
D
D
White
D000199:Actins / Q000235:genetics* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004331:Drosophila melanogaster / Q000235:genetics* / Q000502:physiology; D005426:Flight, Animal; D005971:Glutamates; D018698:Glutamic Acid; D008239:Lysine; D008854:Microscopy, Electron; D009124:Muscle Proteins / Q000302:isolation & purification; D009126:Muscle Relaxation; D009132:Muscles / Q000502:physiology / Q000648:ultrastructure*; D009154:Mutation; D009210:Myofibrils / Q000648:ultrastructure; D010449:Peptide Mapping
D000199:Actins; D005971:Glutamates; D009124:Muscle Proteins; D018698:Glutamic Acid; D008239:Lysine