Mutations affecting the indirect flight muscles of Drosophila melanogaster

Development ◽  
1982 ◽  
Vol 69 (1) ◽  
pp. 61-81
Author(s):  
Ilan I. Deak ◽  
Paul R. Bellamy ◽  
Marianne Bienz ◽  
Yvonne Dubuis ◽  
Elke Fenner ◽  
...  

The development of the indirect flight muscles of Drosophila melanogaster was studied by analysing mutations that cause flightlessness. Twenty-five mutations on the X-chromosome and two on the third chromosome were examined. The X-chromosomal mutations form ten complementation units. The ten loci were assigned preliminary map positions by meiotic recombination and deficiencies and duplications. The two autosomal mutationsrepresent two genes. Gynandromorph analyses suggest that many of these mutations have their primary effect inthe presumptive thoracic muscle region of the embryo. The mutations cause a variety of characteristic defects, such as absence of the bulk of the thoracic muscle mass, or absence of only one of the two fibrillar musclegroups. Electronmicroscopic studies of sixteen mutants representing all twelve loci reveal abnormal myofibrillar organization in some of these mutants, e.g. aberrant or missing Z-bands, or absence of the thin filaments. Mutant protein patterns, obtained by SDS-polyacrylamide gel electrophoresis, show the following differences from wild type: ten mutants are characterized by absence or reduction of the 54 K protein, and most mutants exhibit a reduction and modification of the 80 and 90 K proteins. The absence or reduction of the 54 K proteinwas strongly correlated with aberrant Z-bands.

1986 ◽  
Vol 47 (1) ◽  
pp. 59-70 ◽  
Author(s):  
Trudy F. C. Mackay

SummaryForty-one third chromosomes extracted from a natural population of Drosophila melanogaster were assessed for net fitness and for the quantitative characters viability, net fertility, female productivity, male weight, abdominal bristle number, and sternopleural bristle number. Net homozygous and heterozygous fitness of the third chromosomes was estimated by competition against a marked balancer third chromosome. Average fitness of the homozygous lines relative to wild-type heterozygotes was 0·13, indicating substantial inbreeding depression for net fitness. All significant correlations of quantitative characters with fitness and with each other were high and positive. Homozygous fitness is strongly correlated with net fertility, viability, and female productivity, moderately associated with male weight, and not significantly associated with bristle traits. The combination of metric traits which best predicts homozygous fitness is the simple multiple of viability and female productivity. Heterozygous fitness is not correlated with homozygous fitness; furthermore, the relative contribution of metric traits to fitness in a heterozygous population is likely to be different from that deduced from homozygous lines. These observations are consistent with a model of genetic variation for fitness in natural populations caused by segregation of rare deleterious recessive alleles.


1989 ◽  
Vol 35 (10) ◽  
pp. 960-967 ◽  
Author(s):  
René De Mot ◽  
Jos Vanderleyden

Phenol-extracted total proteins from wild-type and mutant strains of Azospirillum brasilense and A. lipoferum were subjected to two-dimensional polyacrylamide gel electrophoresis. The protein patterns of both species could be readily distinguished by specific configurations of a limited number of spots. The resolving power of the technique allowed its application for strain fingerprinting. Thus, near-identity of A. brasilense strains Sp7 and Cd was demonstrated. In addition, minor changes in protein profiles resulting from spontaneous, chemically induced, or transposon-mediated mutations in the A. brasilense genome were evidenced. For a nitrate reductase negative mutant of strain Sp245 and a Sp7 mutant affected in exopolysaccharide synthesis, production of a truncated protein was involved. In Tn5-generated mutants, a gene dosage effect for the transposon-encoded neomycine phosphotransferase was observed.Key words: Azospirillum, two-dimensional protein analysis, fingerprinting.


Genetics ◽  
1994 ◽  
Vol 136 (1) ◽  
pp. 173-182 ◽  
Author(s):  
N S Petersen ◽  
D H Lankenau ◽  
H K Mitchell ◽  
P Young ◽  
V G Corces

Abstract The forked (f) gene of Drosophila melanogaster encodes six different transcripts 6.4, 5.6, 5.4, 2.5, 1.9, and 1.1 kb long. These transcripts arise by the use of alternative promoters. A polyclonal antibody raised against a domain common to all of the forked-encoded products has been used to identify forked proteins on two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and in Drosophila pupal tissues. The antibody stains fiber bundles present in bristle cells for about 15 hr during normal pupal development. Electron microscopy shows that these fibers are present from 40 to 53 hr in bristles of wild-type flies but are absent in the null f36a mutant. The forked protein(s) thus appear to be an essential part of the bristle fibers. The phenotype of the f36a mutation can be rescued by a 13-kb fragment of the forked locus containing the coding regions for the 2.5, 1.9, and 1.1-kb transcripts, suggesting that the proteins encoded by the three large forked RNAs are dispensable during bristle development. Increasing the copy number of a P[w+,f+] construct containing the 13-kb fragment induces a hypermorphic bristle phenotype whose severity correlates with the number of copies of P[w+,f+] present. These results indicate that alterations in the ratios among the forked proteins, or between forked products and other components of the fiber, result in abnormal assembly of the fibrillar cytoplasmic structures necessary for bristle morphogenesis.


1987 ◽  
Vol 7 (9) ◽  
pp. 3084-3091
Author(s):  
C C Karlik ◽  
D L Saville ◽  
E A Fyrberg

We have characterized two extant mutations of the flight muscle-specific act88F actin gene of Drosophila melanogaster. Both defective alleles were recovered from flightless mutants isolated previously (K. Mogami and Y. Hotta, Mol. Gen. Genet. 183:409-417, 1981). By directly sequencing the mutant alleles, we demonstrated that in act88FIfm(3)2 a single G-C to A-T transition converted arginine-28 to cysteine and that in act88FIfm(3)4 a single A-T to T-A transversion changed isoleucine-76 to phenylalanine. We showed that the actins encoded by either allele were strongly antimorphic. Mutant alleles effectively disrupted myofibril structure and function in the flight muscles of strains having the diploid complement of wild-type act88F genes. However, unlike antimorphic actins encoded by three previously characterized act88F alleles, neither that encoded by act88FIfm(3)2 nor that encoded by act88FIfm(3)4 was a strong inducer of heat shock protein synthesis.


Genetics ◽  
1987 ◽  
Vol 116 (3) ◽  
pp. 399-407
Author(s):  
Bruce McKee ◽  
Dan L Lindsley

ABSTRACT Deficiencies encompassing part or all of the X heterochromatin of Drosophila melanogaster have been linked to three abnormalities in male meiosis and spermatogenesis: X-Y nondisjunction, skewed sperm recovery ratios favoring sperm with reduced chromatin content, and sterility in males carrying either a Y-autosome translocation or mal  +  Y. In this study, 18 X heterochromatic deficiencies of varying sizes were tested in XY males for their spermatogenic phenotypes. All 18 proved to be either mutant for all three phenotypes or wild type for all three. Although variable among mutant deficiencies, expression levels of all three phenotypes were strongly correlated. Deficiencies that cause high levels of nondisjunction also cause severe recovery ratio distortion and are completely sterile in conjunction with mal  +  Y. Low nondisjunction deficiencies cause comparable mild effects for the other phenotypes. The same deficiencies were also tested in males carrying a large heterochromatic free X duplication Dp(1;f)3. For all deficiencies which induce nondisjunction in XY males, the Y and free duplication pair regularly and the X fails to pair in XYDp males. Drive levels are constant across deficiencies in these males. Thus elimination of variability in the pairing phenotype also eliminates variability in sperm recovery ratios.


1991 ◽  
Vol 114 (5) ◽  
pp. 941-951 ◽  
Author(s):  
C J Beall ◽  
E Fyrberg

We have investigated the molecular bases of muscle abnormalities in four Drosophila melanogaster heldup mutants. We find that the heldup gene encodes troponin-I, one of the principal regulatory proteins associated with skeletal muscle thin filaments. heldup3, heldup4, and heldup5 mutants, all of which have grossly abnormal flight muscle myofibrils, lack mRNAs encoding one or more troponin-I isoforms. In contrast, heldup2, an especially interesting mutant wherein flight muscles are atrophic, synthesizes the complete mRNA complement. By sequencing mutant troponin-I cDNAs we demonstrate that the molecular basis for muscle degeneration in heldup2 is conversion of an invariant alanine residue to valine. We finally show that degeneration of heldup2 thin filament/Z-disc networks can be prevented by eliminating thick filaments from flight muscles using a null allele of the sarcomeric myosin heavy chain gene. This latter observation suggests that actomyosin interactions exacerbate the structural or functional defect resulting from the troponin-I mutation.


1987 ◽  
Vol 7 (9) ◽  
pp. 3084-3091 ◽  
Author(s):  
C C Karlik ◽  
D L Saville ◽  
E A Fyrberg

We have characterized two extant mutations of the flight muscle-specific act88F actin gene of Drosophila melanogaster. Both defective alleles were recovered from flightless mutants isolated previously (K. Mogami and Y. Hotta, Mol. Gen. Genet. 183:409-417, 1981). By directly sequencing the mutant alleles, we demonstrated that in act88FIfm(3)2 a single G-C to A-T transition converted arginine-28 to cysteine and that in act88FIfm(3)4 a single A-T to T-A transversion changed isoleucine-76 to phenylalanine. We showed that the actins encoded by either allele were strongly antimorphic. Mutant alleles effectively disrupted myofibril structure and function in the flight muscles of strains having the diploid complement of wild-type act88F genes. However, unlike antimorphic actins encoded by three previously characterized act88F alleles, neither that encoded by act88FIfm(3)2 nor that encoded by act88FIfm(3)4 was a strong inducer of heat shock protein synthesis.


1994 ◽  
Vol 126 (3) ◽  
pp. 689-699 ◽  
Author(s):  
R M Cripps ◽  
K D Becker ◽  
M Mardahl ◽  
W A Kronert ◽  
D Hodges ◽  
...  

We have transformed Drosophila melanogaster with a genomic construct containing the entire wild-type myosin heavy-chain gene, Mhc, together with approximately 9 kb of flanking DNA on each side. Three independent lines stably express myosin heavy-chain protein (MHC) at approximately wild-type levels. The MHC produced is functional since it rescues the mutant phenotypes of a number of different Mhc alleles: the amorphic allele Mhc1, the indirect flight muscle and jump muscle-specific amorphic allele Mhc10, and the hypomorphic allele Mhc2. We show that the Mhc2 mutation is due to the insertion of a transposable element in an intron of Mhc. Since a reduction in MHC in the indirect flight muscles alters the myosin/actin protein ratio and results in myofibrillar defects, we determined the effects of an increase in the effective copy number of Mhc. The presence of four copies of Mhc results in overabundance of the protein and a flightless phenotype. Electron microscopy reveals concomitant defects in the indirect flight muscles, with excess thick filaments at the periphery of the myofibrils. Further increases in copy number are lethal. These results demonstrate the usefulness and potential of the transgenic system to study myosin function in Drosophila. They also show that overexpression of wild-type protein in muscle may disrupt the function of not only the indirect flight but also other muscles of the organism.


1980 ◽  
Vol 84 (3) ◽  
pp. 717-738 ◽  
Author(s):  
A Adoutte ◽  
R Ramanathan ◽  
R M Lewis ◽  
R R Dute ◽  
K Y Ling ◽  
...  

As a first step in the biochemical analysis of membrane excitation in wild-type Paramecium and its behavioral mutants we have defined the protein composition of the ciliary membrane of wild-type cells. The techniques for the isolation of cilia and ciliary membrane vesicles were refined. Membranes of high purity and integrity were obtained without the use of detergents. The fractions were characterized by electron microscopy, and the proteins of whole cilia, axonemes, and ciliary membrane vesicles were resolved by SDS polyacrylamide gel electrophoresis and isoelectric focusing in one and two dimensions. Protein patterns and EM appearance of the fractions were highly reproducible. Over 200 polypeptides were present in isolated cilia, most of which were recovered in the axonemal fraction. Trichocysts, which were sometimes present as a minor contaminant in ciliary preparations, were composed of a very distinct set of over 30 polypeptides of mol wt 11,000--19,000. Membrane vesicles contained up to 70 polypeptides of mol wt 15,000--250,000. The major vesicle species were a high molecular weight protein (the "immobilization antigen") and a group of acidic proteins with mol wt similar to or approximately 40,000. These and several other membrane proteins were specifically decreased or totally absent in the axoneme fraction. Tubulin, the major axonemal species, occurred only in trace amounts in isolated vesicles; the same was true for Tetrahymena ciliary membranes prepared by the methods described in this paper. A protein of mol wt 31,000, pI 6.8, was virtually absent in vesicles prepared from cells in exponential growth phase, but became prominent early in stationary phase in good correlation with cellular mating reactivity. This detailed characterization will provide the basis for comparison of the ciliary proteins of wild-type and behavioral mutants and for analysis of topography and function of membrane proteins. It will also be useful in future studies of trichocysts and mating reactions.


2000 ◽  
Vol 151 (7) ◽  
pp. 1483-1500 ◽  
Author(s):  
Mary C. Reedy ◽  
Belinda Bullard ◽  
Jim O. Vigoreaux

Flightin is a multiply phosphorylated, 20-kD myofibrillar protein found in Drosophila indirect flight muscles (IFM). Previous work suggests that flightin plays an essential, as yet undefined, role in normal sarcomere structure and contractile activity. Here we show that flightin is associated with thick filaments where it is likely to interact with the myosin rod. We have created a null mutation for flightin, fln0, that results in loss of flight ability but has no effect on fecundity or viability. Electron microscopy comparing pupa and adult fln0 IFM shows that sarcomeres, and thick and thin filaments in pupal IFM, are 25–30% longer than in wild type. fln0 fibers are abnormally wavy, but sarcomere and myotendon structure in pupa are otherwise normal. Within the first 5 h of adult life and beginning of contractile activity, IFM fibers become disrupted as thick filaments and sarcomeres are variably shortened, and myofibrils are ruptured at the myotendon junction. Unusual empty pockets and granular material interrupt the filament lattice of adult fln0 sarcomeres. Site-specific cleavage of myosin heavy chain occurs during this period. That myosin is cleaved in the absence of flightin is consistent with the immunolocalization of flightin on the thick filament and biochemical and genetic evidence suggesting it is associated with the myosin rod. Our results indicate that flightin is required for the establishment of normal thick filament length during late pupal development and thick filament stability in adult after initiation of contractile activity.


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