Structure of Thick Filaments from Drosophila Indirect Flight Muscle by Cryo-EM

We have determined the molecular defect of the Drosophila melanogaster myosin heavy chain (MHC) mutation Mhc and the mutation's effect on indirect flight muscle, jump muscle, and larval intersegmental muscle. We show that the Mhc1 mutation is essentially a null allele which results in the dominant-flightless and recessive-lethal phenotypes associated with this mutant (Mogami, K., P. T. O'Donnell, S. I. Bernstein, T. R. F. Wright, C. P. Emerson, Jr. 1986. Proc. Natl. Acad. Sci. USA. 83:1393-1397). The mutation is a 101-bp deletion in the MHC gene which removes most of exon 5 and the intron that precedes it. S1 nuclease mapping indicates that mutant transcripts follow two alternative processing pathways. Both pathways result in the production of mature transcripts with altered reading frames, apparently yielding unstable, truncated MHC proteins. Interestingly, the preferred splicing pathway uses the more distal of two available splice donor sites. We present the first ultrastrutural characterization of a completely MHC-null muscle and show that it lacks any discernable thick filaments. Sarcomeres in these muscles are completely disorganized suggesting that thick filaments play a critical role in sarcomere assembly. To understand why the Mhc1 mutation severely disrupts indirect flight muscle and jump muscle function in heterozygotes, but does not seriously affect the function of other muscle types, we examined the muscle ultrastructure of Mhc1/+ heterozygotes. We find that these organisms have a nearly 50% reduction in the number of thick filaments in indirect flight muscle, jump muscle, and larval intersegmental muscle. In addition, aberrantly shaped thick filaments are common in the jump muscle and larval intersegmental muscle. We suggest that the differential sensitivity of muscle function to the Mhc1 mutation is a consequence of the unique myofilament arrays in each of these muscles. The highly variable myofilament array of larval intersegmental muscle makes its function relatively insensitive to changes in thick filament number and morphology. Conversely, the rigid double hexagonal lattice of the indirect flight muscle, and the organized lattice of the jump muscle cannot be perturbed without interfering with the specialized and evolutionarily more complex functions they perform.

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Characterization of components of Z-bands in the fibrillar flight muscle of Drosophila melanogaster.

The Journal of Cell Biology ◽

10.1083/jcb.109.5.2157 ◽

1989 ◽

Vol 109 (5) ◽

pp. 2157-2167 ◽

Cited By ~ 78

Author(s):

J D Saide ◽

S Chin-Bow ◽

J Hogan-Sheldon ◽

L Busquets-Turner ◽

J O Vigoreaux ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Monoclonal Antibodies ◽

Flight Muscle ◽

Cross Reactivity ◽

Antigenic Determinants ◽

Immunogold Staining ◽

Thick Filaments ◽

The Matrix ◽

Flight Muscles

Twelve monoclonal antibodies have been raised against proteins in preparations of Z-disks isolated from Drosophila melanogaster flight muscle. The monoclonal antibodies that recognized Z-band components were identified by immunofluorescence microscopy of flight muscle myofibrils. These antibodies have identified three Z-disk antigens on immunoblots of myofibrillar proteins. Monoclonal antibodies alpha:1-4 recognize a 90-100-kD protein which we identify as alpha-actinin on the basis of cross-reactivity with antibodies raised against honeybee and vertebrate alpha-actinins. Monoclonal antibodies P:1-4 bind to the high molecular mass protein, projectin, a component of connecting filaments that link the ends of thick filaments to the Z-band in insect asynchronous flight muscles. The anti-projectin antibodies also stain synchronous muscle, but, surprisingly, the epitopes here are within the A-bands, not between the A- and Z-bands, as in flight muscle. Monoclonal antibodies Z(210):1-4 recognize a 210-kD protein that has not been previously shown to be a Z-band structural component. A fourth antigen, resolved as a doublet (approximately 400/600 kD) on immunoblots of Drosophila fibrillar proteins, is detected by a cross reacting antibody, Z(400):2, raised against a protein in isolated honeybee Z-disks. On Lowicryl sections of asynchronous flight muscle, indirect immunogold staining has localized alpha-actinin and the 210-kD protein throughout the matrix of the Z-band, projectin between the Z- and A-bands, and the 400/600-kD components at the I-band/Z-band junction. Drosophila alpha-actinin, projectin, and the 400/600-kD components share some antigenic determinants with corresponding honeybee proteins, but no honeybee protein interacts with any of the Z(210) antibodies.

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The Drosophila indirect flight muscle myosin heavy chain isoform is insufficient to transform the jump muscle into a highly stretch-activated muscle type

AJP Cell Physiology ◽

10.1152/ajpcell.00284.2016 ◽

2017 ◽

Vol 312 (2) ◽

pp. C111-C118 ◽

Cited By ~ 4

Author(s):

Cuiping Zhao ◽

Douglas M. Swank

Keyword(s):

Power Generation ◽

Myosin Heavy Chain ◽

Heavy Chain ◽

Flight Muscle ◽

Fold Increase ◽

Isometric Tension ◽

Indirect Flight Muscle ◽

Muscle Type ◽

Myosin Heavy Chain Isoform ◽

Muscle Types

Stretch activation (SA) is a delayed increase in force that enables high power and efficiency from a cyclically contracting muscle. SA exists in various degrees in almost all muscle types. In Drosophila, the indirect flight muscle (IFM) displays exceptionally high SA force production ( FSA), whereas the jump muscle produces only minimal FSA. We previously found that expressing an embryonic (EMB) myosin heavy chain (MHC) isoform in the jump muscle transforms it into a moderately SA muscle type and enables positive cyclical power generation. To investigate whether variation in MHC isoforms is sufficient to produce even higher FSA, we substituted the IFM MHC isoform (IFI) into the jump muscle. Surprisingly, we found that IFI only caused a 1.7-fold increase in FSA, less than half the increase previously observed with EMB, and only at a high Pi concentration, 16 mM. This IFI-induced FSA is much less than what occurs in IFM, relative to isometric tension, and did not enable positive cyclical power generation by the jump muscle. Both isometric tension and FSA of control fibers decreased with increasing Pi concentration. However, for IFI-expressing fibers, only isometric tension decreased. The rate of FSA generation was ~1.5-fold faster for IFI fibers than control fibers, and both rates were Pi dependent. We conclude that MHC isoforms can alter FSA and hence cyclical power generation but that isoforms can only endow a muscle type with moderate FSA. Highly SA muscle types, such as IFM, likely use a different or additional mechanism.

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Isolation and characterization of flightless mutants in Drosophila melanogaster

Development ◽

10.1242/dev.45.1.123 ◽

1978 ◽

Vol 45 (1) ◽

pp. 123-143

Author(s):

Takao Koana ◽

Yoshiki Hotta

Keyword(s):

Drosophila Melanogaster ◽

Flight Muscle ◽

Chromosomal Mapping ◽

Indirect Flight Muscle ◽

Mapping Technique ◽

Isolation And Characterization ◽

Genetic Mosaic ◽

Column Type ◽

Muscle Genes ◽

Development And Differentiation

Since animal behaviour is executed through neuronal circuits including sensory receptors and muscle, genes vital for their development and differentiation must be found among mutants having behavioural anomaly. After mutagenesis with ethyl methanesulphonate (EMS), we screened for X-linked flightless mutants of Drosophila melanogaster by using a column-type flight tester. Approximately 104 individuals were screened and 21 mutant genes were isolated. Chromosomal mapping and complementation experiments revealed that they belong to 15 cistrons randomly located on X chromosome, three cistrons having more than two alleles. Two of the isolated mutants (AtO2 and AtH, which are recessive both behaviourally and morphologically) were analysed with the mosaic fate mapping technique, and both were found to have their primary foci in mesodermal region of blastoderm, suggesting that the genes exert their primary effect in indirect flight muscle. Electronmicroscopic studies on the muscles from four alleles of the AtO2 cistron revealed an abnormality in myofibrillar arrangement. A possible deficit within Z-band components is discussed in relation to wings-up B mutants. The indirect flight muscle of AltH was also examined, and it was found that sarcomere length and diameter of myofibrils were abnormal. It was postulated that a possible factor which controls size of myofibrils is defective in this mutant. These examples indicate the advantage of combining ultrastructural examination with genetic mosaic mapping technique.

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The distribution pattern of indirect flight muscle fibres in a laboratory strain of Aedes togoi (Theobald) (Diptera: Culicidae)

Journal of Entomology Series A General Entomology ◽

10.1111/j.1365-3032.1974.tb00045.x ◽

2009 ◽

Vol 48 (2) ◽

pp. 135-140 ◽

Cited By ~ 2

Author(s):

EVELYN B. BECKETT

Keyword(s):

Distribution Pattern ◽

Flight Muscle ◽

Laboratory Strain ◽

Indirect Flight Muscle ◽

Muscle Fibres ◽

Aedes Togoi

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The continuity of thick filaments between sarcomeres in honey bee flight muscle

Nature ◽

10.1038/281319a0 ◽

1979 ◽

Vol 281 (5729) ◽

pp. 319-320 ◽

Cited By ~ 7

Author(s):

K. Trombitas ◽

A. Tigyi-Sebes

Keyword(s):

Honey Bee ◽

Flight Muscle ◽

Thick Filaments

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Functional and ultrastructural effects of a missense mutation in the indirect flight muscle-specific actin gene of Drosophila melanogaster

Journal of Molecular Biology ◽

10.1016/0022-2836(91)90588-w ◽

1991 ◽

Vol 222 (4) ◽

pp. 963-982 ◽

Cited By ~ 26

Author(s):

John Sparrow ◽

Mary Reedy ◽

Elizabeth Ball ◽

Vassilis Kyrtatas ◽

Justin Molloy ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Missense Mutation ◽

Flight Muscle ◽

Indirect Flight Muscle ◽

Actin Gene

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Ifm(2)2 is a myosin heavy chain allele that disrupts myofibrillar assembly only in the indirect flight muscle of Drosophila melanogaster.

The Journal of Cell Biology ◽

10.1083/jcb.107.6.2613 ◽

1988 ◽

Vol 107 (6) ◽

pp. 2613-2621 ◽

Cited By ~ 38

Author(s):

M Chun ◽

S Falkenthal

Keyword(s):

Myosin Heavy Chain ◽

Heavy Chain ◽

Flight Muscle ◽

Microscopic Analysis ◽

Thick Filament ◽

Indirect Flight Muscle ◽

Chain Gene ◽

Wild Type ◽

Electron Microscopic ◽

Sarcomere Assembly

Using a combination of molecular and genetic techniques we demonstrate that Ifm(2)2 is an allele of the single-copy sarcomeric myosin heavy chain gene. Flies homozygous for this allele accumulate wild-type levels of mRNA and protein in tubular muscle of adults, but fail to accumulate detectable amounts of myosin heavy chain mRNA or protein in the indirect flight muscle. We propose that the mutation interferes with either transcription of the gene or splicing of the primary transcript in the indirect flight muscle and not in other muscle tissues. Biochemical and electron microscopic analysis of flies homozygous for this mutation has revealed that thick filament assembly is abolished in the indirect flight muscle resulting in the instability of wild-type thick filament proteins. In contrast, thin filament and Z disc assembly are marginally affected. We discuss a working hypothesis for sarcomere assembly and define and experimental approach to test the predictions of this proposed pathway for sarcomere assembly.

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