Journal articles: 'Drosophila Troponin I'

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QIU, Feng, Anne LAKEY, Bogos AGIANIAN, Amanda HUTCHINGS, Geoffrey W. BUTCHER, Siegfried LABEIT, Kevin LEONARD, and Belinda BULLARD. "Troponin C in different insect muscle types: identification of two isoforms in Lethocerus, Drosophila and Anopheles that are specific to asynchronous flight muscle in the adult insect." Biochemical Journal 371, no. 3 (May 1, 2003): 811–21. http://dx.doi.org/10.1042/bj20021814.

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The indirect flight muscles (IFMs) of Lethocerus (giant water bug) and Drosophila (fruitfly) are asynchronous: oscillatory contractions are produced by periodic stretches in the presence of a Ca2+ concentration that does not fully activate the muscle. The troponin complex on thin filaments regulates contraction in striated muscle. The complex in IFM has subunits that are specific to this muscle type, and stretch activation may act through troponin. Lethocerus and Drosophila have an unusual isoform of the Ca2+-binding subunit of troponin, troponin C (TnC), with a single Ca2+-binding site near the C-terminus (domain IV); this isoform is only in IFMs, together with a minor isoform with an additional Ca2+-binding site in the N-terminal region (domain II). Lethocerus has another TnC isoform in leg muscle which also has two Ca2+-binding sites. Ca2+ binds more strongly to domain IV than to domain II in two-site isoforms. There are four isoforms in Drosophila and Anopheles (malarial mosquito), three of which are also in adult Lethocerus. A larval isoform has not been identified in Lethocerus. Different TnC isoforms are expressed in the embryonic, larval, pupal and adult stages of Drosophila; the expression of the two IFM isoforms is increased in the pupal stage. Immunoelectron microscopy shows the distribution of the major IFM isoform with one Ca2+-binding site is uniform along Lethocerus thin filaments. We suggest that initial activation of IFM is by Ca2+ binding to troponin with the two-site TnC, and full activation is through the action of stretch on the complex with the one-site isoform.

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Terami, Hiromi, Benjamin D. Williams, Shin-ichi Kitamura, Yasuji Sakube, Shinji Matsumoto, Shima Doi, Takashi Obinata, and Hiroaki Kagawa. "Genomic Organization, Expression, and Analysis of the Troponin C Gene pat-10 of Caenorhabditis elegans." Journal of Cell Biology 146, no. 1 (July 12, 1999): 193–202. http://dx.doi.org/10.1083/jcb.146.1.193.

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We have cloned and characterized the troponin C gene, pat-10 of the nematode Caenorhabditis elegans. At the amino acid level nematode troponin C is most similar to troponin C of Drosophila (45% identity) and cardiac troponin C of vertebrates. Expression studies demonstrate that this troponin is expressed in body wall muscle throughout the life of the animal. Later, vulval muscles and anal muscles also express this troponin C isoform. The structural gene for this troponin is pat-10 and mutations in this gene lead to animals that arrest as twofold paralyzed embryos late in development. We have sequenced two of the mutations in pat-10 and both had identical two mutations in the gene; one changes D64 to N and the other changes W153 to a termination site. The missense alteration affects a calcium-binding site and eliminates calcium binding, whereas the second mutation eliminates binding to troponin I. These combined biochemical and in vivo studies of mutant animals demonstrate that this troponin is essential for proper muscle function during development.

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Mas, José-Antonio, Elena García-Zaragoza, and Margarita Cervera. "Two Functionally Identical Modular Enhancers in Drosophila Troponin T Gene Establish the Correct Protein Levels in Different Muscle Types." Molecular Biology of the Cell 15, no. 4 (April 2004): 1931–45. http://dx.doi.org/10.1091/mbc.e03-10-0729.

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The control of muscle-specific expression is one of the principal mechanisms by which diversity is generated among muscle types. In an attempt to elucidate the regulatory mechanisms that control fiber diversity in any given muscle, we have focused our attention on the transcriptional regulation of the Drosophila Troponin T gene. Two, nonredundant, functionally identical, enhancer-like elements activate Troponin T transcription independently in all major muscles of the embryo and larvae as well as in adult somatic and visceral muscles. Here, we propose that the differential but concerted interaction of these two elements underlies the mechanism by which a particular muscle-type establish the correct levels of Troponin T expression, adapting these levels to their specific needs. This mechanism is not exclusive to the Troponin T gene, but is also relevant to the muscle-specific Troponin I gene. In conjunction with in vivo transgenic studies, an in silico analysis of the Troponin T enhancer-like sequences revealed that both these elements are organized in a modular manner. Extending this analysis to the Troponin I and Tropomyosin regulatory elements, the two other components of the muscle-regulatory complex, we have discovered a similar modular organization of phylogenetically conserved domains.

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Beall, C. J., and E. Fyrberg. "Muscle abnormalities in Drosophila melanogaster heldup mutants are caused by missing or aberrant troponin-I isoforms." Journal of Cell Biology 114, no. 5 (September 1, 1991): 941–51. http://dx.doi.org/10.1083/jcb.114.5.941.

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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.

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Nongthomba, Upendra, Mark Cummins, Samantha Clark, Jim O. Vigoreaux, and John C. Sparrow. "Suppression of Muscle Hypercontraction by Mutations in the Myosin Heavy Chain Gene of Drosophila melanogaster." Genetics 164, no. 1 (May 1, 2003): 209–22. http://dx.doi.org/10.1093/genetics/164.1.209.

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Abstract The indirect flight muscles (IFM) of Drosophila melanogaster provide a good genetic system with which to investigate muscle function. Flight muscle contraction is regulated by both stretch and Ca2+-induced thin filament (actin + tropomyosin + troponin complex) activation. Some mutants in troponin-I (TnI) and troponin-T (TnT) genes cause a “hypercontraction” muscle phenotype, suggesting that this condition arises from defects in Ca2+ regulation and actomyosin-generated tension. We have tested the hypothesis that missense mutations of the myosin heavy chain gene, Mhc, which suppress the hypercontraction of the TnI mutant held-up2 (hdp2), do so by reducing actomyosin force production. Here we show that a “headless” Mhc transgenic fly construct that reduces the myosin head concentration in the muscle thick filaments acts as a dose-dependent suppressor of hypercontracting alleles of TnI, TnT, Mhc, and flightin genes. The data suggest that most, if not all, mutants causing hypercontraction require actomyosin-produced forces to do so. Whether all Mhc suppressors act simply by reducing the force production of the thick filament is discussed with respect to current models of myosin function and thin filament activation by the binding of calcium to the troponin complex.

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Marín, María-Cruz, José-Rodrigo Rodríguez, and Alberto Ferrús. "Transcription of Drosophila Troponin I Gene Is Regulated by Two Conserved, Functionally Identical, Synergistic Elements." Molecular Biology of the Cell 15, no. 3 (March 2004): 1185–96. http://dx.doi.org/10.1091/mbc.e03-09-0663.

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The Drosophila wings-up A gene encodes Troponin I. Two regions, located upstream of the transcription initiation site (upstream regulatory element) and in the first intron (intron regulatory element), regulate gene expression in specific developmental and muscle type domains. Based on LacZ reporter expression in transgenic lines, upstream regulatory element and intron regulatory element yield identical expression patterns. Both elements are required for full expression levels in vivo as indicated by quantitative reverse transcription-polymerase chain reaction assays. Three myocyte enhancer factor-2 binding sites have been functionally characterized in each regulatory element. Using exon specific probes, we show that transvection is based on transcriptional changes in the homologous chromosome and that Zeste and Suppressor of Zeste 3 gene products act as repressors for wings-up A. Critical regions for transvection and for Zeste effects are defined near the transcription initiation site. After in silico analysis in insects (Anopheles and Drosophila pseudoobscura) and vertebrates (Ratus and Coturnix), the regulatory organization of Drosophila seems to be conserved. Troponin I (TnI) is expressed before muscle progenitors begin to fuse, and sarcomere morphogenesis is affected by TnI depletion as Z discs fail to form, revealing a novel developmental role for the protein or its transcripts. Also, abnormal stoichiometry among TnI isoforms, rather than their absolute levels, seems to cause the functional muscle defects.

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Chechenova, Maria B., Sara Maes, Sandy T. Oas, Cloyce Nelson, Kaveh G. Kiani, Anton L. Bryantsev, and Richard M. Cripps. "Functional redundancy and nonredundancy between two Troponin C isoforms inDrosophilaadult muscles." Molecular Biology of the Cell 28, no. 6 (March 15, 2017): 760–70. http://dx.doi.org/10.1091/mbc.e16-07-0498.

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We investigated the functional overlap of two muscle Troponin C (TpnC) genes that are expressed in the adult fruit fly, Drosophila melanogaster: TpnC4 is predominantly expressed in the indirect flight muscles (IFMs), whereas TpnC41C is the main isoform in the tergal depressor of the trochanter muscle (TDT; jump muscle). Using CRISPR/Cas9, we created a transgenic line with a homozygous deletion of TpnC41C and compared its phenotype to a line lacking functional TpnC4. We found that the removal of either of these genes leads to expression of the other isoform in both muscle types. The switching between isoforms occurs at the transcriptional level and involves minimal enhancers located upstream of the transcription start points of each gene. Functionally, the two TpnC isoforms were not equal. Although ectopic TpnC4 in TDT muscles was able to maintain jumping ability, TpnC41C in IFMs could not effectively support flying. Simultaneous functional disruption of both TpnC genes resulted in jump-defective and flightless phenotypes of the survivors, as well as abnormal sarcomere organization. These results indicated that TpnC is required for myofibril assembly, and that there is functional specialization among TpnC isoforms in Drosophila.

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Vicente-Crespo, Marta, Maya Pascual, Juan M. Fernandez-Costa, Amparo Garcia-Lopez, Lidón Monferrer, M. Eugenia Miranda, Lei Zhou, and Ruben D. Artero. "Drosophila Muscleblind Is Involved in troponin T Alternative Splicing and Apoptosis." PLoS ONE 3, no. 2 (February 20, 2008): e1613. http://dx.doi.org/10.1371/journal.pone.0001613.

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Naimi, Benyoussef, Andrew Harrison, Mark Cummins, Upendra Nongthomba, Samantha Clark, Inmaculada Canal, Alberto Ferrus, and John C. Sparrow. "A Tropomyosin-2 Mutation Suppresses a Troponin I Myopathy inDrosophila." Molecular Biology of the Cell 12, no. 5 (May 2001): 1529–39. http://dx.doi.org/10.1091/mbc.12.5.1529.

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A suppressor mutation, D53, of theheld-up 2 allele of the Drosophila melanogaster Troponin I (wupA) gene is described. D53, a missense mutation, S185F, of the tropomyosin-2,Tm2, gene fully suppresses all the phenotypic effects ofheld-up 2, including the destructive hypercontraction of the indirect flight muscles (IFMs), a lack of jumping, the progressive myopathy of the walking muscles, and reductions in larval crawling and feeding behavior. The suppressor restores normal function of the IFMs, but flight ability decreases with age and correlates with an unusual, progressive structural collapse of the myofibrillar lattice starting at the center. The S185F substitution in Tm2 is close to a troponin T binding site on tropomyosin. Models to explain suppression by D53, derived from current knowledge of the vertebrate troponin-tropomyosin complex structure and functions, are discussed. The effects of S185F are compared with those of two mutations in residues 175 and 180 of human α-tropomyosin 1 which cause familial hypertrophic cardiomyopathy (HCM).

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Montana, Enrico S., and J. Troy Littleton. "Characterization of a hypercontraction-induced myopathy in Drosophila caused by mutations in Mhc." Journal of Cell Biology 164, no. 7 (March 29, 2004): 1045–54. http://dx.doi.org/10.1083/jcb.200308158.

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The Myosin heavy chain (Mhc) locus encodes the muscle-specific motor mediating contraction in Drosophila. In a screen for temperature-sensitive behavioral mutants, we have identified two dominant Mhc alleles that lead to a hypercontraction-induced myopathy. These mutants are caused by single point mutations in the ATP binding/hydrolysis domain of Mhc and lead to degeneration of the flight muscles. Electrophysiological analysis in the adult giant fiber flight circuit demonstrates temperature-dependent seizure activity that requires neuronal input, as genetic blockage of neuronal activity suppresses the electrophysiological seizure defects. Intracellular recordings at the third instar neuromuscular junction show spontaneous muscle movements in the absence of neuronal stimulation and extracellular Ca2+, suggesting a dysregulation of intracellular calcium homeostasis within the muscle or an alteration of the Ca2+ dependence of contraction. Characterization of these new Mhc alleles suggests that hypercontraction occurs via a mechanism, which is molecularly distinct from mutants identified previously in troponin I and troponin T.

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Kronert, William A., Angel Acebes, Alberto Ferrús, and Sanford I. Bernstein. "Specific Myosin Heavy Chain Mutations Suppress Troponin I Defects in Drosophila Muscles." Journal of Cell Biology 144, no. 5 (March 8, 1999): 989–1000. http://dx.doi.org/10.1083/jcb.144.5.989.

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We show that specific mutations in the head of the thick filament molecule myosin heavy chain prevent a degenerative muscle syndrome resulting from the hdp2 mutation in the thin filament protein troponin I. One mutation deletes eight residues from the actin binding loop of myosin, while a second affects a residue at the base of this loop. Two other mutations affect amino acids near the site of nucleotide entry and exit in the motor domain. We document the degree of phenotypic rescue each suppressor permits and show that other point mutations in myosin, as well as null mutations, fail to suppress the hdp2 phenotype. We discuss mechanisms by which the hdp2 phenotypes are suppressed and conclude that the specific residues we identified in myosin are important in regulating thick and thin filament interactions. This in vivo approach to dissecting the contractile cycle defines novel molecular processes that may be difficult to uncover by biochemical and structural analysis. Our study illustrates how expression of genetic defects are dependent upon genetic background, and therefore could have implications for understanding gene interactions in human disease.

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Herranz, R. "Expression patterns of the whole Troponin C gene repertoire during Drosophila development." Gene Expression Patterns 4, no. 2 (March 2004): 183–90. http://dx.doi.org/10.1016/j.modgep.2003.09.008.

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SINGH, SALAM HEROJEET, PRABODH KUMAR, NALLUR B. RAMACHANDRA, and UPENDRA NONGTHOMBA. "Roles of the troponin isoforms during indirect flight muscle development in Drosophila." Journal of Genetics 93, no. 2 (August 2014): 379–88. http://dx.doi.org/10.1007/s12041-014-0386-8.

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Fyrberg, Eric, Christine C. Fyrberg, Clifford Beall, and Donna L. Saville. "Drosophila melanogaster troponin-T mutations engender three distinct syndromes of myofibrillar abnormalities." Journal of Molecular Biology 216, no. 3 (December 1990): 657–75. http://dx.doi.org/10.1016/0022-2836(90)90390-8.

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Cao, Tianxin, Alyson Sujkowski, Tyler Cobb, Robert J. Wessells, and Jian-Ping Jin. "The glutamic acid-rich–long C-terminal extension of troponin T has a critical role in insect muscle functions." Journal of Biological Chemistry 295, no. 12 (February 5, 2020): 3794–807. http://dx.doi.org/10.1074/jbc.ra119.012014.

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The troponin complex regulates the Ca2+ activation of myofilaments during striated muscle contraction and relaxation. Troponin genes emerged 500–700 million years ago during early animal evolution. Troponin T (TnT) is the thin-filament–anchoring subunit of troponin. Vertebrate and invertebrate TnTs have conserved core structures, reflecting conserved functions in regulating muscle contraction, and they also contain significantly diverged structures, reflecting muscle type- and species-specific adaptations. TnT in insects contains a highly-diverged structure consisting of a long glutamic acid–rich C-terminal extension of ∼70 residues with unknown function. We found here that C-terminally truncated Drosophila TnT (TpnT–CD70) retains binding of tropomyosin, troponin I, and troponin C, indicating a preserved core structure of TnT. However, the mutant TpnTCD70 gene residing on the X chromosome resulted in lethality in male flies. We demonstrate that this X-linked mutation produces dominant-negative phenotypes, including decreased flying and climbing abilities, in heterozygous female flies. Immunoblot quantification with a TpnT-specific mAb indicated expression of TpnT–CD70 in vivo and normal stoichiometry of total TnT in myofilaments of heterozygous female flies. Light and EM examinations revealed primarily normal sarcomere structures in female heterozygous animals, whereas Z-band streaming could be observed in the jump muscle of these flies. Although TpnT–CD70-expressing flies exhibited lower resistance to cardiac stress, their hearts were significantly more tolerant to Ca2+ overloading induced by high-frequency electrical pacing. Our findings suggest that the Glu-rich long C-terminal extension of insect TnT functions as a myofilament Ca2+ buffer/reservoir and is potentially critical to the high-frequency asynchronous contraction of flight muscles.

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Fyrberg, Christine, Heather Parker, Bernadette Hutchison, and Eric Fyrberg. "Drosophila melanogaster genes encoding three troponin-C isoforms and a calmodulin-related protein." Biochemical Genetics 32, no. 3-4 (April 1994): 119–35. http://dx.doi.org/10.1007/bf00554420.

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PECKHAM, MICHELLE, RICHARD CRIPPS, DAVID WHITE, and BELINDA BULLARD. "Mechanics and Protein Content of Insect Flight Muscles." Journal of Experimental Biology 168, no. 1 (July 1, 1992): 57–76. http://dx.doi.org/10.1242/jeb.168.1.57.

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In asynchronous insect flight muscles, stretch activation may arise from a matching of the helix periodicities of actin target sites to myosin heads and/or a special form of troponin subunit called troponin-H (Tn-H, relative molecular mass 80×103), which has so far only been found in the asynchronous flight muscles of Drosophila (Diptera) and Lethocerus (Hemiptera). The sequence of Tn-H in Drosophila shows it to be a fusion protein of tropomyosin and a hydrophobic proline-rich sequence. Tn-H in Lethocerus is immunologically similar. From immunoblots of synchronous (non-stretch-activated) and asynchronous flight muscles from a wide range of insects, using antibodies against tropomyosin and the hydrophobic sequence of Tn-H, raised against Lethocerus proteins, we found two forms of Tn-H. One, found in most flight muscles, only reacted with antibodies to the hydrophobic sequence. The other, found in asynchronous flight muscles from Diptera and Hemiptera, reacted with antibodies to both the hydrophobic sequence and to tropomyosin, although in the Hemiptera the reaction of Tn-H with anti-tropomyosin was weak. When we compared the mechanics of most of the asynchronous flight muscles used in this study, we found that in the Diptera Ca2+-activated tension was much lower at rest length and stretch-activated tension was more highly dependent on muscle length than in other orders of insects. This suggests that, when Tn-H is in the form found in Diptera, it may be able to modulate Ca2+-activated and stretch-activated tension. We conclude that Tn-H is not sufficient for stretch-activation, but it may enhance stretch-activation, particularly when it is in the form found in the Diptera.

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Barbas, J. A., J. Galceran, I. Krah-Jentgens, J. L. de la Pompa, I. Canal, O. Pongs, and A. Ferrus. "Troponin I is encoded in the haplolethal region of the Shaker gene complex of Drosophila." Genes & Development 5, no. 1 (January 1, 1991): 132–40. http://dx.doi.org/10.1101/gad.5.1.132.

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Prado, A., I. Canal, J. A. Barbas, J. Molloy, and A. Ferrús. "Functional recovery of troponin I in a Drosophila heldup mutant after a second site mutation." Molecular Biology of the Cell 6, no. 11 (November 1995): 1433–41. http://dx.doi.org/10.1091/mbc.6.11.1433.

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To identify proteins that interact in vivo with muscle components we have used a genetic approach based on the isolation of suppressors of mutant alleles of known muscle components. We have applied this system to the case of troponin I (TnI) in Drosophila and its mutant allele heldup2 (hdp2). This mutation causes an alanine to valine substitution at position 116 after a single nucleotide change in a constitutive exon. Among the isolated suppressors, one of them results from a second site mutation at the TnI gene itself. Muscles endowed with TnI mutated at both sites support nearly normal myofibrillar structure, perform notably well in wing beating and flight tests, and isolated muscle fibers produce active force. We show that the structural and functional recovery in this suppressor does not result from a change in the stoichiometric ratio of TnI isoforms. The second site suppression is due to a leucine to phenylalanine change within a heptameric leucine string motif adjacent to the actin binding domain of TnI. These data evidence a structural and functional role for the heptameric leucine string that is most noticeable, if not specific, in the indirect flight muscle.

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Eldred, Catherine C., Anja Katzemich, Georgia Yalanis, Andrea Page-McCaw, Belinda Bullard, and Douglas Swank. "The Influence of Troponin C, Isoform 4 on Drosophila Development, Stretch Activation, and Power Generation." Biophysical Journal 102, no. 3 (January 2012): 155a. http://dx.doi.org/10.1016/j.bpj.2011.11.849.

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Eldred, Catherine C., Anja Katzemich, Monica Patel, Belinda Bullard, and Douglas M. Swank. "The roles of troponin C isoforms in the mechanical function of Drosophila indirect flight muscle." Journal of Muscle Research and Cell Motility 35, no. 3-4 (August 2014): 211–23. http://dx.doi.org/10.1007/s10974-014-9387-8.

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Glasheen, Bernadette M., Catherine C. Eldred, Leah C. Sullivan, Cuiping Zhao, Michael K. Reedy, Robert J. Edwards, and Douglas M. Swank. "Stretch activation properties of Drosophila and Lethocerus indirect flight muscle suggest similar calcium-dependent mechanisms." American Journal of Physiology-Cell Physiology 313, no. 6 (December 1, 2017): C621—C631. http://dx.doi.org/10.1152/ajpcell.00110.2017.

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Muscle stretch activation (SA) is critical for optimal cardiac and insect indirect flight muscle (IFM) power generation. The SA mechanism has been investigated for decades with many theories proposed, but none proven. One reason for the slow progress could be that multiple SA mechanisms may have evolved in multiple species or muscle types. Laboratories studying IFM SA in the same or different species have reported differing SA functional properties which would, if true, suggest divergent mechanisms. However, these conflicting results might be due to different experimental methodologies. Thus, we directly compared SA characteristics of IFMs from two SA model systems, Drosophila and Lethocerus, using two different fiber bathing solutions. Compared with Drosophila IFM, Lethocerus IFM isometric tension is 10- or 17-fold higher and SA tension was 5- or 10-fold higher, depending on the bathing solution. However, the rate of SA tension generation was 9-fold faster for Drosophila IFM. The inverse differences between rate and tension in the two species causes maximum power output to be similar, where Drosophila power is optimized in the bathing solution that favors faster muscle kinetics and Lethocerus in the solution that favors greater tension generation. We found that isometric tension and SA tension increased with calcium concentration for both species in both solutions, reaching a maximum plateau around pCa 5.0. Our results favor a similar mechanism for both species, perhaps involving a troponin complex that does not fully calcium activate the thin filament thus leaving room for further tension generation by SA.

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MARCO-FERRERES, Raquel, Juan J. ARREDONDO, Benito FRAILE, and Margarita CERVERA. "Overexpression of troponin T in Drosophila muscles causes a decrease in the levels of thin-filament proteins." Biochemical Journal 386, no. 1 (February 8, 2005): 145–52. http://dx.doi.org/10.1042/bj20041240.

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Formation of the contractile apparatus in muscle cells requires co-ordinated activation of several genes and the proper assembly of their products. To investigate the role of TnT (troponin T) in the mechanisms that control and co-ordinate thin-filament formation, we generated transgenic Drosophila lines that overexpress TnT in their indirect flight muscles. All flies that overexpress TnT were unable to fly, and the loss of thin filaments themselves was coupled with ultrastructural perturbations of the sarcomere. In contrast, thick filaments remained largely unaffected. Biochemical analysis of these lines revealed that the increase in TnT levels could be detected only during the early stages of adult muscle formation and was followed by a profound decrease in the amount of this protein as well as that of other thin-filament proteins such as tropomyosin, troponin I and actin. The decrease in thin-filament proteins is not only due to degradation but also due to a decrease in their synthesis, since accumulation of their mRNA transcripts was also severely diminished. This decrease in expression levels of the distinct thin-filament components led us to postulate that any change in the amount of TnT transcripts might trigger the down-regulation of other co-regulated thin-filament components. Taken together, these results suggest the existence of a mechanism that tightly co-ordinates the expression of thin-filament genes and controls the correct stoichiometry of these proteins. We propose that the high levels of unassembled protein might act as a sensor in this process.

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Nongthomba, Upendra, Maqsood Ansari, Divesh Thimmaiya, Meg Stark, and John Sparrow. "Aberrant Splicing of an Alternative Exon in the Drosophila Troponin-T Gene Affects Flight Muscle Development." Genetics 177, no. 1 (July 1, 2007): 295–306. http://dx.doi.org/10.1534/genetics.106.056812.

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Gajewski, Kathleen M., Jianbo Wang, and Robert A. Schulz. "Calcineurin function is required for myofilament formation and troponin I isoform transition in Drosophila indirect flight muscle." Developmental Biology 289, no. 1 (January 2006): 17–29. http://dx.doi.org/10.1016/j.ydbio.2005.09.039.

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Casas-Tintó, Sergio, and Alberto Ferrús. "The haplolethality paradox of the wupA gene in Drosophila." PLOS Genetics 17, no. 3 (March 19, 2021): e1009108. http://dx.doi.org/10.1371/journal.pgen.1009108.

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Haplolethals (HL) are regions of diploid genomes that in one dose are fatal for the organism. Their biological meaning is obscure because heterozygous loss-of-function mutations result in dominant lethality (DL) and, consequently, should be under strong negative selection. We report an in depth study of the HL associated to the gene wings up A (wupA). It encodes 13 transcripts (A-M) that yield 11 protein isoforms (A-K) of Troponin I (TnI). They are functionally diverse in their control of muscle contraction, cell polarity and cell proliferation. Isoform K transfers to the nucleus where it increases transcription of the cell proliferation related genes CDK2, CDK4, Rap and Rab5. The nuclear translocation of isoform K is prevented by the co-expression of A or B isoforms, which illustrates isoform interactions. The corresponding DL mutations are, either DNA rearrangements clustered towards the gene 3’ end, thus affecting the genomic organization of all transcripts, or CRISPR-induced mutations in one of the two ATG sites which eliminate a subset of wupA products. The joint elimination of isoforms C, F, G and H, however, do not cause DL phenotypes. Genetically driven expression of single isoforms rescue neither DL nor any of the mutants known in the gene, suggesting that normal function requires properly regulated expression of specific combinations, rather than single, TnI isoforms. We conclude that the wupA associated HL results from the combined haploinsufficiency of a large set of TnI isoforms. The qualitative and quantitative normal expression of which, requires the chromosomal integrity of the wupA genomic region. Since all fly TnI isoforms are encoded in the same gene, its HL condition becomes unavoidable. These wupA features are comparable to those of dpp, the only other HL studied to some extent, and reveal a scenario of strict dosage dependence with implications for gene expression regulation and splitting.

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Cozhimuttam Viswanathan, Meera, William Lehman, and Anthony Cammarato. "A Troponin-T Mutation Initiates Cardiac and Skeletal Myopathy due to Impaired Inhibition of Contraction in Drosophila Melanogaster." Biophysical Journal 104, no. 2 (January 2013): 311a. http://dx.doi.org/10.1016/j.bpj.2012.11.1727.

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Prado, Antonio, Inmaculada Canal, and Alberto Ferrús. "The Haplolethal Region at the 16F Gene Cluster of Drosophila melanogaster: Structure and Function." Genetics 151, no. 1 (January 1, 1999): 163–75. http://dx.doi.org/10.1093/genetics/151.1.163.

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Abstract Extensive aneuploid analyses had shown the existence of a few haplolethal (HL) regions and one triplolethal region in the genome of Drosophila melanogaster. Since then, only two haplolethals, 22F1-2 and 16F, have been directly linked to identified genes, dpp and wupA, respectively. However, with the possible exception of dpp, the actual bases for this dosage sensitivity remain unknown. We have generated and characterized dominant-lethal mutations and chromosomal rearrangements in 16F and studied them in relation to the genes in the region. This region extends along 100 kb and includes at least 14 genes. The normal HL function depends on the integrity of a critical 4-kb window of mostly noncoding sequences within the wupA transcription unit that encodes the muscle protein troponin I (TNI). All dominant lethals are breakpoints within that window, which prevent the functional expression of TNI and other adjacent genes in the proximal direction. However, independent mutations in these genes result in recessive lethal phenotypes only. We propose that the HL at 16F represents a long-range cis regulatory region that acts upon a number of functionally related genes whose combined haploidy would yield the dominant-lethal effect.

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Barbas, J. A., J. Galceran, L. Torroja, A. Prado, and A. Ferrús. "Abnormal muscle development in the heldup3 mutant of Drosophila melanogaster is caused by a splicing defect affecting selected troponin I isoforms." Molecular and Cellular Biology 13, no. 3 (March 1993): 1433–39. http://dx.doi.org/10.1128/mcb.13.3.1433-1439.1993.

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The troponin I (TnI) gene of Drosophila melanogaster encodes a family of 10 isoforms resulting from the differential splicing of 13 exons. Four of these exons (6a1, 6a2, 6b1, and 6b2) are mutually exclusive and very similar in sequence. TnI isoforms show qualitative specificity whereby each muscle expresses a selected repertoire of them. In addition, TnI isoforms show quantitative specificity whereby each muscle expresses characteristic amounts of each isoform. In the mutant heldup3, the development of the thoracic muscles DLM, DVM, and TDT is aborted. The mutation consists of a one-nucleotide displacement of the 3' AG splice site at the intron preceding exon 6b1, resulting in the failure to produce all exon 6b1-containing TnI isoforms. These molecular changes in a constituent of the thin filaments cause the selective failure to develop the DLM, DVM, and TDT muscles while having no visible effect on other muscles wherein exon 6b1 expression is minor.

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30

Barbas, J. A., J. Galceran, L. Torroja, A. Prado, and A. Ferrús. "Abnormal muscle development in the heldup3 mutant of Drosophila melanogaster is caused by a splicing defect affecting selected troponin I isoforms." Molecular and Cellular Biology 13, no. 3 (March 1993): 1433–39. http://dx.doi.org/10.1128/mcb.13.3.1433.

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The troponin I (TnI) gene of Drosophila melanogaster encodes a family of 10 isoforms resulting from the differential splicing of 13 exons. Four of these exons (6a1, 6a2, 6b1, and 6b2) are mutually exclusive and very similar in sequence. TnI isoforms show qualitative specificity whereby each muscle expresses a selected repertoire of them. In addition, TnI isoforms show quantitative specificity whereby each muscle expresses characteristic amounts of each isoform. In the mutant heldup3, the development of the thoracic muscles DLM, DVM, and TDT is aborted. The mutation consists of a one-nucleotide displacement of the 3' AG splice site at the intron preceding exon 6b1, resulting in the failure to produce all exon 6b1-containing TnI isoforms. These molecular changes in a constituent of the thin filaments cause the selective failure to develop the DLM, DVM, and TDT muscles while having no visible effect on other muscles wherein exon 6b1 expression is minor.

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Wang, Shuoshuo, Elizabeth Stoops, Unnikannan CP, Barak Markus, Adriana Reuveny, Elly Ordan, and Talila Volk. "Mechanotransduction via the LINC complex regulates DNA replication in myonuclei." Journal of Cell Biology 217, no. 6 (April 12, 2018): 2005–18. http://dx.doi.org/10.1083/jcb.201708137.

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Nuclear mechanotransduction has been implicated in the control of chromatin organization; however, its impact on functional contractile myofibers is unclear. We found that deleting components of the linker of nucleoskeleton and cytoskeleton (LINC) complex in Drosophila melanogaster larval muscles abolishes the controlled and synchronized DNA endoreplication, typical of nuclei across myofibers, resulting in increased and variable DNA content in myonuclei of individual myofibers. Moreover, perturbation of LINC-independent mechanical input after knockdown of β-Integrin in larval muscles similarly led to increased DNA content in myonuclei. Genome-wide RNA-polymerase II occupancy analysis in myofibers of the LINC mutant klar indicated an altered binding profile, including a significant decrease in the chromatin regulator barrier-to-autointegration factor (BAF) and the contractile regulator Troponin C. Importantly, muscle-specific knockdown of BAF led to increased DNA content in myonuclei, phenocopying the LINC mutant phenotype. We propose that mechanical stimuli transmitted via the LINC complex act via BAF to regulate synchronized cell-cycle progression of myonuclei across single myofibers.

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Firdaus, H., J. Mohan, S. Naz, P. Arathi, S. R. Ramesh, and U. Nongthomba. "A cis-Regulatory Mutation in Troponin-I of Drosophila Reveals the Importance of Proper Stoichiometry of Structural Proteins During Muscle Assembly." Genetics 200, no. 1 (March 5, 2015): 149–65. http://dx.doi.org/10.1534/genetics.115.175604.

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Chechenova, Maria B., Sara Maes, and Richard M. Cripps. "Expression of the Troponin C at 41C Gene in Adult Drosophila Tubular Muscles Depends upon Both Positive and Negative Regulatory Inputs." PLOS ONE 10, no. 12 (December 7, 2015): e0144615. http://dx.doi.org/10.1371/journal.pone.0144615.

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Nikonova, Elena, Amartya Mukherjee, Ketaki Kamble, Christiane Barz, Upendra Nongthomba, and Maria L. Spletter. "Rbfox1 is required for myofibril development and maintaining fiber type–specific isoform expression in Drosophila muscles." Life Science Alliance 5, no. 4 (January 7, 2022): e202101342. http://dx.doi.org/10.26508/lsa.202101342.

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Protein isoform transitions confer muscle fibers with distinct properties and are regulated by differential transcription and alternative splicing. RNA-binding Fox protein 1 (Rbfox1) can affect both transcript levels and splicing, and is known to contribute to normal muscle development and physiology in vertebrates, although the detailed mechanisms remain obscure. In this study, we report that Rbfox1 contributes to the generation of adult muscle diversity in Drosophila. Rbfox1 is differentially expressed among muscle fiber types, and RNAi knockdown causes a hypercontraction phenotype that leads to behavioral and eclosion defects. Misregulation of fiber type–specific gene and splice isoform expression, notably loss of an indirect flight muscle–specific isoform of Troponin-I that is critical for regulating myosin activity, leads to structural defects. We further show that Rbfox1 directly binds the 3′-UTR of target transcripts, regulates the expression level of myogenic transcription factors myocyte enhancer factor 2 and Salm, and both modulates expression of and genetically interacts with the CELF family RNA-binding protein Bruno1 (Bru1). Rbfox1 and Bru1 co-regulate fiber type–specific alternative splicing of structural genes, indicating that regulatory interactions between FOX and CELF family RNA-binding proteins are conserved in fly muscle. Rbfox1 thus affects muscle development by regulating fiber type–specific splicing and expression dynamics of identity genes and structural proteins.

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Schultheiss, T. M., S. Xydas, and A. B. Lassar. "Induction of avian cardiac myogenesis by anterior endoderm." Development 121, no. 12 (December 1, 1995): 4203–14. http://dx.doi.org/10.1242/dev.121.12.4203.

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An experimental system was devised to study the mechanisms by which cells become committed to the cardiac myocyte lineage during avian development. Chick tissues from outside the fate map of the heart (in the posterior primitive streak (PPS) of a Hamburger & Hamilton stage 4 embryo) were combined with potential inducing tissues from quail embryos and cultured in vitro. Species-specific RT-PCR was employed to detect the appearance of the cardiac muscle markers chick Nkx-2.5 (cNkx-2.5), cardiac troponin C and ventricular myosin heavy chain in the chick responder tissues. Using this procedure, we found that stage 4–5 anterior lateral (AL) endoderm and anterior central (AC) mesendoderm, but not AL mesoderm or posterior lateral mesendoderm, induced cells of the PPS to differentiate as cardiac myocytes. Induction of cardiogenesis was accompanied by a marked decrease in the expression of rho-globin, implying that PPS cells were being induced by anterior endoderm to become cardiac myocytes instead of blood-forming tissue. These results suggest that anterior endoderm contains signaling molecules that can induce cardiac myocyte specification of early primitive streak cells. One of the cardiac muscle markers induced by anterior endoderm, cNkx-2.5, is here described for the first time. cNkx-2.5 is a chick homeobox-containing gene that shares extensive sequence similarity with the Drosophila gene tinman, which is required for Drosophila heart formation. The mesodermal component of cNkx-2.5 expression from stage 5 onward, as determined by in situ hybridization, is strikingly in accord with the fate map of the avian heart. By the time the myocardium and endocardium form distinct layers, cNkx-2.5 is found only in the myocardium. cNkx-2.5 thus appears to be the earliest described marker of avian mesoderm fated to give rise to cardiac muscle.

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Coulter, L. R., M. A. Landree, and T. A. Cooper. "Identification of a new class of exonic splicing enhancers by in vivo selection." Molecular and Cellular Biology 17, no. 4 (April 1997): 2143–50. http://dx.doi.org/10.1128/mcb.17.4.2143.

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In vitro selection strategies have typically been used to identify a preferred ligand, usually an RNA, for an identified protein. Ideally, one would like to know RNA consensus sequences preferred in vivo for as-yet-unidentified factors. The ability to select RNA-processing signals would be particularly beneficial in the analysis of exon enhancer sequences that function in exon recognition during pre-mRNA splicing. Exon enhancers represent a class of potentially ubiquitous RNA-processing signals whose actual prevalence is unknown. To establish an approach for in vivo selection, we developed an iterative scheme to select for exon sequences that enhance exon inclusion. This approach is modeled on the in vitro SELEX procedure and uses transient transfection in an iterative procedure to enrich RNA-processing signals in cultured vertebrate cells. Two predominant sequence motifs were enriched after three rounds of selection: a purine-rich motif that resembles previously identified splicing enhancers and a class of A/C-rich splicing enhancers (ACEs). Individual selected ACEs enhanced splicing in vivo and in vitro. ACE splicing activity was competed by RNAs containing the purine-rich splicing enhancer from cardiac troponin T exon 5. Thus, ACE activity is likely to require a subset of the SR splicing factors previously shown to mediate activity of this purine-rich enhancer. ACE motifs are found in two vertebrate exons previously demonstrated to contain splicing enhancer activity as well as in the well-characterized Drosophila doublesex (dsx) splicing enhancer. We demonstrate that one copy of the dsx repeat enhances splicing of a vertebrate exon in vertebrate cells and that this enhancer activity requires the ACE motif. We suggest the possibility that the dsx enhancer is a member of a previously unrecognized family of ACEs.

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Strehler, E. E., M. Periasamy, M. A. Strehler-Page, and B. Nadal-Ginard. "Myosin light-chain 1 and 3 gene has two structurally distinct and differentially regulated promoters evolving at different rates." Molecular and Cellular Biology 5, no. 11 (November 1985): 3168–82. http://dx.doi.org/10.1128/mcb.5.11.3168-3182.1985.

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DNA fragments located 10 kilobases apart in the genome and containing, respectively, the first myosin light chain 1 (MLC1f) and the first myosin light chain 3 (MLC3f) specific exon of the rat myosin light chain 1 and 3 gene, together with several hundred base pairs of upstream flanking sequences, have been shown in runoff in vitro transcription assays to direct initiation of transcription at the cap sites of MLC1f and MLC3f mRNAs used in vivo. These results establish the presence of two separate, functional promoters within that gene. A comparison of the nucleotide sequence of the rat MLC1f/3f gene with the corresponding sequences from mouse and chicken shows that: the MLC1f promoter regions have been highly conserved up to position -150 from the cap site while the MLC3f promoter regions display a very poor degree of homology and even the absence or poor conservation of typical eucaryotic promoter elements such as TATA and CAT boxes; the exon/intron structure of this gene has been completely conserved in the three species; and corresponding exons, except for the regions encoding most of the 5' and 3' untranslated sequences, show greater than 75% homology while corresponding introns are similar in size but considerably divergent in sequence. The above findings indicate that the overall structure of the MLC1f/3f genes has been maintained between avian and mammalian species and that these genes contain two functional and widely spaced promoters. The fact that the structures of the alkali light chain gene from Drosophila melanogaster and of other related genes of the troponin C supergene family resemble a MLC3f gene without an upstream promoter and first exon strongly suggests that the present-day MLC1f/3f genes of higher vertebrates arose from a primordial alkali light chain gene through the addition of a far-upstream MLC1f-specific promoter and first exon. The two promoters have evolved at different rates, with the MLC1f promoter being more conserved than the MLC3f promoter. This discrepant evolutionary rate might reflect different mechanisms of promoter activation for the transcription of MLC1f and MLC3f RNA.

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Strehler, E. E., M. Periasamy, M. A. Strehler-Page, and B. Nadal-Ginard. "Myosin light-chain 1 and 3 gene has two structurally distinct and differentially regulated promoters evolving at different rates." Molecular and Cellular Biology 5, no. 11 (November 1985): 3168–82. http://dx.doi.org/10.1128/mcb.5.11.3168.

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DNA fragments located 10 kilobases apart in the genome and containing, respectively, the first myosin light chain 1 (MLC1f) and the first myosin light chain 3 (MLC3f) specific exon of the rat myosin light chain 1 and 3 gene, together with several hundred base pairs of upstream flanking sequences, have been shown in runoff in vitro transcription assays to direct initiation of transcription at the cap sites of MLC1f and MLC3f mRNAs used in vivo. These results establish the presence of two separate, functional promoters within that gene. A comparison of the nucleotide sequence of the rat MLC1f/3f gene with the corresponding sequences from mouse and chicken shows that: the MLC1f promoter regions have been highly conserved up to position -150 from the cap site while the MLC3f promoter regions display a very poor degree of homology and even the absence or poor conservation of typical eucaryotic promoter elements such as TATA and CAT boxes; the exon/intron structure of this gene has been completely conserved in the three species; and corresponding exons, except for the regions encoding most of the 5' and 3' untranslated sequences, show greater than 75% homology while corresponding introns are similar in size but considerably divergent in sequence. The above findings indicate that the overall structure of the MLC1f/3f genes has been maintained between avian and mammalian species and that these genes contain two functional and widely spaced promoters. The fact that the structures of the alkali light chain gene from Drosophila melanogaster and of other related genes of the troponin C supergene family resemble a MLC3f gene without an upstream promoter and first exon strongly suggests that the present-day MLC1f/3f genes of higher vertebrates arose from a primordial alkali light chain gene through the addition of a far-upstream MLC1f-specific promoter and first exon. The two promoters have evolved at different rates, with the MLC1f promoter being more conserved than the MLC3f promoter. This discrepant evolutionary rate might reflect different mechanisms of promoter activation for the transcription of MLC1f and MLC3f RNA.

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Eldred, Catherine J., Laura Koppes, Kevin Georgek, Andrea Page-McCaw, Belinda Bullard, and Douglas M. Swank. "The Influence of Troponin C Isoforms on Drosophils Stretch Activation and Power Generation." Biophysical Journal 100, no. 3 (February 2011): 113a. http://dx.doi.org/10.1016/j.bpj.2010.12.824.

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Singh, Salam Herojeet, Prabodh Kumar, Nallur B. Ramachandra, and Upendra Nongthomba. "Correction to: Roles of the troponin isoforms during indirect flight muscle development in Drosophila." Journal of Genetics 98, no. 5 (November 14, 2019). http://dx.doi.org/10.1007/s12041-019-1150-x.

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Journal articles on the topic 'Drosophila Troponin I'

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