Summary
Ubiquitin was localized by immunofluorescence microscopy during post-mating histolysis of fibrillar flight muscle in female fire ants, Solenopsis spp. Normal muscles, as well as histolysing muscles from artificially inseminated and haemolymph-injected females contained ubiquitin in association with nuclei, Z-lines, myofilaments and mitochondria. However, the density of the ubiquitin immunoreaction was markedly increased in the nuclei, Z-lines and mitochondria of degenerating tissues 6, 12 and 24 h post treatment. At these times the heaviest immunoreactivity for ubiquitin was seen in association with the nuclei, Z-lines and mitochondria. Immuno-controls, incubated in the absence of the primary antibody, showed no similar immunostaining. When insemination was preceded by the injection of actinomycin D, muscle degradation was significantly depressed after a 24-h period. Also, ubiquitin immunofluorescence was markedly reduced in tissues pre-treated with actinomycin D. These observations suggest that insemination increases the ubiquitination of specific myofibrillar proteins destined for degradation.
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Ball, E., Karlik, C. C., Beall, C. J., Saville, D. L., Sparrow, J. C., Bullard, B. & Fyrberg, E. A. (1987) Arthrin, a myofibrillar protein of flight muscle, is an actin-ubiquitin conjugate Cell 51, 221–8.
Ciechanover, A., Finely, D. & Varshavsky, A. (1984) The ubiquitin-dependent pathway and mechanisms of energy-dependent intracellular protein degradation. J. Cell Biochem. 24, 27–53.
Davis, W. L., Jones, R. G. & Farmer, G. R. (1989) Insect hemolymph factor promotes histolysis in Solenopsis. Anat. Rec. 224, 473–8.
Davis, W. L., Jacoby, B. H., Jones, R. G. & Goodman, D. B. P. (1993a) Superoxide formation preceding flight muscle degeneration in Solenopsis: fine structural cytochemistry and biochemistry. Histochem. J. 25, 478–490.
Davis, W. L., Hagler, H. K., Jacoby, B. H., Konrad, R. J. & Goodman, D. B. P. (1993b) TNF-a induced changes in the fine structure, ion distribution and oxyradical formation in insect flight muscle. Histochem. J. (submitted for publication).
Goldstein, G., Scheid, M., Hammerling, U., Boyse, E. A., Schelsinger, D. H. & Niall, H. D. (1975) Isolation of a polypeptide that has lymphocyte differentiating properties and is probably present universally in living cells. Proc. Natl Acad. Sci. USA 75, 11–15.
Haas, A. L. & Bright, P. M. (1985) The immunochemical detection and quantitation of intracellular ubiquitin-protein conjugates. J. Biol Chem. 260, 12 464–73.
Herschko, A., Eytan, E., Ciechanover, A. & Haas, A. L. (1982) Immunochemical analysis of the turnover of the ubiquitin-protein conjugates in intact cells: relationship to the breakdown of abnormal proteins. J. Biol. Chem. 257, 13 964–70.
Herschko, A. & Ciechanover, A. (1983) Mechanisms of intracellular protein breakdown. Ann. Rev. Biochem. 51, 335–64.
Hilenski, L. L., Terracio, L., Haas, A. L. & Borg, T. K. (1992) Immunolocalization of ubiquitin conjugates at Z-bands and intercalated discs of rat cardiomyocytes in vitro and in vivo. J. Histochem. Cytochem. 40, 1037–42.
Jones, R. G., Davis, W. L., Hung, A. C. F. & Vinson, S. B. (1978) Insemination-induced histolysis of the flight muscle in fire ants (Solenopsis, spp.): an ultrastructural study. Am. J. Anat. 151, 603–10.
Jones, R. G., Davis, W. L. & Vinson, S. B. (1982) A histochemical and microanalysis study of calcium changes in insect flight muscle degeneration in Solenopsis, the queen fire ant. J. Histochem. Cytochem. 30, 293–304.
Jones, R. G. & Davis, W. L. (1985a) Muscle membranes from early degeneration muscle cell fibers in Solenopsis are leaky to lanthanum: electron microscopy and x-ray analysis. Anat. Rec. 212, 123–8.
Jones, R. G. & Davis, W. L. (1985b) Leupeptin, a protease inhibitor blocks insemination-induced flight muscle histolysis in the fire ant Solenopsis. Tissue Cell 17, 111–6.
Knowles, S. E. & Ballard, F. J. (1976) Selective control of the degradation of normal and aberrant proteins in Reuber H35 hepatoma cells. Biochem. J. 156, 609–17.
Libby, P. & Goldberg, A. L. (1978) Leupeptin, a protease inhibitor, decreases protein degradation in normal and diseased muscles. Science 199, 534–6.
Mortimore, G. E. (1982) Mechanisms of cellular protein catabolism. Nutr. Rev. 40, 1–12.
Obinata, T. (1981) Dynamics of protein turnover in vertebrate skeletal muscle. Muscle Nerve 4, 456–88.
Pringle, J. W. S. (1949) The excitation and contraction of the flight muscles of insects. J. Physiol. 108, 225–32.
Pringle, J. W. S. (1978) Stretch activation of muscle: function and mechanism. Proc. Roy. Soc. Lond. Ser. B 201, 107–31.
Rechsteiner, M. (1988) Ubiquitin. New York: Plenum Press.
Riley, D. A., Bain, J. L. W. & Haas, A. L. (1986) Increased ubiquitin conjugation of proteins during skeletal muscle atrophy. J. Cell Biol. 103, 401A.
Riley, D. A., Bain, J. L. W., Ellis, S. & Haas, A. L. (1988) Quantitation and immunocytochemical localization of ubiquitin conjugates within rat red and white skeletal muscles. J. Histochem. Cytochem. 36, 621–32.
Riley, D. A., Ilyina-Kakueva, E. L., Ellis, S., Bain, J. L.W., Slocum, G. R. & Sedlak, F. R. (1990) Skeletal muscle fibers, nerve, and blood vessel breakdown in space-flown rats. FASEB J. 4, 84–91.
Sanger, J. W., Mittal, B. & Sanger, J. M. (1984) Analysis of myofibrillar structure and assembly using fluorescein labeled contractile proteins. J. Cell Biol. 98, 825–833.
Schwartz, L. M. & Truman, J. W. (1982) Peptide and steroid regulation of muscle degeneration in an insect. Science 215, 1420–1.
Schwartz, L. M. & Truman, J. W. (1983) Hormonal control rates of metamorphic development in the Tobacco Hookworm Manduca sexta. Dev. Biol. 99, 103–14.
Schwartz, L. M., Kosz, L. & Kay, B. K. (1990) Gene activation is required for developmentally programmed cell death. Proc. Natl Acad. Sci. USA 87, 6594–8.
Seufert, W. & Jentsch, S. (1990) Ubiquitin-conjugating enzymes UBC4 and UBC5 mediate selective degradation of short-lived and abnormal proteins. EMBO J. 9, 543–550.
Stracher, A., Mcgowan, E. B. & Shafiq, S. A. (1978) Muscular dystrophy: inhibition of degeneration in vivo with protease inhibitors. Science 200, 50–51.
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Davis, W.L., Jacoby, B.H. & Goodman, D.B.P. Immunolocalization of ubiquitin in degenerating insect flight muscle. Histochem J 26, 298–305 (1994). https://doi.org/10.1007/BF00157762
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DOI: https://doi.org/10.1007/BF00157762