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. 1986 Dec 1;103(6):2163–2171. doi: 10.1083/jcb.103.6.2163

Formation and alignment of Z lines in living chick myotubes microinjected with rhodamine-labeled alpha-actinin

PMCID: PMC2114583  PMID: 3782295

Abstract

We have used fluorescence analogue cytochemistry in conjunction with time lapse recording to study the dynamics of alpha-actinin, a major component of the Z line, during myofibrillogenesis. Rhodamine-labeled alpha-actinin microinjected into living cultured chick skeletal myotubes became localized in discrete cellular structures within 1 h and remained specifically associated with structures for up to 4 d, allowing individual identified structures to be followed during development. In the most immature cells used, alpha-actinin was found in diffuse aggregates, some of which displayed sarcomeric periodicity. Aggregates were observed to coalesce into better defined structures (Z bands) that were approximately 1.0-micron wide. Z bands condensed into narrow, more intensely fluorescent Z lines in 4-48 h. During this period, Z lines grew laterally, primarily by the addition of small beads of alpha-actinin to existing Z lines or by the merging of small Z lines. In more mature cells, alpha-actinin added to Z lines without going through a visible intermediary structure. Mean sarcomere length did not change significantly during the stages examined, although the variability of sarcomere length did decrease markedly over time for identified sets of sarcomeres. At early stages, myofibrils frequently shifted position in both the longitudinal and lateral directions. Neighboring myofibrils were frequently associated for one or more sarcomeres sporadically along their length, such that the intervening sarcomeres were often misaligned. Associations between myofibrils were often transitory. Shifts in myofibril location in conjunction with the formation, breaking, and reformation of lateral associations between myofibrils facilitated the alignment of Z lines through a trial and error process.

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Selected References

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  1. Allen E. R. Sarcomere formation in chick striated muscle. Z Zellforsch Mikrosk Anat. 1973 Nov 29;145(2):167–170. doi: 10.1007/BF00307385. [DOI] [PubMed] [Google Scholar]
  2. Allen R. E., Stromer M. H., Goll D. E., Robson R. M. Accumulation of myosin, actin, tropomyosin, and alpha-actinin in cultured muscles cells. Dev Biol. 1979 Apr;69(2):655–660. doi: 10.1016/0012-1606(79)90318-x. [DOI] [PubMed] [Google Scholar]
  3. Amato P. A., Unanue E. R., Taylor D. L. Distribution of actin in spreading macrophages: a comparative study on living and fixed cells. J Cell Biol. 1983 Mar;96(3):750–761. doi: 10.1083/jcb.96.3.750. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Antin P. B., Tokunaka S., Nachmias V. T., Holtzer H. Role of stress fiber-like structures in assembling nascent myofibrils in myosheets recovering from exposure to ethyl methanesulfonate. J Cell Biol. 1986 Apr;102(4):1464–1479. doi: 10.1083/jcb.102.4.1464. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Busch W. A., Stromer M. H., Goll D. E., Suzuki A. Ca 2+ -specific removal of Z lines from rabbit skeletal muscle. J Cell Biol. 1972 Feb;52(2):367–381. doi: 10.1083/jcb.52.2.367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chowrashi P. K., Pepe F. A. The Z-band: 85,000-dalton amorphin and alpha-actinin and their relation to structure. J Cell Biol. 1982 Sep;94(3):565–573. doi: 10.1083/jcb.94.3.565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Croop J., Toyama Y., Dlugosz A. A., Holtzer H. Selective effects of phorbol 12-myristate 13-acetate on myofibrils and 10-nm filaments. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5273–5277. doi: 10.1073/pnas.77.9.5273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Devlin R. B., Emerson C. P., Jr Coordinate regulation of contractile protein synthesis during myoblast differentiation. Cell. 1978 Apr;13(4):599–611. doi: 10.1016/0092-8674(78)90211-8. [DOI] [PubMed] [Google Scholar]
  9. Doetschman T. C., Eppenberger H. M. Comparison of M-line and other myofibril components during reversible phorbol ester treatment. Eur J Cell Biol. 1984 Mar;33(2):265–274. [PubMed] [Google Scholar]
  10. Fischman D. A. The synthesis and assembly of myofibrils in embryonic muscle. Curr Top Dev Biol. 1970;5:235–280. doi: 10.1016/s0070-2153(08)60057-5. [DOI] [PubMed] [Google Scholar]
  11. Gard D. L., Lazarides E. The synthesis and distribution of desmin and vimentin during myogenesis in vitro. Cell. 1980 Jan;19(1):263–275. doi: 10.1016/0092-8674(80)90408-0. [DOI] [PubMed] [Google Scholar]
  12. Goli D. E., Suzuki A., Temple J., Holmes G. R. Studies on purified -actinin. I. Effect of temperature and tropomyosin on the -actinin-F-actin interaction. J Mol Biol. 1972 Jun 28;67(3):469–488. doi: 10.1016/0022-2836(72)90464-0. [DOI] [PubMed] [Google Scholar]
  13. Hill C. S., Lemanski L. F. Immunoelectron microscopic localization of alpha-actinin and actin in embryonic hamster heart cells. Eur J Cell Biol. 1986 Jan;39(2):300–312. [PubMed] [Google Scholar]
  14. Isobe Y., Shimada Y. Myofibrillogenesis in vitro as seen with the scanning electron microscope. Cell Tissue Res. 1983;231(3):481–494. doi: 10.1007/BF00218107. [DOI] [PubMed] [Google Scholar]
  15. Jockusch H., Jockusch B. M. Structural organization of the Z-line protein, alpha-actinin, in developing skeletal muscle cells. Dev Biol. 1980 Mar;75(1):231–238. doi: 10.1016/0012-1606(80)90158-x. [DOI] [PubMed] [Google Scholar]
  16. Kaneko H., Okamoto M., Goshima K. Structural change of myofibrils during mitosis of newt embryonic myocardial cells in culture. Exp Cell Res. 1984 Aug;153(2):483–498. doi: 10.1016/0014-4827(84)90615-3. [DOI] [PubMed] [Google Scholar]
  17. Kelly D. E. Myofibrillogenesis and Z-band differentiation. Anat Rec. 1969 Mar;163(3):403–425. doi: 10.1002/ar.1091630305. [DOI] [PubMed] [Google Scholar]
  18. Konigsberg I. R. Skeletal myoblasts in culture. Methods Enzymol. 1979;58:511–527. doi: 10.1016/s0076-6879(79)58166-x. [DOI] [PubMed] [Google Scholar]
  19. Kreis T. E., Birchmeier W. Stress fiber sarcomeres of fibroblasts are contractile. Cell. 1980 Nov;22(2 Pt 2):555–561. doi: 10.1016/0092-8674(80)90365-7. [DOI] [PubMed] [Google Scholar]
  20. Markwald R. R. Distribution and relationship of precursor Z material to organizing myofibrillar bundles in embryonic rat and hamster ventricular myocytes. J Mol Cell Cardiol. 1973 Aug;5(4):341–350. doi: 10.1016/0022-2828(73)90026-6. [DOI] [PubMed] [Google Scholar]
  21. Masaki T., Endo M., Ebashi S. Localization of 6S component of a alpha-actinin at Z-band. J Biochem. 1967 Nov;62(5):630–632. doi: 10.1093/oxfordjournals.jbchem.a128717. [DOI] [PubMed] [Google Scholar]
  22. McKenna N. M., Meigs J. B., Wang Y. L. Exchangeability of alpha-actinin in living cardiac fibroblasts and muscle cells. J Cell Biol. 1985 Dec;101(6):2223–2232. doi: 10.1083/jcb.101.6.2223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Meigs J. B., Wang Y. L. Reorganization of alpha-actinin and vinculin induced by a phorbol ester in living cells. J Cell Biol. 1986 Apr;102(4):1430–1438. doi: 10.1083/jcb.102.4.1430. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Morkin E. Postnatal muscle fiber assembly: localization of newly synthesized myofibrillar proteins. Science. 1970 Mar 13;167(3924):1499–1501. doi: 10.1126/science.167.3924.1499. [DOI] [PubMed] [Google Scholar]
  25. Peng H. B., Wolosewick J. J., Cheng P. C. The development of myofibrils in cultured muscle cells: a whole-mount and thin-section electron microscopic study. Dev Biol. 1981 Nov;88(1):121–136. doi: 10.1016/0012-1606(81)90224-4. [DOI] [PubMed] [Google Scholar]
  26. Puri E. C., Caravatti M., Perriard J. C., Turner D. C., Eppenberger H. M. Anchorage-independent muscle cell differentiation. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5297–5301. doi: 10.1073/pnas.77.9.5297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Sanger J. W., Mittal B., Sanger J. M. Analysis of myofibrillar structure and assembly using fluorescently labeled contractile proteins. J Cell Biol. 1984 Mar;98(3):825–833. doi: 10.1083/jcb.98.3.825. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Sanger J. W., Mittal B., Sanger J. M. Formation of myofibrils in spreading chick cardiac myocytes. Cell Motil. 1984;4(6):405–416. doi: 10.1002/cm.970040602. [DOI] [PubMed] [Google Scholar]
  29. Singer R. H., Pudney J. A. Filament-directed intercellular contacts during differentiation of cultured chick myoblasts. Tissue Cell. 1984;16(1):17–29. doi: 10.1016/0040-8166(84)90015-6. [DOI] [PubMed] [Google Scholar]
  30. Taniguchi M., Ishikawa H. In situ reconstitution of myosin filaments within the myosin-extracted myofibril in cultured skeletal muscle cells. J Cell Biol. 1982 Feb;92(2):324–332. doi: 10.1083/jcb.92.2.324. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Tokuyasu K. T., Maher P. A., Singer S. J. Distributions of vimentin and desmin in developing chick myotubes in vivo. I. Immunofluorescence study. J Cell Biol. 1984 Jun;98(6):1961–1972. doi: 10.1083/jcb.98.6.1961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Traeger L., Goldstein M. A. Thin filaments are not of uniform length in rat skeletal muscle. J Cell Biol. 1983 Jan;96(1):100–103. doi: 10.1083/jcb.96.1.100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vandenburgh H. H. Dynamic mechanical orientation of skeletal myofibers in vitro. Dev Biol. 1982 Oct;93(2):438–443. doi: 10.1016/0012-1606(82)90131-2. [DOI] [PubMed] [Google Scholar]
  34. Wang K., Feramisco J. R., Ash J. F. Fluorescent localization of contractile proteins in tissue culture cells. Methods Enzymol. 1982;85(Pt B):514–562. doi: 10.1016/0076-6879(82)85050-7. [DOI] [PubMed] [Google Scholar]

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