Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1986 Dec 1;103(6):2185–2196. doi: 10.1083/jcb.103.6.2185

Titin and myosin, but not desmin, are linked during myofibrillogenesis in postmitotic mononucleated myoblasts

PMCID: PMC2114608  PMID: 3536962

Abstract

Monoclonal antibodies specific for the muscle protein titin have been used in conjunction with muscle-specific antibodies against myofibrillar myosin heavy chains (MHCs) and desmin to study myogenesis in cultured cells. Desmin synthesis is initiated in replicating presumptive myoblasts, whereas the synthesis of titin and MHC is initiated simultaneously in their progeny, the postmitotic, mononucleated myoblasts. Both titin and MHC are briefly localized to nonstriated and thereafter to definitively striated myofibrils. At no stage during myofibrillogenesis is either protein observed as part of a sequence of mini-sarcomeres. Titin antibodies bind to the A-I junction, MHC antibodies to the A bands in nascent, maturing, and mature myofibrils. In contrast, desmin remains distributed as longitudinal filaments until well after the definitive myofibrils have aligned laterally. This tight temporal and topographical linkage between titin and myosin is also observed in postmitotic, mononucleated myoblasts and multinucleated myotubes when myofibrillogenesis is perturbed with Colcemid or taxol. Colcemid induces elongating postmitotic mononucleated myoblasts and multinucleated myotubes to round up and form Colcemid myosacs. The myofibrils that emerge in these rounded cells are deployed in convoluted circles. The time required for their nonstriated myofibrils to transform into striated myofibrils is greatly protracted. Furthermore, as Colcemid induces immense desmin intermediate filament cables, the normal spatial relationships between emerging individual myofibrils is distorted. Despite these disturbances at all stages, the characteristic temporal and spatial relationship observed in normal myofibrils between titin and MHC is observed in myofibrils assembling in Colcemid-treated cells. Newly born postmitotic mononucleated myoblasts, or maturing myotubes, reared in taxol acquire a star-shaped configuration and are induced to assemble "pseudo- striated myofibrils." Pseudo-striated myofibrils consist of laterally aggregated 1.6-micron long, thick filaments that interdigitate, not with thin filaments, but with long microtubules. These atypical myofibrils lack Z bands. Despite the absence of thin filaments and Z bands, titin localizes with its characteristics sarcomeric periodicity in pseudo-striated myofibrils. We conclude that the initiation and subsequent regulation of titin and myosin synthesis, and their spatial deployment within developing sarcomeres are tightly coupled events. These findings are discussed in terms of a model that proposes interaction between two relatively autonomous "organizing centers" in the assembly of each sarcomere.

Full Text

The Full Text of this article is available as a PDF (4.3 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Altmannsberger M., Osborn M., Treuner J., Hölscher A., Weber K., Shauer A. Diagnosis of human childhood rhabdomyosarcoma of antibodies to desmin, the structural protein of muscle specific intermediate filaments. Virchows Arch B Cell Pathol Incl Mol Pathol. 1982;39(2):203–215. doi: 10.1007/BF02892848. [DOI] [PubMed] [Google Scholar]
  2. Altmannsberger M., Weber K., Droste R., Osborn M. Desmin is a specific marker for rhabdomyosarcomas of human and rat origin. Am J Pathol. 1985 Jan;118(1):85–95. [PMC free article] [PubMed] [Google Scholar]
  3. Antin P. B., Forry-Schaudies S., Friedman T. M., Tapscott S. J., Holtzer H. Taxol induces postmitotic myoblasts to assemble interdigitating microtubule-myosin arrays that exclude actin filaments. J Cell Biol. 1981 Aug;90(2):300–308. doi: 10.1083/jcb.90.2.300. [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. Bennett G. S., Fellini S. A., Holtzer H. Immunofluorescent visualization of 100 A filaments in different cultured chick embryo cell types. Differentiation. 1978;12(2):71–82. doi: 10.1111/j.1432-0436.1979.tb00992.x. [DOI] [PubMed] [Google Scholar]
  6. Bennett G. S., Fellini S. A., Toyama Y., Holtzer H. Redistribution of intermediate filament subunits during skeletal myogenesis and maturation in vitro. J Cell Biol. 1979 Aug;82(2):577–584. doi: 10.1083/jcb.82.2.577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chi J. C., Fellini S. A., Holtzer H. Differences among myosins synthesized in non-myogenic cells, presumptive myoblasts, and myoblasts. Proc Natl Acad Sci U S A. 1975 Dec;72(12):4999–5003. doi: 10.1073/pnas.72.12.4999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chi J. C., Rubinstein N., Strahs K., Holtzer H. Synthesis of myosin heavy and light chains in muscle cultures. J Cell Biol. 1975 Dec;67(3):523–537. doi: 10.1083/jcb.67.3.523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Croop J., Holtzer H. Response of myogenic and fibrogenic cells to cytochalasin B and to colcemid. I. Light microscope observations. J Cell Biol. 1975 May;65(2):271–285. doi: 10.1083/jcb.65.2.271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dlugosz A. A., Antin P. B., Nachmias V. T., Holtzer H. The relationship between stress fiber-like structures and nascent myofibrils in cultured cardiac myocytes. J Cell Biol. 1984 Dec;99(6):2268–2278. doi: 10.1083/jcb.99.6.2268. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dlugosz A. A., Tapscott S. J., Holtzer H. Effects of phorbol 12-myristate 13-acetate on the differentiation program of embryonic chick skeletal myoblasts. Cancer Res. 1983 Jun;43(6):2780–2789. [PubMed] [Google Scholar]
  12. Endo T., Masaki T. Differential expression and distribution of chicken skeletal- and smooth-muscle-type alpha-actinins during myogenesis in culture. J Cell Biol. 1984 Dec;99(6):2322–2332. doi: 10.1083/jcb.99.6.2322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fellini S. A., Bennett G. S., Holtzer H. Selective binding of antibody against gizzard 10-nm filaments to different cell types in myogenic cultures. Am J Anat. 1978 Nov;153(3):451–457. doi: 10.1002/aja.1001530308. [DOI] [PubMed] [Google Scholar]
  14. Forry-Schaudies S., Murray J. M., Toyama Y., Holtzer H. Effects of colcemid and taxol on microtubules and intermediate filaments in chick embryo fibroblasts. Cell Motil Cytoskeleton. 1986;6(3):324–338. doi: 10.1002/cm.970060309. [DOI] [PubMed] [Google Scholar]
  15. Grove B. K., Cerny L., Perriard J. C., Eppenberger H. M. Myomesin and M-protein: expression of two M-band proteins in pectoral muscle and heart during development. J Cell Biol. 1985 Oct;101(4):1413–1421. doi: 10.1083/jcb.101.4.1413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gruen L. C., King N. L., Kurth L., McKenzie L. J. Studies on the structure of connectin in muscle. Int J Pept Protein Res. 1982 Nov;20(5):401–407. doi: 10.1111/j.1399-3011.1982.tb03059.x. [DOI] [PubMed] [Google Scholar]
  17. HOLTZER H., MARSHALL J. M., Jr, FINCK H. An analysis of myogenesis by the use of fluorescent antimyosin. J Biophys Biochem Cytol. 1957 Sep 25;3(5):705–724. doi: 10.1083/jcb.3.5.705. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hill C., Weber K. Monoclonal antibodies distinguish titins from heart and skeletal muscle. J Cell Biol. 1986 Mar;102(3):1099–1108. doi: 10.1083/jcb.102.3.1099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Holtzer H., Bennett G. S., Tapscott S. J., Croop J. M., Toyama Y. Intermediate-size filaments: changes in synthesis and distribution in cells of the myogenic and neurogenic lineages. Cold Spring Harb Symp Quant Biol. 1982;46(Pt 1):317–329. doi: 10.1101/sqb.1982.046.01.033. [DOI] [PubMed] [Google Scholar]
  20. Holtzer H., Croop J., Dienstman S., Ishikawa H., Somlyo A. P. Effects of cytochaslasin B and colcemide on myogenic cultures. Proc Natl Acad Sci U S A. 1975 Feb;72(2):513–517. doi: 10.1073/pnas.72.2.513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Holtzer H., Forry-Schaudies S., Dlugosz A., Antin P., Dubyak G. Interactions between IFs, microtubules, and myofibrils in fibrogenic and myogenic cells. Ann N Y Acad Sci. 1985;455:106–125. doi: 10.1111/j.1749-6632.1985.tb50407.x. [DOI] [PubMed] [Google Scholar]
  22. Ishikawa H., Bischoff R., Holtzer H. Mitosis and intermediate-sized filaments in developing skeletal muscle. J Cell Biol. 1968 Sep;38(3):538–555. doi: 10.1083/jcb.38.3.538. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Lazarides E. Intermediate filaments as mechanical integrators of cellular space. Nature. 1980 Jan 17;283(5744):249–256. doi: 10.1038/283249a0. [DOI] [PubMed] [Google Scholar]
  25. Maruyama K., Yoshioka T., Higuchi H., Ohashi K., Kimura S., Natori R. Connectin filaments link thick filaments and Z lines in frog skeletal muscle as revealed by immunoelectron microscopy. J Cell Biol. 1985 Dec;101(6):2167–2172. doi: 10.1083/jcb.101.6.2167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Moss P. S., Strohman R. C. Myosin synthesis by fusion-arrested chick embryo myoblasts in cell culture. Dev Biol. 1976 Feb;48(2):431–437. doi: 10.1016/0012-1606(76)90104-4. [DOI] [PubMed] [Google Scholar]
  27. Okazaki K., Holtzer H. An analysis of myogenesis in vitro using fluorescein-labeled antimyosin. J Histochem Cytochem. 1965 Nov-Dec;13(8):726–739. doi: 10.1177/13.8.726. [DOI] [PubMed] [Google Scholar]
  28. Quinn L. S., Holtzer H., Nameroff M. Generation of chick skeletal muscle cells in groups of 16 from stem cells. Nature. 1985 Feb 21;313(6004):692–694. doi: 10.1038/313692a0. [DOI] [PubMed] [Google Scholar]
  29. Rubinstein N., Chi J., Holtzer H. Coordinated synthesis and degradation of actin and myosin in a variety of myogenic and non-myogenic cells. Exp Cell Res. 1976 Feb;97(2):387–393. doi: 10.1016/0014-4827(76)90630-3. [DOI] [PubMed] [Google Scholar]
  30. Sanger J. M., Mittal B., Pochapin M. B., Sanger J. W. Myofibrillogenesis in living cells microinjected with fluorescently labeled alpha-actinin. J Cell Biol. 1986 Jun;102(6):2053–2066. doi: 10.1083/jcb.102.6.2053. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Shimada Y., Obinata T. Polarity of actin filaments at the initial stage of myofibril assembly in myogenic cells in vitro. J Cell Biol. 1977 Mar;72(3):777–785. doi: 10.1083/jcb.72.3.777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Suzuki S., Pollack G. H. Bridgelike interconnections between thick filaments in stretched skeletal muscle fibers observed by the freeze-fracture method. J Cell Biol. 1986 Mar;102(3):1093–1098. doi: 10.1083/jcb.102.3.1093. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Tapscott S. J., Bennett G. S., Toyama Y., Kleinbart F., Holtzer H. Intermediate filament proteins in the developing chick spinal cord. Dev Biol. 1981 Aug;86(1):40–54. doi: 10.1016/0012-1606(81)90313-4. [DOI] [PubMed] [Google Scholar]
  34. Tokuyasu K. T., Maher P. A., Dutton A. H., Singer S. J. Intermediate filaments in skeletal and cardiac muscle tissue in embryonic and adult chicken. Ann N Y Acad Sci. 1985;455:200–212. doi: 10.1111/j.1749-6632.1985.tb50413.x. [DOI] [PubMed] [Google Scholar]
  35. Tokuyasu K. T., Maher P. A., Singer S. J. Distributions of vimentin and desmin in developing chick myotubes in vivo. II. Immunoelectron microscopic study. J Cell Biol. 1985 Apr;100(4):1157–1166. doi: 10.1083/jcb.100.4.1157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Toyama Y., Forry-Schaudies S., Hoffman B., Holtzer H. Effects of taxol and Colcemid on myofibrillogenesis. Proc Natl Acad Sci U S A. 1982 Nov;79(21):6556–6560. doi: 10.1073/pnas.79.21.6556. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Trinick J., Knight P., Whiting A. Purification and properties of native titin. J Mol Biol. 1984 Dec 5;180(2):331–356. doi: 10.1016/s0022-2836(84)80007-8. [DOI] [PubMed] [Google Scholar]
  38. Vertel B. M., Fischman D. A. Myosin accumulation in mononucleated cells of chick muscle cultures. Dev Biol. 1976 Feb;48(2):438–446. doi: 10.1016/0012-1606(76)90105-6. [DOI] [PubMed] [Google Scholar]
  39. Wang K. Sarcomere-associated cytoskeletal lattices in striated muscle. Review and hypothesis. Cell Muscle Motil. 1985;6:315–369. doi: 10.1007/978-1-4757-4723-2_10. [DOI] [PubMed] [Google Scholar]
  40. Wang K., Williamson C. L. Identification of an N2 line protein of striated muscle. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3254–3258. doi: 10.1073/pnas.77.6.3254. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wang S. M., Greaser M. L. Immunocytochemical studies using a monoclonal antibody to bovine cardiac titin on intact and extracted myofibrils. J Muscle Res Cell Motil. 1985 Jun;6(3):293–312. doi: 10.1007/BF00713171. [DOI] [PubMed] [Google Scholar]
  42. Yeoh G. C., Holtzer H. The effect of cell density, conditioned medium and cytosine arabinoside on myogenesis in primary and secondary cultures. Exp Cell Res. 1977 Jan;104(1):63–78. doi: 10.1016/0014-4827(77)90069-6. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES