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. 1993 Nov 2;123(4):823–835. doi: 10.1083/jcb.123.4.823

Type 2X-myosin heavy chain is coded by a muscle fiber type-specific and developmentally regulated gene

PMCID: PMC2200149  PMID: 8227143

Abstract

We have previously reported the identification of a distinct myosin heavy chain (MyHC) isoform in a major subpopulation of rat skeletal muscle fibers, referred to as 2X fibers (Schiaffino, S., L. Gorza, S. Sartore, L. Saggin, M. Vianello, K. Gundersen, and T. Lomo. 1989. J. Muscle Res. Cell Motil. 10:197-205). However, it was not known whether 2X-MyHC is the product of posttranslational modification of other MyHCs or is coded by a distinct mRNA. We report here the isolation and characterization of cDNAs coding a MyHC isoform that is expressed in type 2X skeletal muscle fibers. 2X-MyHC transcripts differ from other MyHC transcripts in their restriction map and 3' end sequence and are thus derived from a distinct gene. In situ hybridization analyses show that 2X-MyHC transcripts are expressed at high levels in the diaphragm and fast hindlimb muscles and can be coexpressed either with 2B- or 2A- MyHC transcripts in a number of fibers. At the single fiber level the distribution of each MyHC mRNA closely matches that of the corresponding protein, determined by specific antibodies on serial sections. In hindlimb muscles 2X-, 2A-, and 2B-MyHC transcripts are first detected by postnatal day 2-5 and display from the earliest stages a distinct pattern of distribution in different muscles and different fibers. The emergence of type 2 MyHC isoforms thus defines a distinct neonatal phase of fiber type differentiation during muscle development. The functional significance of MyHC isoforms is discussed with particular reference to the velocity of shortening of skeletal muscle fibers.

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Selected References

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  1. Aigner S., Gohlsch B., Hämäläinen N., Staron R. S., Uber A., Wehrle U., Pette D. Fast myosin heavy chain diversity in skeletal muscles of the rabbit: heavy chain IId, not IIb predominates. Eur J Biochem. 1993 Jan 15;211(1-2):367–372. doi: 10.1111/j.1432-1033.1993.tb19906.x. [DOI] [PubMed] [Google Scholar]
  2. Ausoni S., Gorza L., Schiaffino S., Gundersen K., Lømo T. Expression of myosin heavy chain isoforms in stimulated fast and slow rat muscles. J Neurosci. 1990 Jan;10(1):153–160. doi: 10.1523/JNEUROSCI.10-01-00153.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Biral D., Betto R., Danieli-Betto D., Salviati G. Myosin heavy chain composition of single fibres from normal human muscle. Biochem J. 1988 Feb 15;250(1):307–308. doi: 10.1042/bj2500307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bottinelli R., Schiaffino S., Reggiani C. Force-velocity relations and myosin heavy chain isoform compositions of skinned fibres from rat skeletal muscle. J Physiol. 1991 Jun;437:655–672. doi: 10.1113/jphysiol.1991.sp018617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brooke M. H., Kaiser K. K. Muscle fiber types: how many and what kind? Arch Neurol. 1970 Oct;23(4):369–379. doi: 10.1001/archneur.1970.00480280083010. [DOI] [PubMed] [Google Scholar]
  6. Butler-Browne G. S., Whalen R. G. Myosin isozyme transitions occurring during the postnatal development of the rat soleus muscle. Dev Biol. 1984 Apr;102(2):324–334. doi: 10.1016/0012-1606(84)90197-0. [DOI] [PubMed] [Google Scholar]
  7. Bär A., Pette D. Three fast myosin heavy chains in adult rat skeletal muscle. FEBS Lett. 1988 Aug 1;235(1-2):153–155. doi: 10.1016/0014-5793(88)81253-5. [DOI] [PubMed] [Google Scholar]
  8. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  9. Condon K., Silberstein L., Blau H. M., Thompson W. J. Development of muscle fiber types in the prenatal rat hindlimb. Dev Biol. 1990 Apr;138(2):256–274. doi: 10.1016/0012-1606(90)90196-p. [DOI] [PubMed] [Google Scholar]
  10. Cox R. D., Weydert A., Barlow D., Buckingham M. E. Three linked myosin heavy chain genes clustered within 370 kb of each other show independent transcriptional and post-transcriptional regulation during differentiation of a mouse muscle cell line. Dev Biol. 1991 Jan;143(1):36–43. doi: 10.1016/0012-1606(91)90052-5. [DOI] [PubMed] [Google Scholar]
  11. Dhoot G. K. Selective synthesis and degradation of slow skeletal myosin heavy chains in developing muscle fibers. Muscle Nerve. 1986 Feb;9(2):155–164. doi: 10.1002/mus.880090209. [DOI] [PubMed] [Google Scholar]
  12. Eddinger T. J., Moss R. L. Mechanical properties of skinned single fibers of identified types from rat diaphragm. Am J Physiol. 1987 Aug;253(2 Pt 1):C210–C218. doi: 10.1152/ajpcell.1987.253.2.C210. [DOI] [PubMed] [Google Scholar]
  13. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  14. Gorza L., Gundersen K., Lømo T., Schiaffino S., Westgaard R. H. Slow-to-fast transformation of denervated soleus muscles by chronic high-frequency stimulation in the rat. J Physiol. 1988 Aug;402:627–649. doi: 10.1113/jphysiol.1988.sp017226. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gorza L. Identification of a novel type 2 fiber population in mammalian skeletal muscle by combined use of histochemical myosin ATPase and anti-myosin monoclonal antibodies. J Histochem Cytochem. 1990 Feb;38(2):257–265. doi: 10.1177/38.2.2137154. [DOI] [PubMed] [Google Scholar]
  16. Greaser M. L., Moss R. L., Reiser P. J. Variations in contractile properties of rabbit single muscle fibres in relation to troponin T isoforms and myosin light chains. J Physiol. 1988 Dec;406:85–98. doi: 10.1113/jphysiol.1988.sp017370. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Gubler U. A one tube reaction for the synthesis of blunt-ended double-stranded cDNA. Nucleic Acids Res. 1988 Mar 25;16(6):2726–2726. doi: 10.1093/nar/16.6.2726. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gubler U., Hoffman B. J. A simple and very efficient method for generating cDNA libraries. Gene. 1983 Nov;25(2-3):263–269. doi: 10.1016/0378-1119(83)90230-5. [DOI] [PubMed] [Google Scholar]
  19. Gunning P., Hardeman E. Multiple mechanisms regulate muscle fiber diversity. FASEB J. 1991 Dec;5(15):3064–3070. doi: 10.1096/fasebj.5.15.1835946. [DOI] [PubMed] [Google Scholar]
  20. Harris A. J., Fitzsimons R. B., McEwan J. C. Neural control of the sequence of expression of myosin heavy chain isoforms in foetal mammalian muscles. Development. 1989 Dec;107(4):751–769. doi: 10.1242/dev.107.4.751. [DOI] [PubMed] [Google Scholar]
  21. Izumo S., Nadal-Ginard B., Mahdavi V. All members of the MHC multigene family respond to thyroid hormone in a highly tissue-specific manner. Science. 1986 Feb 7;231(4738):597–600. doi: 10.1126/science.3945800. [DOI] [PubMed] [Google Scholar]
  22. Kavinsky C. J., Umeda P. K., Sinha A. M., Elzinga M., Tong S. W., Zak R., Jakovcic S., Rabinowitz M. Cloned mRNA sequences for two types of embryonic myosin heavy chains from chick skeletal muscle. I. DNA and derived amino acid sequence of light meromyosin. J Biol Chem. 1983 Apr 25;258(8):5196–5205. [PubMed] [Google Scholar]
  23. Kelly A. M., Rosser B. W., Hoffman R., Panettieri R. A., Schiaffino S., Rubinstein N. A., Nemeth P. M. Metabolic and contractile protein expression in developing rat diaphragm muscle. J Neurosci. 1991 May;11(5):1231–1242. doi: 10.1523/JNEUROSCI.11-05-01231.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Klitgaard H., Zhou M., Schiaffino S., Betto R., Salviati G., Saltin B. Ageing alters the myosin heavy chain composition of single fibres from human skeletal muscle. Acta Physiol Scand. 1990 Sep;140(1):55–62. doi: 10.1111/j.1748-1716.1990.tb08975.x. [DOI] [PubMed] [Google Scholar]
  25. LaFramboise W. A., Daood M. J., Guthrie R. D., Moretti P., Schiaffino S., Ontell M. Electrophoretic separation and immunological identification of type 2X myosin heavy chain in rat skeletal muscle. Biochim Biophys Acta. 1990 Jul 20;1035(1):109–112. doi: 10.1016/0304-4165(90)90181-u. [DOI] [PubMed] [Google Scholar]
  26. LaFramboise W. A., Daood M. J., Guthrie R. D., Schiaffino S., Moretti P., Brozanski B., Ontell M. P., Butler-Browne G. S., Whalen R. G., Ontell M. Emergence of the mature myosin phenotype in the rat diaphragm muscle. Dev Biol. 1991 Mar;144(1):1–15. doi: 10.1016/0012-1606(91)90473-g. [DOI] [PubMed] [Google Scholar]
  27. Larsson L., Ansved T., Edström L., Gorza L., Schiaffino S. Effects of age on physiological, immunohistochemical and biochemical properties of fast-twitch single motor units in the rat. J Physiol. 1991 Nov;443:257–275. doi: 10.1113/jphysiol.1991.sp018833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Larsson L., Biral D., Campione M., Schiaffino S. An age-related type IIB to IIX myosin heavy chain switching in rat skeletal muscle. Acta Physiol Scand. 1993 Feb;147(2):227–234. doi: 10.1111/j.1748-1716.1993.tb09493.x. [DOI] [PubMed] [Google Scholar]
  29. Larsson L., Edström L., Lindegren B., Gorza L., Schiaffino S. MHC composition and enzyme-histochemical and physiological properties of a novel fast-twitch motor unit type. Am J Physiol. 1991 Jul;261(1 Pt 1):C93–101. doi: 10.1152/ajpcell.1991.261.1.C93. [DOI] [PubMed] [Google Scholar]
  30. Leinwand L. A., Fournier R. E., Nadal-Ginard B., Shows T. B. Multigene family for sarcomeric myosin heavy chain in mouse and human DNA: localization on a single chromosome. Science. 1983 Aug 19;221(4612):766–769. doi: 10.1126/science.6879174. [DOI] [PubMed] [Google Scholar]
  31. Lyons G. E., Haselgrove J., Kelly A. M., Rubinstein N. A. Myosin transitions in developing fast and slow muscles of the rat hindlimb. Differentiation. 1983;25(2):168–175. doi: 10.1111/j.1432-0436.1984.tb01352.x. [DOI] [PubMed] [Google Scholar]
  32. Maeda K., Sczakiel G., Wittinghofer A. Characterization of cDNA coding for the complete light meromyosin portion of a rabbit fast skeletal muscle myosin heavy chain. Eur J Biochem. 1987 Aug 17;167(1):97–102. doi: 10.1111/j.1432-1033.1987.tb13308.x. [DOI] [PubMed] [Google Scholar]
  33. Mahdavi V., Izumo S., Nadal-Ginard B. Developmental and hormonal regulation of sarcomeric myosin heavy chain gene family. Circ Res. 1987 Jun;60(6):804–814. doi: 10.1161/01.res.60.6.804. [DOI] [PubMed] [Google Scholar]
  34. Narusawa M., Fitzsimons R. B., Izumo S., Nadal-Ginard B., Rubinstein N. A., Kelly A. M. Slow myosin in developing rat skeletal muscle. J Cell Biol. 1987 Mar;104(3):447–459. doi: 10.1083/jcb.104.3.447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Parker-Thornburg J., Bauer B., Palermo J., Robbins J. Structural and developmental analysis of two linked myosin heavy chain genes. Dev Biol. 1992 Mar;150(1):99–107. doi: 10.1016/0012-1606(92)90010-e. [DOI] [PubMed] [Google Scholar]
  36. Reiser P. J., Moss R. L., Giulian G. G., Greaser M. L. Shortening velocity in single fibers from adult rabbit soleus muscles is correlated with myosin heavy chain composition. J Biol Chem. 1985 Aug 5;260(16):9077–9080. [PubMed] [Google Scholar]
  37. Russell S. D., Cambon N., Nadal-Ginard B., Whalen R. G. Thyroid hormone induces a nerve-independent precocious expression of fast myosin heavy chain mRNA in rat hindlimb skeletal muscle. J Biol Chem. 1988 May 5;263(13):6370–6374. [PubMed] [Google Scholar]
  38. Saez L., Leinwand L. A. Characterization of diverse forms of myosin heavy chain expressed in adult human skeletal muscle. Nucleic Acids Res. 1986 Apr 11;14(7):2951–2969. doi: 10.1093/nar/14.7.2951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sassoon D. A., Garner I., Buckingham M. Transcripts of alpha-cardiac and alpha-skeletal actins are early markers for myogenesis in the mouse embryo. Development. 1988 Sep;104(1):155–164. doi: 10.1242/dev.104.1.155. [DOI] [PubMed] [Google Scholar]
  41. Schiaffino S., Gorza L., Sartore S., Saggin L., Ausoni S., Vianello M., Gundersen K., Lømo T. Three myosin heavy chain isoforms in type 2 skeletal muscle fibres. J Muscle Res Cell Motil. 1989 Jun;10(3):197–205. doi: 10.1007/BF01739810. [DOI] [PubMed] [Google Scholar]
  42. Schiaffino S., Samuel J. L., Sassoon D., Lompré A. M., Garner I., Marotte F., Buckingham M., Rappaport L., Schwartz K. Nonsynchronous accumulation of alpha-skeletal actin and beta-myosin heavy chain mRNAs during early stages of pressure-overload--induced cardiac hypertrophy demonstrated by in situ hybridization. Circ Res. 1989 May;64(5):937–948. doi: 10.1161/01.res.64.5.937. [DOI] [PubMed] [Google Scholar]
  43. Strehler E. E., Strehler-Page M. A., Perriard J. C., Periasamy M., Nadal-Ginard B. Complete nucleotide and encoded amino acid sequence of a mammalian myosin heavy chain gene. Evidence against intron-dependent evolution of the rod. J Mol Biol. 1986 Aug 5;190(3):291–317. doi: 10.1016/0022-2836(86)90003-3. [DOI] [PubMed] [Google Scholar]
  44. Sweeney H. L., Kushmerick M. J., Mabuchi K., Sréter F. A., Gergely J. Myosin alkali light chain and heavy chain variations correlate with altered shortening velocity of isolated skeletal muscle fibers. J Biol Chem. 1988 Jun 25;263(18):9034–9039. [PubMed] [Google Scholar]
  45. Termin A., Staron R. S., Pette D. Changes in myosin heavy chain isoforms during chronic low-frequency stimulation of rat fast hindlimb muscles. A single-fiber study. Eur J Biochem. 1989 Dec 22;186(3):749–754. doi: 10.1111/j.1432-1033.1989.tb15269.x. [DOI] [PubMed] [Google Scholar]
  46. Termin A., Staron R. S., Pette D. Myosin heavy chain isoforms in histochemically defined fiber types of rat muscle. Histochemistry. 1989;92(6):453–457. doi: 10.1007/BF00524756. [DOI] [PubMed] [Google Scholar]
  47. Weydert A., Barton P., Harris A. J., Pinset C., Buckingham M. Developmental pattern of mouse skeletal myosin heavy chain gene transcripts in vivo and in vitro. Cell. 1987 Apr 10;49(1):121–129. doi: 10.1016/0092-8674(87)90762-8. [DOI] [PubMed] [Google Scholar]
  48. Weydert A., Daubas P., Caravatti M., Minty A., Bugaisky G., Cohen A., Robert B., Buckingham M. Sequential accumulation of mRNAs encoding different myosin heavy chain isoforms during skeletal muscle development in vivo detected with a recombinant plasmid identified as coding for an adult fast myosin heavy chain from mouse skeletal muscle. J Biol Chem. 1983 Nov 25;258(22):13867–13874. [PubMed] [Google Scholar]
  49. Wieczorek D. F., Periasamy M., Butler-Browne G. S., Whalen R. G., Nadal-Ginard B. Co-expression of multiple myosin heavy chain genes, in addition to a tissue-specific one, in extraocular musculature. J Cell Biol. 1985 Aug;101(2):618–629. doi: 10.1083/jcb.101.2.618. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Yoon S. J., Seiler S. H., Kucherlapati R., Leinwand L. Organization of the human skeletal myosin heavy chain gene cluster. Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):12078–12082. doi: 10.1073/pnas.89.24.12078. [DOI] [PMC free article] [PubMed] [Google Scholar]

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