• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of jcellbiolHomeThe Rockefeller University PressEditorsContactInstructions for AuthorsThis issue
J Cell Biol. May 1, 1988; 106(5): 1563–1572.
PMCID: PMC2115059

The organization of titin filaments in the half-sarcomere revealed by monoclonal antibodies in immunoelectron microscopy: a map of ten nonrepetitive epitopes starting at the Z line extends close to the M line

Abstract

mAbs specific for titin or nebulin were characterized by immunoblotting and fluorescence microscopy. Immunoelectron microscopy on relaxed chicken breast muscle revealed unique transverse striping patterns. Each of the 10 distinct titin antibodies provided a pair of delicate decoration lines per sarcomere. The position of these pairs was centrally symmetric to the M line and was antibody dependent. The results provided a linear epitope map, which starts at the Z line (antibody T20), covers five distinct positions along the I band (T21, T12, T4, T1, T11), the A-I junction (T3), and three distinct positions within the A band (T10, T22, T23). The epitope of T23 locates 0.2 micron before the M line. In immunoblots, the two antibodies decorating at or just before the Z line (T20, T21) specifically recognized the insoluble titin TI component but did not recognize TII, a proteolytic derivative. All other titin antibodies recognized TI and TII. Thus titin molecules appear as polar structures lacking over large regions repetitive epitopes. One physical end seems related to Z line anchorage, while the other may bind close to the M line. Titin epitopes influenced by the contractional state of the sarcomere locate between the N1 line and the A-I junction (T4, T1, T11). We discuss the results in relation to titin molecules having half-sarcomere lengths. The three nebulin antibodies so far characterized again give rise to distinct pairs of stripes. These locate close to the N2 line.

Full Text

The Full Text of this article is available as a PDF (4.2M).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Debus E, Weber K, Osborn M. Monoclonal antibodies to desmin, the muscle-specific intermediate filament protein. EMBO J. 1983;2(12):2305–2312. [PMC free article] [PubMed]
  • Dennis JE, Shimizu T, Reinach FC, Fischman DA. Localization of C-protein isoforms in chicken skeletal muscle: ultrastructural detection using monoclonal antibodies. J Cell Biol. 1984 Apr;98(4):1514–1522. [PMC free article] [PubMed]
  • Fürst D, Nave R, Osborn M, Weber K, Bardosi A, Archidiacono N, Ferro M, Romano V, Romeo G. Nebulin and titin expression in Duchenne muscular dystrophy appears normal. FEBS Lett. 1987 Nov 16;224(1):49–53. [PubMed]
  • Hashimoto K, Kamitani T, Wada Y, Tatsumi N. Presence of connectin-like protein in white blood cells and platelets. Tohoku J Exp Med. 1984 May;143(1):59–70. [PubMed]
  • Henderson LE, Oroszlan S, Konigsberg W. A micromethod for complete removal of dodecyl sulfate from proteins by ion-pair extraction. Anal Biochem. 1979 Feb;93(1):153–157. [PubMed]
  • Hill C, Weber K. Monoclonal antibodies distinguish titins from heart and skeletal muscle. J Cell Biol. 1986 Mar;102(3):1099–1108. [PMC free article] [PubMed]
  • Hoffman EP, Knudson CM, Campbell KP, Kunkel LM. Subcellular fractionation of dystrophin to the triads of skeletal muscle. Nature. 1987 Dec 24;330(6150):754–758. [PubMed]
  • Hu DH, Kimura S, Maruyama K. Sodium dodecyl sulfate gel electrophoresis studies of connectin-like high molecular weight proteins of various types of vertebrate and invertebrate muscles. J Biochem. 1986 May;99(5):1485–1492. [PubMed]
  • Itoh Y, Kimura S, Suzuki T, Ohashi K, Maruyama K. Native connectin from porcine cardiac muscle. J Biochem. 1986 Aug;100(2):439–447. [PubMed]
  • Kimura S, Maruyama K. Preparation of native connectin from chicken breast muscle. J Biochem. 1983 Dec;94(6):2083–2085. [PubMed]
  • Kimura S, Yoshidomi H, Maruyama K. Proteolytic fragments of connectin cause aggregation of myosin filaments but not of actin filaments. J Biochem. 1984 Dec;96(6):1947–1950. [PubMed]
  • Knight PJ, Trinick JA. Preparation of myofibrils. Methods Enzymol. 1982;85(Pt B):9–12. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Locker RH, Wild DJ. The N-lines of skeletal muscle. J Ultrastruct Res. 1984 Sep;88(3):207–222. [PubMed]
  • Locker RH, Wild DJ. A comparative study of high molecular weight proteins in various types of muscle across the animal kingdom. J Biochem. 1986 May;99(5):1473–1484. [PubMed]
  • Loewy AG, Wilson FJ, Taggart NM, Greene EA, Frasca P, Kaufman HS, Sorrell MJ. A covalently cross-linked matrix in skeletal muscle fibers. Cell Motil. 1983;3(5-6):463–483. [PubMed]
  • Maruyama K. Connectin, an elastic filamentous protein of striated muscle. Int Rev Cytol. 1986;104:81–114. [PubMed]
  • Maruyama K, Kimura S, Ohashi K, Kuwano Y. Connectin, an elastic protein of muscle. Identification of "titin" with connectin. J Biochem. 1981 Mar;89(3):701–709. [PubMed]
  • Maruyama K, Kimura S, Yoshidomi H, Sawada H, Kikuchi M. Molecular size and shape of beta-connectin, an elastic protein of striated muscle. J Biochem. 1984 May;95(5):1423–1433. [PubMed]
  • Maruyama K, Matsubara S, Natori R, Nonomura Y, Kimura S. Connectin, an elastic protein of muscle. Characterization and Function. J Biochem. 1977 Aug;82(2):317–337. [PubMed]
  • Maruyama K, Sawada H, Kimura S, Ohashi K, Higuchi H, Umazume Y. Connectin filaments in stretched skinned fibers of frog skeletal muscle. J Cell Biol. 1984 Oct;99(4 Pt 1):1391–1397. [PMC free article] [PubMed]
  • 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. [PMC free article] [PubMed]
  • Monaco AP, Neve RL, Colletti-Feener C, Bertelson CJ, Kurnit DM, Kunkel LM. Isolation of candidate cDNAs for portions of the Duchenne muscular dystrophy gene. Nature. 1986 Oct 16;323(6089):646–650. [PubMed]
  • Muguruma M, Kobayashi K, Fukazawa T, Ohashi K, Maruyama K. A new 220,000 dalton protein located in the Z lines of vertebrate skeletal muscle. J Biochem. 1981 Jun;89(6):1981–1984. [PubMed]
  • Osborn M, Hill C, Altmannsberger M, Weber K. Monoclonal antibodies to titin in conjunction with antibodies to desmin separate rhabdomyosarcomas from other tumor types. Lab Invest. 1986 Jul;55(1):101–108. [PubMed]
  • Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. [PMC free article] [PubMed]
  • Trinick JA. End-filaments: a new structural element of vertebrate skeletal muscle thick filaments. J Mol Biol. 1981 Sep 15;151(2):309–314. [PubMed]
  • Trinick J, Knight P, Whiting A. Purification and properties of native titin. J Mol Biol. 1984 Dec 5;180(2):331–356. [PubMed]
  • Wang K. Purification of titin and nebulin. Methods Enzymol. 1982;85(Pt B):264–274. [PubMed]
  • Wang K. Cytoskeletal matrix in striated muscle: the role of titin, nebulin and intermediate filaments. Adv Exp Med Biol. 1984;170:285–305. [PubMed]
  • Wang K. Sarcomere-associated cytoskeletal lattices in striated muscle. Review and hypothesis. Cell Muscle Motil. 1985;6:315–369. [PubMed]
  • Wang SM, Greaser ML. 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. [PubMed]
  • Wang K, Williamson CL. Identification of an N2 line protein of striated muscle. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3254–3258. [PMC free article] [PubMed]
  • Wang K, McClure J, Tu A. Titin: major myofibrillar components of striated muscle. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3698–3702. [PMC free article] [PubMed]
  • Wang K, Ramirez-Mitchell R, Palter D. Titin is an extraordinarily long, flexible, and slender myofibrillar protein. Proc Natl Acad Sci U S A. 1984 Jun;81(12):3685–3689. [PMC free article] [PubMed]
  • Wood DS, Zeviani M, Prelle A, Bonilla E, Salviati G, Miranda AF, DiMauro S, Rowland LP. Is nebulin the defective gene product in Duchenne muscular dystrophy? N Engl J Med. 1987 Jan 8;316(2):107–108. [PubMed]

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

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Cited in Books
    Cited in Books
    PubMed Central articles cited in books
  • MedGen
    MedGen
    Related information in MedGen
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...