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. 1993 Jul;12(7):2863–2871. doi: 10.1002/j.1460-2075.1993.tb05948.x

Kettin, a large modular protein in the Z-disc of insect muscles.

A Lakey 1, S Labeit 1, M Gautel 1, C Ferguson 1, D P Barlow 1, K Leonard 1, B Bullard 1
PMCID: PMC413539  PMID: 8335002

Abstract

Z-discs of insect flight muscle contain a large protein of 500-700 kDa. Monoclonal antibodies label an epitope in the molecule at the Z-disc in Drosophila and Lethocerus (waterbug). A partial cDNA of 1.6 kb from the Drosophila gene has been cloned and sequenced. The corresponding amino acid sequence has a modular structure composed of four conserved repeats of 95 amino acids homologous to immunoglobulin C2 domains (called class II domains in muscle proteins), separated by less conserved linker sequences of 35 amino acids. An expressed class II domain with flanking linker sequences binds to actin and alpha-actinin but not to myosin. Single molecules of the protein would be large enough to span the Z-disc. We suggest that the protein acts as scaffolding in the Z-disc and we call the protein kettin. The Ca2+ activated protease, calpain, disrupts the Z-disc of striated muscle, releasing alpha-actinin intact. Calpain digests kettin to a series of peptides of between 30 and 170 kDa which are released from the myofibril. Digestion of kettin may cause disintegration of the Z-disc and alpha-actinin release which lead to disassembly of the myofibril.

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Selected References

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

  1. Arakawa N., Robson R. M., Goll D. E. An improved method for the preparation of alpha-actinin from rabbit striated muscle. Biochim Biophys Acta. 1970 Feb 17;200(2):284–295. doi: 10.1016/0005-2795(70)90172-8. [DOI] [PubMed] [Google Scholar]
  2. Ayme-Southgate A., Vigoreaux J., Benian G., Pardue M. L. Drosophila has a twitchin/titin-related gene that appears to encode projectin. Proc Natl Acad Sci U S A. 1991 Sep 15;88(18):7973–7977. doi: 10.1073/pnas.88.18.7973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ball E., Karlik C. C., Beall C. J., Saville D. L., Sparrow J. C., Bullard B., Fyrberg E. A. Arthrin, a myofibrillar protein of insect flight muscle, is an actin-ubiquitin conjugate. Cell. 1987 Oct 23;51(2):221–228. doi: 10.1016/0092-8674(87)90149-8. [DOI] [PubMed] [Google Scholar]
  4. Benian G. M., Kiff J. E., Neckelmann N., Moerman D. G., Waterston R. H. Sequence of an unusually large protein implicated in regulation of myosin activity in C. elegans. Nature. 1989 Nov 2;342(6245):45–50. doi: 10.1038/342045a0. [DOI] [PubMed] [Google Scholar]
  5. Blitzer P. H. A consensus statement by radiation oncologists regarding radiotherapy for bone metastases. Am J Hosp Palliat Care. 1993 Jan-Feb;10(1):3–6. [PubMed] [Google Scholar]
  6. Bullard B., Bell J., Craig R., Leonard K. Arthrin: a new actin-like protein in insect flight muscle. J Mol Biol. 1985 Apr 5;182(3):443–454. doi: 10.1016/0022-2836(85)90203-7. [DOI] [PubMed] [Google Scholar]
  7. Bullard B., Leonard K., Larkins A., Butcher G., Karlik C., Fyrberg E. Troponin of asynchronous flight muscle. J Mol Biol. 1988 Dec 5;204(3):621–637. doi: 10.1016/0022-2836(88)90360-9. [DOI] [PubMed] [Google Scholar]
  8. Bullard B., Sainsbury G., Miller N. Digestion of proteins associated with the Z-disc by calpain. J Muscle Res Cell Motil. 1990 Jun;11(3):271–279. doi: 10.1007/BF01843580. [DOI] [PubMed] [Google Scholar]
  9. Cheng N. Q., Deatherage J. F. Three-dimensional reconstruction of the Z disk of sectioned bee flight muscle. J Cell Biol. 1989 May;108(5):1761–1774. doi: 10.1083/jcb.108.5.1761. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  11. Cong J., Goll D. E., Peterson A. M., Kapprell H. P. The role of autolysis in activity of the Ca2+-dependent proteinases (mu-calpain and m-calpain). J Biol Chem. 1989 Jun 15;264(17):10096–10103. [PubMed] [Google Scholar]
  12. Dayton W. R., Reville W. J., Goll D. E., Stromer M. H. A Ca2+-activated protease possibly involved in myofibrillar protein turnover. Partial characterization of the purified enzyme. Biochemistry. 1976 May 18;15(10):2159–2167. doi: 10.1021/bi00655a020. [DOI] [PubMed] [Google Scholar]
  13. Deatherage J. F., Cheng N. Q., Bullard B. Arrangement of filaments and cross-links in the bee flight muscle Z disk by image analysis of oblique sections. J Cell Biol. 1989 May;108(5):1775–1782. doi: 10.1083/jcb.108.5.1775. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Einheber S., Fischman D. A. Isolation and characterization of a cDNA clone encoding avian skeletal muscle C-protein: an intracellular member of the immunoglobulin superfamily. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2157–2161. doi: 10.1073/pnas.87.6.2157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Feinberg A. P., Vogelstein B. "A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity". Addendum. Anal Biochem. 1984 Feb;137(1):266–267. doi: 10.1016/0003-2697(84)90381-6. [DOI] [PubMed] [Google Scholar]
  17. Fyrberg C. C., Labeit S., Bullard B., Leonard K., Fyrberg E. Drosophila projectin: relatedness to titin and twitchin and correlation with lethal(4) 102 CDa and bent-dominant mutants. Proc Biol Sci. 1992 Jul 22;249(1324):33–40. doi: 10.1098/rspb.1992.0080. [DOI] [PubMed] [Google Scholar]
  18. Fürst D. O., Osborn M., Nave R., Weber K. 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. J Cell Biol. 1988 May;106(5):1563–1572. doi: 10.1083/jcb.106.5.1563. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Goll D. E., Dayton W. R., Singh I., Robson R. M. Studies of the alpha-actinin/actin interaction in the Z-disk by using calpain. J Biol Chem. 1991 May 5;266(13):8501–8510. [PubMed] [Google Scholar]
  20. Hammond K. S., Goll D. E. Purification of insect myosin and alpha-actinin. Biochem J. 1975 Oct;151(1):189–192. doi: 10.1042/bj1510189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kimura S., Matsuura T., Ohtsuka S., Nakauchi Y., Matsuno A., Maruyama K. Characterization and localization of alpha-connectin (titin 1): an elastic protein isolated from rabbit skeletal muscle. J Muscle Res Cell Motil. 1992 Feb;13(1):39–47. doi: 10.1007/BF01738426. [DOI] [PubMed] [Google Scholar]
  22. Labeit S., Barlow D. P., Gautel M., Gibson T., Holt J., Hsieh C. L., Francke U., Leonard K., Wardale J., Whiting A. A regular pattern of two types of 100-residue motif in the sequence of titin. Nature. 1990 May 17;345(6272):273–276. doi: 10.1038/345273a0. [DOI] [PubMed] [Google Scholar]
  23. Labeit S., Gautel M., Lakey A., Trinick J. Towards a molecular understanding of titin. EMBO J. 1992 May;11(5):1711–1716. doi: 10.1002/j.1460-2075.1992.tb05222.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  25. Lakey A., Ferguson C., Labeit S., Reedy M., Larkins A., Butcher G., Leonard K., Bullard B. Identification and localization of high molecular weight proteins in insect flight and leg muscle. EMBO J. 1990 Nov;9(11):3459–3467. doi: 10.1002/j.1460-2075.1990.tb07554.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Le Grice S. F., Grüninger-Leitch F. Rapid purification of homodimer and heterodimer HIV-1 reverse transcriptase by metal chelate affinity chromatography. Eur J Biochem. 1990 Jan 26;187(2):307–314. doi: 10.1111/j.1432-1033.1990.tb15306.x. [DOI] [PubMed] [Google Scholar]
  27. Maher P. A., Cox G. F., Singer S. J. Zeugmatin: a new high molecular weight protein associated with Z lines in adult and early embryonic striated muscle. J Cell Biol. 1985 Nov;101(5 Pt 1):1871–1883. doi: 10.1083/jcb.101.5.1871. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mierendorf R. C., Percy C., Young R. A. Gene isolation by screening lambda gt11 libraries with antibodies. Methods Enzymol. 1987;152:458–469. doi: 10.1016/0076-6879(87)52054-7. [DOI] [PubMed] [Google Scholar]
  29. Nave R., Weber K. A myofibrillar protein of insect muscle related to vertebrate titin connects Z band and A band: purification and molecular characterization of invertebrate mini-titin. J Cell Sci. 1990 Apr;95(Pt 4):535–544. doi: 10.1242/jcs.95.4.535. [DOI] [PubMed] [Google Scholar]
  30. Newman R., Butcher G. W., Bullard B., Leonard K. R. A method for determining the periodicity of a troponin component in isolated insect flight muscle thin filaments by gold/Fab labelling. J Cell Sci. 1992 Mar;101(Pt 3):503–508. doi: 10.1242/jcs.101.3.503. [DOI] [PubMed] [Google Scholar]
  31. Noguchi J., Yanagisawa M., Imamura M., Kasuya Y., Sakurai T., Tanaka T., Masaki T. Complete primary structure and tissue expression of chicken pectoralis M-protein. J Biol Chem. 1992 Oct 5;267(28):20302–20310. [PubMed] [Google Scholar]
  32. Olson N. J., Pearson R. B., Needleman D. S., Hurwitz M. Y., Kemp B. E., Means A. R. Regulatory and structural motifs of chicken gizzard myosin light chain kinase. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2284–2288. doi: 10.1073/pnas.87.6.2284. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Reddy M. K., Etlinger J. D., Rabinowitz M., Fischman D. A., Zak R. Removal of Z-lines and alpha-actinin from isolated myofibrils by a calcium-activated neutral protease. J Biol Chem. 1975 Jun 10;250(11):4278–4284. [PubMed] [Google Scholar]
  34. Reedy M. K., Leonard K. R., Freeman R., Arad T. Thick myofilament mass determination by electron scattering measurements with the scanning transmission electron microscope. J Muscle Res Cell Motil. 1981 Mar;2(1):45–64. doi: 10.1007/BF00712061. [DOI] [PubMed] [Google Scholar]
  35. Saide J. D., Chin-Bow S., Hogan-Sheldon J., Busquets-Turner L., Vigoreaux J. O., Valgeirsdottir K., Pardue M. L. Characterization of components of Z-bands in the fibrillar flight muscle of Drosophila melanogaster. J Cell Biol. 1989 Nov;109(5):2157–2167. doi: 10.1083/jcb.109.5.2157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Saide J. D., Chin-Bow S., Hogan-Sheldon J., Busquets-Turner L. Z-band proteins in the flight muscle and leg muscle of the honeybee. J Muscle Res Cell Motil. 1990 Apr;11(2):125–136. doi: 10.1007/BF01766491. [DOI] [PubMed] [Google Scholar]
  37. Saiki R. K., Scharf S., Faloona F., Mullis K. B., Horn G. T., Erlich H. A., Arnheim N. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science. 1985 Dec 20;230(4732):1350–1354. doi: 10.1126/science.2999980. [DOI] [PubMed] [Google Scholar]
  38. 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]
  39. Shoemaker M. O., Lau W., Shattuck R. L., Kwiatkowski A. P., Matrisian P. E., Guerra-Santos L., Wilson E., Lukas T. J., Van Eldik L. J., Watterson D. M. Use of DNA sequence and mutant analyses and antisense oligodeoxynucleotides to examine the molecular basis of nonmuscle myosin light chain kinase autoinhibition, calmodulin recognition, and activity. J Cell Biol. 1990 Sep;111(3):1107–1125. doi: 10.1083/jcb.111.3.1107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Staros J. V., Wright R. W., Swingle D. M. Enhancement by N-hydroxysulfosuccinimide of water-soluble carbodiimide-mediated coupling reactions. Anal Biochem. 1986 Jul;156(1):220–222. doi: 10.1016/0003-2697(86)90176-4. [DOI] [PubMed] [Google Scholar]
  41. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  42. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  43. 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]
  44. Vigoreaux J. O., Saide J. D., Pardue M. L. Structurally different Drosophila striated muscles utilize distinct variants of Z-band-associated proteins. J Muscle Res Cell Motil. 1991 Aug;12(4):340–354. doi: 10.1007/BF01738589. [DOI] [PubMed] [Google Scholar]
  45. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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