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. 2000 Dec;79(6):3226–3234. doi: 10.1016/S0006-3495(00)76555-6

Extensibility of isoforms of cardiac titin: variation in contour length of molecular subsegments provides a basis for cellular passive stiffness diversity.

K Trombitás 1, A Redkar 1, T Centner 1, Y Wu 1, S Labeit 1, H Granzier 1
PMCID: PMC1301197  PMID: 11106626

Abstract

Titin is a giant polypeptide that spans between the Z- and M-lines of the cardiac muscle sarcomere and that develops force when extended. This force arises from titin's extensible I-band region, which consists mainly of three segment types: serially linked immunoglobulin-like domains (Ig segments), interrupted by the PEVK segment, and the N2B unique sequence. Recently it was reported that the myocardium of large mammals co-expresses small (N2B) and large (N2BA) cardiac isoforms and that the passive stiffness of cardiac myocytes varies with the isoform expression ratio. To understand the molecular basis of the differences in passive stiffness we investigated titin's extensibility in bovine atrium, which expresses predominantly N2BA titin, and compared it to that of rat, which expresses predominantly N2B titin. Immunoelectron microscopy was used with antibodies that flank the Ig segments, the PEVK segment, and the unique sequence of the N2B element. The extension of the various segments was then determined as a function of sarcomere length (SL). When slack sarcomeres of bovine atrium were stretched, the PEVK segment extended much more steeply and the unique N2B sequence less steeply than in rat, while the Ig segments behaved similarly in both species. However, the extensions normalized with the segment's contour length (i.e., the fractional extensions) of Ig, PEVK, and unique sequence segments all increase less steeply with SL in cow than in rat. Considering that fractional extension determines the level of entropic force, these differences in fractional extension are expected to result in shallow and steep passive force-SL curves in myocytes that express high levels of N2BA and N2B titin, respectively. Thus, the findings provide a molecular basis for passive stiffness diversity.

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Selected References

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  1. Cazorla O., Freiburg A., Helmes M., Centner T., McNabb M., Wu Y., Trombitás K., Labeit S., Granzier H. Differential expression of cardiac titin isoforms and modulation of cellular stiffness. Circ Res. 2000 Jan 7;86(1):59–67. doi: 10.1161/01.res.86.1.59. [DOI] [PubMed] [Google Scholar]
  2. Fabiato A., Fabiato F. Myofilament-generated tension oscillations during partial calcium activation and activation dependence of the sarcomere length-tension relation of skinned cardiac cells. J Gen Physiol. 1978 Nov;72(5):667–699. doi: 10.1085/jgp.72.5.667. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Freiburg A., Trombitas K., Hell W., Cazorla O., Fougerousse F., Centner T., Kolmerer B., Witt C., Beckmann J. S., Gregorio C. C. Series of exon-skipping events in the elastic spring region of titin as the structural basis for myofibrillar elastic diversity. Circ Res. 2000 Jun 9;86(11):1114–1121. doi: 10.1161/01.res.86.11.1114. [DOI] [PubMed] [Google Scholar]
  4. Granzier H. L., Irving T. C. Passive tension in cardiac muscle: contribution of collagen, titin, microtubules, and intermediate filaments. Biophys J. 1995 Mar;68(3):1027–1044. doi: 10.1016/S0006-3495(95)80278-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Granzier H. L., Wang K. Gel electrophoresis of giant proteins: solubilization and silver-staining of titin and nebulin from single muscle fiber segments. Electrophoresis. 1993 Jan-Feb;14(1-2):56–64. doi: 10.1002/elps.1150140110. [DOI] [PubMed] [Google Scholar]
  6. Granzier H., Helmes M., Trombitás K. Nonuniform elasticity of titin in cardiac myocytes: a study using immunoelectron microscopy and cellular mechanics. Biophys J. 1996 Jan;70(1):430–442. doi: 10.1016/S0006-3495(96)79586-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Granzier H., Kellermayer M., Helmes M., Trombitás K. Titin elasticity and mechanism of passive force development in rat cardiac myocytes probed by thin-filament extraction. Biophys J. 1997 Oct;73(4):2043–2053. doi: 10.1016/S0006-3495(97)78234-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Gregorio C. C., Granzier H., Sorimachi H., Labeit S. Muscle assembly: a titanic achievement? Curr Opin Cell Biol. 1999 Feb;11(1):18–25. doi: 10.1016/s0955-0674(99)80003-9. [DOI] [PubMed] [Google Scholar]
  9. Grimm A. F., Lin H. L., Grimm B. R. Left ventricular free wall and intraventricular pressure-sarcomere length distributions. Am J Physiol. 1980 Jul;239(1):H101–H107. doi: 10.1152/ajpheart.1980.239.1.H101. [DOI] [PubMed] [Google Scholar]
  10. Helmes M., Trombitás K., Centner T., Kellermayer M., Labeit S., Linke W. A., Granzier H. Mechanically driven contour-length adjustment in rat cardiac titin's unique N2B sequence: titin is an adjustable spring. Circ Res. 1999 Jun 11;84(11):1339–1352. doi: 10.1161/01.res.84.11.1339. [DOI] [PubMed] [Google Scholar]
  11. Kellermayer M. S., Smith S. B., Granzier H. L., Bustamante C. Folding-unfolding transitions in single titin molecules characterized with laser tweezers. Science. 1997 May 16;276(5315):1112–1116. doi: 10.1126/science.276.5315.1112. [DOI] [PubMed] [Google Scholar]
  12. Kentish J. C., ter Keurs H. E., Ricciardi L., Bucx J. J., Noble M. I. Comparison between the sarcomere length-force relations of intact and skinned trabeculae from rat right ventricle. Influence of calcium concentrations on these relations. Circ Res. 1986 Jun;58(6):755–768. doi: 10.1161/01.res.58.6.755. [DOI] [PubMed] [Google Scholar]
  13. Labeit S., Kolmerer B., Linke W. A. The giant protein titin. Emerging roles in physiology and pathophysiology. Circ Res. 1997 Feb;80(2):290–294. doi: 10.1161/01.res.80.2.290. [DOI] [PubMed] [Google Scholar]
  14. Labeit S., Kolmerer B. Titins: giant proteins in charge of muscle ultrastructure and elasticity. Science. 1995 Oct 13;270(5234):293–296. doi: 10.1126/science.270.5234.293. [DOI] [PubMed] [Google Scholar]
  15. Linke W. A., Rudy D. E., Centner T., Gautel M., Witt C., Labeit S., Gregorio C. C. I-band titin in cardiac muscle is a three-element molecular spring and is critical for maintaining thin filament structure. J Cell Biol. 1999 Aug 9;146(3):631–644. doi: 10.1083/jcb.146.3.631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Maruyama K. Connectin/titin, giant elastic protein of muscle. FASEB J. 1997 Apr;11(5):341–345. doi: 10.1096/fasebj.11.5.9141500. [DOI] [PubMed] [Google Scholar]
  17. Neuhoff V., Arold N., Taube D., Ehrhardt W. Improved staining of proteins in polyacrylamide gels including isoelectric focusing gels with clear background at nanogram sensitivity using Coomassie Brilliant Blue G-250 and R-250. Electrophoresis. 1988 Jun;9(6):255–262. doi: 10.1002/elps.1150090603. [DOI] [PubMed] [Google Scholar]
  18. Trinick J., Tskhovrebova L. Titin: a molecular control freak. Trends Cell Biol. 1999 Oct;9(10):377–380. doi: 10.1016/s0962-8924(99)01641-4. [DOI] [PubMed] [Google Scholar]
  19. Trombitás K., Freiburg A., Centner T., Labeit S., Granzier H. Molecular dissection of N2B cardiac titin's extensibility. Biophys J. 1999 Dec;77(6):3189–3196. doi: 10.1016/S0006-3495(99)77149-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Trombitás K., Greaser M., French G., Granzier H. PEVK extension of human soleus muscle titin revealed by immunolabeling with the anti-titin antibody 9D10. J Struct Biol. 1998;122(1-2):188–196. doi: 10.1006/jsbi.1998.3984. [DOI] [PubMed] [Google Scholar]
  21. Trombitás K., Greaser M., Labeit S., Jin J. P., Kellermayer M., Helmes M., Granzier H. Titin extensibility in situ: entropic elasticity of permanently folded and permanently unfolded molecular segments. J Cell Biol. 1998 Feb 23;140(4):853–859. doi: 10.1083/jcb.140.4.853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wang K. Titin/connectin and nebulin: giant protein rulers of muscle structure and function. Adv Biophys. 1996;33:123–134. [PubMed] [Google Scholar]

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