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© The Rockefeller University Press, 0021-9525/1999//631 $5.00
The Journal of Cell Biology, Volume 146, Number 3, , 1999 631-644

I-Band Titin in Cardiac Muscle Is a Three-Element Molecular Spring and Is Critical for Maintaining Thin Filament Structure

^a Physiologisches Institut II, Universität Heidelberg, D-69120 Heidelberg, Germany
^b Department of Cell Biology and Anatomy, University of Arizona, Tucson, Arizona 85724
^c European Molecular Biology Laboratory, D-69012 Heidelberg, Germany
^d Max-Planck-Institut für molekulare Physiologie, D-44202 Dortmund, Germany
^e Institut für Anästhesiologie und Operative Intensivmedizin, Universitätsklinikum Mannheim, D-68167 Mannheim, Germany
^f Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona 85721
Physiologisches Institut II, Universität Heidelberg, Im Neuenheimer Feld 326, D-69120 Heidelberg, Germany.49-6221-54404949-6221-544054

wolfgang.linke{at}urz.uni-heidelberg.de

In cardiac muscle, the giant protein titin exists in different length isoforms expressed in the molecule's I-band region. Both isoforms, termed N2-A and N2-B, comprise stretches of Ig-like modules separated by the PEVK domain. Central I-band titin also contains isoform-specific Ig-motifs and nonmodular sequences, notably a longer insertion in N2-B. We investigated the elastic behavior of the I-band isoforms by using single-myofibril mechanics, immunofluorescence microscopy, and immunoelectron microscopy of rabbit cardiac sarcomeres stained with sequence-assigned antibodies. Moreover, we overexpressed constructs from the N2-B region in chick cardiac cells to search for possible structural properties of this cardiac-specific segment.

We found that cardiac titin contains three distinct elastic elements: poly-Ig regions, the PEVK domain, and the N2-B sequence insertion, which extends 60 nm at high physiological stretch. Recruitment of all three elements allows cardiac titin to extend fully reversibly at physiological sarcomere lengths, without the need to unfold Ig domains. Overexpressing the entire N2-B region or its NH₂ terminus in cardiac myocytes greatly disrupted thin filament, but not thick filament structure. Our results strongly suggest that the NH₂-terminal N2-B domains are necessary to stabilize thin filament integrity. N2-B–titin emerges as a unique region critical for both reversible extensibility and structural maintenance of cardiac myofibrils.

Key Words: heart muscle • connectin • elasticity • transfection • sarcomere

© 1999 The Rockefeller University Press

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