Titin-actin interaction in mouse myocardium: passive tension modulation and its regulation by calcium/S100A1.
Passive tension in striated muscles derives primarily from the extension of the giant protein titin. However, several studies have suggested that, in cardiac muscle, interactions between titin and actin might also contribute to passive tension. We expressed recombinant fragments representing the subdomains of the extensible region of cardiac N2B titin (tandem-Ig segments, the N2B splice element, and the PEVK domain), and assayed them for binding to F-actin. The PEVK fragment bound F-actin, but no binding was detected for the other fragments. Comparison with a skeletal muscle PEVK fragment revealed that only the cardiac PEVK binds actin at physiological ionic strengths. The significance of PEVK-actin interaction was investigated using in vitro motility and single-myocyte mechanics. As F-actin slid relative to titin in the motility assay, a dynamic interaction between the PEVK domain and F-actin retarded filament sliding. Myocyte results suggest that a similar interaction makes a significant contribution to the passive tension. We also investigated the effect of calcium on PEVK-actin interaction. Although calcium alone had no effect, S100A1, a soluble calcium-binding protein found at high concentrations in the myocardium, inhibited PEVK-actin interaction in a calcium-dependent manner. Gel overlay analysis revealed that S100A1 bound the PEVK region in vitro in a calcium-dependent manner, and S100A1 binding was observed at several sites along titin's extensible region in situ, including the PEVK domain. In vitro motility results indicate that S100A1-PEVK interaction reduces the force that arises as F-actin slides relative to the PEVK domain, and we speculate that S100A1 may provide a mechanism to free the thin filament from titin and reduce titin-based tension before active contraction.
Yamasaki
R
R
Department of Veterinary and Comparative Anatomy, Pharmacology, and Physiology, Washington State University, Pullman, Washington 99164-6520, USA.
Berri
M
M
Wu
Y
Y
Trombitás
K
K
McNabb
M
M
Kellermayer
M S
MS
Witt
C
C
Labeit
D
D
Labeit
S
S
Greaser
M
M
Granzier
H
H
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites / Q000502:physiology; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000378:metabolism* / Q000494:pharmacology; D064211:Connectin; D051379:Mice; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009124:Muscle Proteins / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism*; D009994:Osmolar Concentration; D011494:Protein Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary / Q000502:physiology; D011994:Recombinant Proteins / Q000378:metabolism; D009418:S100 Proteins
D000199:Actins; D002135:Calcium-Binding Proteins; D064211:Connectin; D009124:Muscle Proteins; D011994:Recombinant Proteins; D009418:S100 Proteins; C110294:S100A1 protein; D011494:Protein Kinases; D002118:Calcium