TY - JOUR
T1 - Dependence of thick filament structure in relaxed mammalian skeletal muscle on temperature and interfilament spacing
AU - Caremani, Marco
AU - Fusi, Luca
AU - Linari, Marco
AU - Reconditi, Massimo
AU - Piazzesi, Gabriella
AU - Irving, Thomas C.
AU - Narayanan, Theyencheri
AU - Irving, Malcolm
AU - Lombardi, Vincenzo
AU - Brunello, Elisabetta
N1 - Funding Information:
This work and some investigators were supported by Fondo per gli Investimenti della Ricerca di Base (Futuro in Ricerca project grant RBFR08JAMZ to E. Brunello, Italy); Progetti di Rilevante Interesse Nazionale-Ministero dell’Istruzione, dell’Università e della Ricerca, Italy and Telethon (Italy); Consorzio Nazionale In-teruniversitario per le Scienze Fisiche della Materia (Progetto d’Innesco della Ricerca Esplorativa 2007; Italy), Fondazione Cassa di Risparmio di Firenze (2016-2018; Italy); UK Medical Research Council (grant MR/M026655/1); and the European Synchrotron Radiation Facility. This research used resources of the Advanced Photon Source, a U.S. Department of Energy Office of Science User Facility operated for the Department of Energy Office of Science by Argonne National Laboratory under contract no. DE-AC02-06CH11357. The project was also supported by National Institutes of Health/National Institute of General Medical Sciences grant P41 GM103622 (BioCAT). M. Caremani was funded by University of Florence (competitive project marcocaremani_rictd1819; Italy). E. Brunello was funded by a British Heart Foundation Intermediate Basic Science Research Fellowship FS/17/3/32604. L. Fusi was funded by a Sir Henry Dale Fellowship awarded by the Wellcome Trust and the Royal Society (fellowship 210464/Z/18/Z). The authors declare no competing financial interests.
Publisher Copyright:
© 2021 Caremani et al. This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).
Copyright:
Copyright 2021 Elsevier B.V., All rights reserved.
PY - 2021/3/1
Y1 - 2021/3/1
N2 - Contraction of skeletal muscle is regulated by structural changes in both actin-containing thin filaments and myosin-containing thick filaments, but myosin-based regulation is unlikely to be preserved after thick filament isolation, and its structural basis remains poorly characterized. Here, we describe the periodic features of the thick filament structure in situ by high-resolution small-angle x-ray diffraction and interference. We used both relaxed demembranated fibers and resting intact muscle preparations to assess whether thick filament regulation is preserved in demembranated fibers, which have been widely used for previous studies. We show that the thick filaments in both preparations exhibit two closely spaced axial periodicities, 43.1 nm and 45.5 nm, at near-physiological temperature. The shorter periodicity matches that of the myosin helix, and x-ray interference between the two arrays of myosin in the bipolar filament shows that all zones of the filament follow this periodicity. The 45.5-nm repeat has no helical component and originates from myosin layers closer to the filament midpoint associated with the titin super-repeat in that region. Cooling relaxed or resting muscle, which partially mimics the effects of calcium activation on thick filament structure, disrupts the helical order of the myosin motors, and they move out from the filament backbone. Compression of the filament lattice of demembranated fibers by 5% Dextran, which restores interfilament spacing to that in intact muscle, stabilizes the higher-temperature structure. The axial periodicity of the filament backbone increases on cooling, but in lattice-compressed fibers the periodicity of the myosin heads does not follow the extension of the backbone. Thick filament structure in lattice-compressed demembranated fibers at near-physiological temperature is similar to that in intact resting muscle, suggesting that the native structure of the thick filament is largely preserved after demembranation in these conditions, although not in the conditions used for most previous studies with this preparation.
AB - Contraction of skeletal muscle is regulated by structural changes in both actin-containing thin filaments and myosin-containing thick filaments, but myosin-based regulation is unlikely to be preserved after thick filament isolation, and its structural basis remains poorly characterized. Here, we describe the periodic features of the thick filament structure in situ by high-resolution small-angle x-ray diffraction and interference. We used both relaxed demembranated fibers and resting intact muscle preparations to assess whether thick filament regulation is preserved in demembranated fibers, which have been widely used for previous studies. We show that the thick filaments in both preparations exhibit two closely spaced axial periodicities, 43.1 nm and 45.5 nm, at near-physiological temperature. The shorter periodicity matches that of the myosin helix, and x-ray interference between the two arrays of myosin in the bipolar filament shows that all zones of the filament follow this periodicity. The 45.5-nm repeat has no helical component and originates from myosin layers closer to the filament midpoint associated with the titin super-repeat in that region. Cooling relaxed or resting muscle, which partially mimics the effects of calcium activation on thick filament structure, disrupts the helical order of the myosin motors, and they move out from the filament backbone. Compression of the filament lattice of demembranated fibers by 5% Dextran, which restores interfilament spacing to that in intact muscle, stabilizes the higher-temperature structure. The axial periodicity of the filament backbone increases on cooling, but in lattice-compressed fibers the periodicity of the myosin heads does not follow the extension of the backbone. Thick filament structure in lattice-compressed demembranated fibers at near-physiological temperature is similar to that in intact resting muscle, suggesting that the native structure of the thick filament is largely preserved after demembranation in these conditions, although not in the conditions used for most previous studies with this preparation.
UR - http://www.scopus.com/inward/record.url?scp=85099698377&partnerID=8YFLogxK
U2 - 10.1085/jgp.202012713
DO - 10.1085/jgp.202012713
M3 - Article
C2 - 33416833
AN - SCOPUS:85099698377
SN - 0022-1295
VL - 153
JO - The Journal of general physiology
JF - The Journal of general physiology
IS - 3
M1 - e202012713
ER -