TY - JOUR
T1 - Nebulin stiffens the thin filament and augments cross-bridge interaction in skeletal muscle
AU - Kiss, Balázs
AU - Lee, Eun Jeong
AU - Ma, Weikang
AU - Li, Frank W.
AU - Tonino, Paola
AU - Mijailovich, Srboljub M.
AU - Irving, Thomas C.
AU - Granzier, Henk L.
N1 - Funding Information:
ACKNOWLEDGMENTS. This research was supported by Grants R01AR053897 and R01AR073179 from the NIH/National Institute of Arthritis and Musculoskeletal and Skin Diseases and used resources of the Advanced Photon Source, operated under Department of Energy Contract DE-AC02-06CH11357 and supported by Grant 9 P41 GM103622 from the NIH/National Institute of General Medical Sciences (NIGMS). Use of the Pilatus 3 1M detector was provided by Grant 1S10OD018090-01 from the NIH/NIGMS. H.L.G. is the Allan and Alfie Norville Endowed Chair for Heart Disease in Women Research.
Publisher Copyright:
© 2018 National Academy of Sciences. All rights reserved.
PY - 2018/10/9
Y1 - 2018/10/9
N2 - Nebulin is a giant sarcomeric protein that spans along the actin filament in skeletal muscle, from the Z-disk to near the thin filament pointed end. Mutations in nebulin cause muscle weakness in nemaline myopathy patients, suggesting that nebulin plays important roles in force generation, yet little is known about nebulin's influence on thin filament structure and function. Here, we used small-angle X-ray diffraction and compared intact muscle deficient in nebulin (using a conditional nebulin-knockout, Neb cKO) with control (Ctrl) muscle. When muscles were activated, the spacing of the actin subunit repeat (27 Å) increased in both genotypes; when converted to thin filament stiffness, the obtained value was 30 pN/nm in Ctrl muscle and 10 pN/nm in Neb cKO muscle; that is, the thin filament was approximately threefold stiffer when nebulin was present. In contrast, the thick filament stiffness was not different between the genotypes. A significantly shorter left-handed (59 Å) thin filament helical pitch was found in passive and contracting Neb cKO muscles, as well as impaired tropomyosin and troponin movement. Additionally, a reduced myosin mass transfer toward the thin filament in contracting Neb cKO muscle was found, suggesting reduced crossbridge interaction. We conclude that nebulin is critically important for physiological force levels, as it greatly stiffens the skeletal muscle thin filament and contributes to thin filament activation and cross-bridge recruitment.
AB - Nebulin is a giant sarcomeric protein that spans along the actin filament in skeletal muscle, from the Z-disk to near the thin filament pointed end. Mutations in nebulin cause muscle weakness in nemaline myopathy patients, suggesting that nebulin plays important roles in force generation, yet little is known about nebulin's influence on thin filament structure and function. Here, we used small-angle X-ray diffraction and compared intact muscle deficient in nebulin (using a conditional nebulin-knockout, Neb cKO) with control (Ctrl) muscle. When muscles were activated, the spacing of the actin subunit repeat (27 Å) increased in both genotypes; when converted to thin filament stiffness, the obtained value was 30 pN/nm in Ctrl muscle and 10 pN/nm in Neb cKO muscle; that is, the thin filament was approximately threefold stiffer when nebulin was present. In contrast, the thick filament stiffness was not different between the genotypes. A significantly shorter left-handed (59 Å) thin filament helical pitch was found in passive and contracting Neb cKO muscles, as well as impaired tropomyosin and troponin movement. Additionally, a reduced myosin mass transfer toward the thin filament in contracting Neb cKO muscle was found, suggesting reduced crossbridge interaction. We conclude that nebulin is critically important for physiological force levels, as it greatly stiffens the skeletal muscle thin filament and contributes to thin filament activation and cross-bridge recruitment.
KW - Muscle biology
KW - Physiology
KW - Skeletal myopathy
KW - X-ray diffraction
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U2 - 10.1073/pnas.1804726115
DO - 10.1073/pnas.1804726115
M3 - Article
C2 - 30249654
AN - SCOPUS:85053084942
SN - 0027-8424
VL - 115
SP - 10369
EP - 10374
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 41
ER -