title: Nebulin alters cross-bridge cycling kinetics and increases thin filament activation: a novel mechanism for increasing tension and reducing tension cost.
abstract: Nebulin is a giant filamentous F-actin-binding protein ( approximately 800 kDa) that binds along the thin filament of the skeletal muscle sarcomere. Nebulin is one of the least well understood major muscle proteins. Although nebulin is usually viewed as a structural protein, here we investigated whether nebulin plays a role in muscle contraction by using skinned muscle fiber bundles from a nebulin knock-out (NEB KO) mouse model. We measured force-pCa (-log[Ca(2+)]) and force-ATPase relations, as well as the rate of tension re-development (k(tr)) in tibialis cranialis muscle fibers. To rule out any alterations in troponin (Tn) isoform expression and/or status of Tn phosphorylation, we studied fiber bundles that had been reconstituted with bacterially expressed fast skeletal muscle recombinant Tn. We also performed a detailed analysis of myosin heavy chain, myosin light chain, and myosin light chain 2 phosphorylation, which showed no significant differences between wild type and NEB KO. Our mechanical studies revealed that NEB KO fibers had increased tension cost (5.9 versus 4.4 pmol millinewtons(-1) mm(-1) s(-1)) and reductions in k(tr) (4.7 versus 7.3 s(-1)), calcium sensitivity (pCa(50) 5.74 versus 5.90), and cooperativity of activation (n(H) 3.64 versus 4.38). Our findings indicate the following: 1) in skeletal muscle nebulin increases thin filament activation, and 2) through altering cross-bridge cycling kinetics, nebulin increases force and efficiency of contraction. These novel properties of nebulin add a new level of understanding of skeletal muscle function and provide a mechanism for the severe muscle weakness in patients with nebulin-based nemaline myopathy.
authors.affiliation: Department of Physiology, Sarver Molecular Cardiovascular Research Program, University of Arizona, Tucson, Arizona 85724-5217, USA.
authors.forename: Murali
authors.initials: M
authors.lastname: Chandra
authors.forename: Ranganath
authors.initials: R
authors.lastname: Mamidi
authors.forename: Steven
authors.initials: S
authors.lastname: Ford
authors.forename: Carlos
authors.initials: C
authors.lastname: Hidalgo
authors.forename: Christian
authors.initials: C
authors.lastname: Witt
authors.forename: Coen
authors.initials: C
authors.lastname: Ottenheijm
authors.forename: Siegfried
authors.initials: S
authors.lastname: Labeit
authors.forename: Henk
authors.initials: H
authors.lastname: Granzier
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D018345:Mice, Knockout; D009119:Muscle Contraction; D018485:Muscle Fibers, Skeletal / Q000737:chemistry / Q000502:physiology; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D009129:Muscle Tonus; D012518:Sarcomeres / Q000737:chemistry* / Q000502:physiology*
chemical_list: D009124:Muscle Proteins; C035620:nebulin; D002118:Calcium