|
|
|
|
|
|
|
|
|||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | |||||
First published online 6 January 2004
doi: 10.1242/jcs.00899
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Research Article |
Department of Pathology and Laboratory Medicine, Robert Wood Johnson Medical School, University of Medicine and Dentistry of New Jersey, Piscataway, NJ 08854, USA
* Author for correspondence (e-mail: daw{at}pleiad.umdnj.edu)
Accepted 18 September 2003
De novo folding and assembly of striated muscle myosin was analyzed by expressing a GFP-tagged embryonic myosin heavy chain (GFP-myosin) in post-mitotic C2C12 myocytes using replication defective adenoviruses. In the early stages of muscle differentiation, the GFP-myosin accumulates in bright globular foci and short filamentous structures that are later replaced by brightly fluorescent myofibrils. Time-lapse microscopy shows that the intermediates are dynamic and are present in elongating and fusing myocytes and in multinucleated myotubes. Immunostaining reveals the co-localization of the molecular chaperones Hsc70 and Hsp90 with the GFP-myosin in the intermediates, but not in the mature myofibrils. Uninfected cells have similar intermediates suggesting a common pathway for myosin maturation. Two conformation-sensitive antibodies that bind the unfolded motor domain and the coiled-coil conformation of the rod demonstrate that in the intermediates, the myosin rod is folded but the motor domain is not folded. Electron microscopy reveals that the intermediates contain loose filament bundles surrounded by a protein rich matrix. Geldanamycin, a specific inhibitor of Hsp90, reversibly blocks myofibril assembly and triggers accumulation of myosin folding intermediates. We conclude that multimeric complexes of nascent myosin filaments associated with Hsc70 and Hsp90 are intermediates in the folding and assembly pathway of muscle myosin.
Key words: Myosin, Muscle, Folding, Molecular chaperone, GFP
This article has been cited by other articles:
|
|
 |
|
  L. Liu, R. Srikakulam, and D. A. Winkelmann Unc45 Activates Hsp90-dependent Folding of the Myosin Motor Domain J. Biol. Chem., May 9, 2008; 283(19): 13185 - 13193. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Etard, U. Roostalu, and U. Strahle Shuttling of the chaperones Unc45b and Hsp90a between the A band and the Z line of the myofibril J. Cell Biol., March 24, 2008; 180(6): 1163 - 1175. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Y. Boateng and P. H. Goldspink Assembly and maintenance of the sarcomere night and day Cardiovasc Res, March 1, 2008; 77(4): 667 - 675. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. J. Du, H. Li, Y. Bian, and Y. Zhong Heat-shock protein 90{alpha}1 is required for organized myofibril assembly in skeletal muscles of zebrafish embryos PNAS, January 15, 2008; 105(2): 554 - 559. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Watanabe, S. Kondo, M. Hayasaka, and K. Hanaoka Functional analysis of homeodomain-containing transcription factor Lbx1 in satellite cells of mouse skeletal muscle J. Cell Sci., December 1, 2007; 120(23): 4178 - 4187. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Landsverk, S. Li, A. H. Hutagalung, A. Najafov, T. Hoppe, J. M. Barral, and H. F. Epstein The UNC-45 chaperone mediates sarcomere assembly through myosin degradation in Caenorhabditis elegans J. Cell Biol., April 23, 2007; 177(2): 205 - 210. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. Agbulut, A. Huet, N. Niederlander, M. Puceat, P. Menasche, and C. Coirault Green Fluorescent Protein Impairs Actin-Myosin Interactions by Binding to the Actin-binding Site of Myosin J. Biol. Chem., April 6, 2007; 282(14): 10465 - 10471. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Hamelin, T. Sayd, C. Chambon, J. Bouix, B. Bibe, D. Milenkovic, H. Leveziel, M. Georges, A. Clop, P. Marinova, et al. Proteomic analysis of ovine muscle hypertrophy J Anim Sci, December 1, 2006; 84(12): 3266 - 3276. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Chang, Y. Shav-Tal, T. Trcek, R. H. Singer, and R. D. Goldman Assembling an intermediate filament network by dynamic cotranslation J. Cell Biol., February 27, 2006; 172(5): 747 - 758. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Seo, K. Lee, K. Park, K. Bae, and I. Choi A Proteomic Assessment of Muscle Contractile Alterations during Unloading and Reloading J. Biochem., January 1, 2006; 139(1): 71 - 80. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Mishra, V. M. D'souza, K. C. Chang, Y. Huang, and M. K. Balasubramanian Hsp90 Protein in Fission Yeast Swo1p and UCS Protein Rng3p Facilitate Myosin II Assembly and Function Eukaryot. Cell, March 1, 2005; 4(3): 567 - 576. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Lord and T. D. Pollard UCS protein Rng3p activates actin filament gliding by fission yeast myosin-II J. Cell Biol., October 25, 2004; 167(2): 315 - 325. [Abstract] [Full Text] [PDF]
|
|
