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Posttranslational control of membrane-skeleton (ankyrin and alpha beta- spectrin) assembly in early myogenesis

Nelson, W. James and Lazarides, Elias (1985) Posttranslational control of membrane-skeleton (ankyrin and alpha beta- spectrin) assembly in early myogenesis. Journal of Cell Biology, 100 (5). pp. 1726-1735. ISSN 0021-9525. http://resolver.caltech.edu/CaltechAUTHORS:NELjcb85

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Abstract

Adult chicken skeletal muscle cells express polypeptides that are antigenically related to alpha-spectrin (Mr 240,000) and beta-spectrin (Mr 220,000-225,000), the major components of the erythrocyte membrane- skeleton, and to ankyrin (Mr 237,000; also termed goblin in chicken erythrocytes), which binds spectrin to the transmembrane anion transporter in erythrocytes. Comparative immunoblotting of SDS- solubilized extracts of presumptive myoblasts and fully differentiated myotubes cultured in vitro demonstrated that there is a dramatic accumulation of ankyrin and alpha- and beta-spectrin during myogenesis and a concomitant switch in the subunit composition of spectrin from alpha gamma to alpha beta. Analysis of early time points in myogenesis (12-96 h) revealed that these changes occur shortly after the main burst of cell fusion. To determine the temporal relationship between cell fusion and the accumulation of ankyrin and alpha- and beta- spectrin, we treated presumptive myoblasts with 2 mM EGTA, which resulted in the complete inhibition of cell fusion. The incorporation of [35S]methionine into total protein and, specifically, into alpha-, gamma-, and beta-spectrin remained the same in EGTA-treated and control cells. Analysis by immunoblotting of the amounts of ankyrin and alpha- and beta-spectrin in fusion-blocked cells revealed that there was no effect on accumulation for the first 19 h. However, there was then a dramatic cessation in their accumulation, and thereafter, the amount of each protein at steady state remained constant. Upon release from the EGTA block, the cells fused rapidly (less than 11 h), and the accumulation of ankyrin and alpha- and beta-spectrin was reinitiated after a lag period of 3-5 h at a rate similar to that in control cells. The inhibition in the accumulation of newly synthesized ankyrin, alpha- spectrin, and beta-spectrin in EGTA-treated myoblasts was not characteristic of all structural proteins, since the accumulation of the muscle-specific intermediate filament protein desmin was the same in control and fusion-blocked cells. These results show that in myogenesis, the synthesis of ankyrin and alpha- and beta-spectrin and their accumulation as a complex, although concurrent, are not coupled events. We hypothesize that the extent of assembly of these components of the membrane-skeleton in muscle cells is determined by a control mechanism(s) operative at the posttranslational level that is triggered near the time of cell fusion and the onset of terminal differentiation.


Item Type:Article
Additional Information:Copyright © 1985 by The Rockefeller University Press. RUP grants the public the non-exclusive right to copy, distribute, or display the Work under a Creative Commons Attribution-Noncommercial-Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ and http://creativecommons.org/licenses/by-nc-sa/3.0/legalcode. Received for publication 18 December 1984, and in revised form 14 February 1985. We thank Ilga Lielausis for technical assistance with the cell culture, and John Ngai for helpful discussions and critical comments on the manuscript. This work was supported by grants from the National Institutes of Health, the National Science Foundation, and the Muscular Dystrophy Association of America. Dr. Nelson was supported also by a Senior Investigator Fellowship from the American Heart Association Los Angeles Affiliate and a grant from the National Institutes of Health. Dr. Lazarides is a recipient of a Research Career Development Award from the National Institutes of Health.
Record Number:CaltechAUTHORS:NELjcb85
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:NELjcb85
Alternative URL:http://www.jcb.org/cgi/content/abstract/100/5/1726
Usage Policy:RUP grants the public the non-exclusive right to copy, distribute, or display the Work under a Creative Commons Attribution-Noncommercial-Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/ and http://creativecommons.org/licenses/by-nc-sa/3.0/legalcode.
ID Code:10996
Collection:CaltechAUTHORS
Deposited By: Archive Administrator
Deposited On:22 Jun 2008
Last Modified:26 Dec 2012 10:07

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