Departmental Papers (ASC)

Document Type

Technical Report

Date of this Version

3-2007

Publication Source

Journal of Biological Chemistry

Volume

282

Issue

13

Start Page

9932

Last Page

9940

DOI

10.1074/jbc.M608714200

Abstract

Rapsyn, a cytoplasmic receptor-associated protein, is required for the clustering of acetylcholine receptors (AChRs). Although AChR dynamics have been extensively studied, little is known about the dynamics of rapsyn. Here, we used a rapsyn-green fluorescent protein (GFP) fusion protein and quantitative fluorescent imaging to study the dynamics of rapsyn in transfected C2C12 myotubes. First, we found that rapsyn-GFP expression at clusters did not alter AChR aggregation, function, or turnover. Quantification of rapsyn immunofluorescence indicated that the expression of rapsyn-GFP proteins at clusters does not increase the overall rapsyn density compared with untransfected myotube clusters. Using time lapse imaging and fluorescence recovery after photobleaching, we demonstrated that the recovery of rapsyn-GFP fluorescence at clusters was very fast, with a halftime of about ∼1.5 h (∼3 times faster than AChRs). Inhibition of protein kinase C significantly altered receptor insertion, but it had no effect on rapsyn insertion. When cells were treated with the broad spectrum kinase inhibitor staurosporine, receptor insertion was decreased even further. However, inhibition of protein kinase A had no effect on insertion of either rapsyn or receptors. Finally, when cells were treated with neural agrin, rapsyn and AChRs were both directed away from preexisting clusters and accumulated together in new small clusters. These results demonstrate the remarkable dynamism of rapsyn, which may underlie the stability and maintenance of the postsynaptic scaffold and suggest that the insertion of different postsynaptic proteins may be operating independently.

Copyright/Permission Statement

This research was originally published in the Journal of Biological Chemistry. Bruneau, E. & Akaaboune, M. The Dynamics of the Rapsyn Scaffolding Protein at Individual Acetylcholine Receptor Clusters. Journal of Biological Chemistry. 2007; 282(13): 9932-40. © the American Society for Biochemistry and Molecular Biology.

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Date Posted: 15 June 2018

This document has been peer reviewed.