ACETYLCHOLINE-RECEPTOR CLUSTERING ASSOCIATES WITH PROTEOGLYCAN BIOSYNTHESIS IN C2 VARIANT AND HETERKARYON MUSCLE-CELLS

Several lines of evidence have suggested roles for proteoglycans (PGs) in acetylcholine receptor (AChR) clustering on muscle cells. One line of evidence comes from the correlation between a defect in the biosyn thesis of glycosaminoglycans (GAGs), the defining carbohydrates of PGs , and the failure of spontaneous AChR clustering in the S27 cell line, a genetic variant of the C2 muscle cell line. Two approaches were use d in the present study to investigate whether GAG and AChR clustering defects are causally linked. First, the formation of AChR clusters was examined in two more variant lines, S11 and S26, also isolated from t he C2 muscle cell line on the basis of deficiencies in GAG biosynthesi s. S11 and S26, like S27, are also defective in AChR clustering. Ion e xchange analysis of the GAGs made by the S11, S26, and S27 lines revea led that the defects in GAG biosynthesis differ between the three line s. Second, heterokaryon myotubes formed between pairs of the GAG defec tive variants were tested for complementation in both AChR clustering and GAG biosynthesis. AChR clusters were conspicuous on individual het erokaryon myotubes, and GAG biosynthesis was restored to near wild typ e levels in the heterokaryon cultures. Complementation in GAG biosynth esis corroborates the biochemical data that the relevant mutations in the genetic variants are in different genes and establishes that the d efects are not dominant. The consistent correlation between GAG defect s and the failure of AChR clustering across three independent genetic variants and the complementary association of GAG biosynthesis with AC hR clustering in heterokaryon myotubes argues against a chance relatio nship between PGs and AChR clustering. A prominent chondroitin sulfate peak correlated with AChR clustering in the heterokaryon cultures. Th is is consistent with earlier results suggesting that chondroitin sulf ate in general is required for the spontaneous clustering of AChRs in C2 cultures and further suggests that a particular chondroitin sulfate proteoglycan may be essential for the clustering process. (C) 1996 Jo hn Wiley & Sons, Inc.