The zinc finger domain of the 43-kDa receptor-associated protein, rapsyn: Role in acetylcholine receptor clustering

We injected rat myotubes with proteins and antibodies to assess the importance of the zinc finger (ZnF) domain of the 43-kDa receptor-associated protein, rapsyn, in clustering acetylcholine receptors (AChR). Injection of rat myotubes with a fusion protein containing the ZnF domain of rapsyn disrupted AChR clusters. Clusters were unaffected by a fusion protein containing a double mutant that does not bind zinc. Similar results were obtained with the purified wild type and mutant ZnF domains. The ZnF of HIV-1 nucleocapsid protein had no effect. AChR clusters were also disrupted in myotubes injected with antibodies to the ZnF domain, followed by injection of anti-antibodies. Injection of antibodies directed against a different rapsyn epitope or against the cytoplasmic domain of the AChR had no effect. In transfection experiments with HEK 293 cells, the ZnF domain failed to associate with membrane aggregates containing full-length rapsyn, AChR, or rapsyn and AChR together. We conclude that the ZnF domain of rapsyn provides a binding site essential for AChR clustering, but that this site is unlikely to be involved in high affinity binding of rapsyn to itself or to AChR.