Marangi P A, Forsayeth J R, Mittaud P, Erb-Vögtli S, Blake D J, Moransard M, Sander A, Fuhrer C
Department of Neurochemistry, Brain Research Institute, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.
EMBO J. 2001 Dec 17;20(24):7060-73. doi: 10.1093/emboj/20.24.7060.
We have investigated the role of acetylcholine receptors (AChRs) in an early step of postsynaptic assembly at the neuromuscular synapse, the clustering of postsynaptic proteins induced by nerve-released agrin. To achieve this, we used two variants of C2 myotubes virtually lacking AChRs and C2 cells in which surface AChRs were down-regulated by AChR antibodies. In all cases, agrin caused normal clustering of the agrin receptor component MuSK, alpha-dystrobrevin and utrophin, but failed to aggregate AChRs, alpha- and beta-dystroglycan, syntrophin isoforms and rapsyn, an AChR-anchoring protein necessary for postsynaptic assembly and AChR clustering. In C2 variants, the stability of rapsyn was decreased, whereas in antibody-treated cells, rapsyn efficiently co-localized with remaining AChRs in microaggregates. Upon ectopic injection into myofibers in vivo, rapsyn did not form clusters in the absence of AChRs. These results show that AChRs and rapsyn are interdependent components of a pre-assembled protein complex that is required for agrin-induced clustering of a full set of postsynaptic proteins, thus providing evidence for an active role of AChRs in postsynaptic assembly.
我们研究了乙酰胆碱受体（AChRs）在神经肌肉突触后突触组装早期步骤中的作用，即神经释放的聚集蛋白诱导的突触后蛋白聚集。为了实现这一目标，我们使用了两种变体：几乎缺乏AChRs的C2肌管和通过AChR抗体下调表面AChRs的C2细胞。在所有情况下，聚集蛋白都会导致聚集蛋白受体成分MuSK、α-肌营养不良蛋白短链和肌养蛋白正常聚集，但无法使AChRs、α-和β-肌营养不良聚糖、肌营养不良蛋白异构体和rapsyn（一种突触后组装和AChR聚集所必需的AChR锚定蛋白）聚集。在C2变体中，rapsyn的稳定性降低，而在抗体处理的细胞中，rapsyn在微聚集体中与剩余的AChRs有效共定位。在体内异位注射到肌纤维后，在没有AChRs的情况下，rapsyn不会形成簇。这些结果表明，AChRs和rapsyn是预组装蛋白复合物的相互依赖成分，该复合物是聚集蛋白诱导全套突触后蛋白聚集所必需的，从而为AChRs在突触后组装中的积极作用提供了证据。
