Nuclear factor kappaB controls acetylcholine receptor clustering at the neuromuscular junction.
At the vertebrate neuromuscular junction (NMJ), acetylcholine receptor (AChR) clustering is stimulated by motor neuron-derived glycoprotein Agrin and requires a number of intracellular signal or structural proteins, including AChR-associated scaffold protein Rapsyn. Here, we report a role of nuclear factor kappaB (NF-kappaB), a well known transcription factor involved in a variety of immune responses, in regulating AChR clustering at the NMJ. We found that downregulating the expression of RelA/p65 subunit of NF-kappaB or inhibiting NF-kappaB activity by overexpression of mutated form of IkappaB (inhibitor kappaB), which is resistant to proteolytic degradation and thus constitutively keeps NF-kappaB inactive in the cytoplasma, impeded the formation of AChR clusters in cultured C2C12 muscle cells stimulated by Agrin. In contrast, overexpression of RelA/p65 promoted AChR clustering. Furthermore, we investigated the mechanism by which NF-kappaB regulates AChR clustering. Interestingly, we found that downregulating the expression of RelA/p65 caused a marked reduction in the protein and mRNA level of Rapsyn and upregulation of RelA/p65 enhanced Rapsyn promoter activity. Mutation of NF-kappaB binding site on Rapsyn promoter prevented responsiveness to RelA/p65 regulation. Moreover, forced expression of Rapsyn in RelA/p65 downregulated muscle cells partially rescued AChR clusters, suggesting that NF-kappaB regulates AChR clustering, at least partially through the transcriptional regulation of Rapsyn. In line with this notion, genetic ablation of RelA/p65 selectively in the skeletal muscle caused a reduction of AChR density at the NMJ and a decrease in the level of Rapsyn. Thus, NF-kappaB signaling controls AChR clustering through transcriptional regulation of synaptic protein Rapsyn.
Wang
Jia
J
Institute of Neuroscience and State Key Laboratory of Neuroscience, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China.
Fu
Xiu-Qing
XQ
Lei
Wen-Liang
WL
Wang
Tong
T
Sheng
Ai-Li
AL
Luo
Zhen-Ge
ZG
D018171:Agrin / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D005786:Gene Expression Regulation; D020672:I-kappa B Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism; D018482:Muscle, Skeletal / Q000254:growth & development / Q000378:metabolism; D009154:Mutation; D032446:Myoblasts / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D009469:Neuromuscular Junction / Q000254:growth & development / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D011950:Receptors, Cholinergic / Q000378:metabolism*; D051996:Transcription Factor RelA / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
D018171:Agrin; D020672:I-kappa B Proteins; D009124:Muscle Proteins; D016328:NF-kappa B; D012333:RNA, Messenger; D011950:Receptors, Cholinergic; C498144:Rela protein, mouse; D051996:Transcription Factor RelA; C051609:peripheral membrane protein 43K