A single pulse of agrin triggers a pathway that acts to cluster acetylcholine receptors.
Agrin triggers signaling mechanisms of high temporal and spatial specificity to achieve phosphorylation, clustering, and stabilization of postsynaptic acetylcholine receptors (AChRs). Agrin transiently activates the kinase MuSK; MuSK activation has largely vanished when AChR clusters appear. Thus, a tyrosine kinase cascade acts downstream from MuSK, as illustrated by the agrin-evoked long-lasting activation of Src family kinases (SFKs) and their requirement for AChR cluster stabilization. We have investigated this cascade and report that pharmacological inhibition of SFKs reduces early but not later agrin-induced phosphorylation of MuSK and AChRs, while inhibition of Abl kinases reduces late phosphorylation. Interestingly, SFK inhibition applied selectively during agrin-induced AChR cluster formation caused rapid cluster dispersal later upon agrin withdrawal. We also report that a single 5-min agrin pulse, followed by extensive washing, triggered long-lasting MuSK and AChR phosphorylation and efficient AChR clustering. Following the pulse, MuSK phosphorylation increased and, beyond a certain level, caused maximal clustering. These data reveal novel temporal aspects of tyrosine kinase action in agrin signaling. First, during AChR cluster formation, SFKs initiate early phosphorylation and an AChR stabilization program that acts much later. Second, a kinase mechanism rapidly activated by agrin acts thereafter autonomously in agrin's absence to further increase MuSK phosphorylation and cluster AChRs.
Mittaud
Peggy
P
Brain Research Institute, University of Zürich, Winterthurerstrasse 190, CH-8057 Zürich, Switzerland.
Camilleri
Alain A
AA
Willmann
Raffaella
R
Erb-Vögtli
Susanne
S
Burden
Steven J
SJ
Fuhrer
Christian
C
D018171:Agrin / Q000008:administration & dosage / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002999:Clone Cells; D004789:Enzyme Activation / Q000187:drug effects; D051379:Mice; D032446:Myoblasts / Q000187:drug effects / Q000378:metabolism; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D016315:Proto-Oncogene Proteins c-abl / Q000378:metabolism; D051076:Proto-Oncogene Proteins c-fyn; D011940:Receptor Aggregation / Q000187:drug effects*; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D011950:Receptors, Cholinergic / Q000187:drug effects* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013569:Synapses / Q000378:metabolism; D019061:src-Family Kinases / Q000737:chemistry / Q000378:metabolism
D018171:Agrin; D011518:Proto-Oncogene Proteins; D011950:Receptors, Cholinergic; D011994:Recombinant Proteins; C095714:MUSK protein, human; D020794:Receptor Protein-Tyrosine Kinases; C494960:Fyn protein, mouse; D016315:Proto-Oncogene Proteins c-abl; D051076:Proto-Oncogene Proteins c-fyn; D019061:src-Family Kinases