ATP potentiates agrin-induced AChR aggregation in cultured myotubes: activation of RhoA in P2Y1 nucleotide receptor signaling at vertebrate neuromuscular junctions.
At vertebrate neuromuscular junctions, ATP is known to stabilize acetylcholine in the synaptic vesicles and to be co-released with it. We have shown previously that a nucleotide receptor, P2Y(1) receptor, is localized at the nmjs, and we propose that this mediates a trophic role for synaptic ATP there. In cultured myotubes, the activation of P2Y(1) receptors modulated agrin-induced acetylcholine receptor (AChR) aggregation in a potentiation manner. This potentiation effect in agrin-induced AChR aggregation was reduced by antagonizing the P2Y(1) receptors. The guanosine triphosphatase RhoA was shown to be responsible for this P2Y(1)-potentiated effect. The localization of RhoA in rat and chicken skeletal muscles was restricted at the neuromuscular junctions. Application of P2Y(1) agonists in cultured myotubes induced RhoA activation, which showed an additive effect with agrin-induced RhoA activation. Over-expression of dominant-negative mutant of RhoA in cultured myotubes diminished the agrin-induced AChR aggregation, as well as the potentiation effect of P2Y(1)-specific agonist. Application of UTP in the cultures also triggered similar responses as did 2-methylthioadenosine 5'-diphosphate, suggesting the involvement of other subtypes of P2Y receptors. These results demonstrate that RhoA could serve as a downstream mediator of signaling mediated by P2Y(1) receptor and agrin, which therefore synergizes the effects of the two neuron-derived trophic factors in modulating the formation and/or maintenance of post-synaptic apparatus at the neuromuscular junctions.
Ling
Karen K Y
KK
Department of Biology and Molecular Neuroscience Center, Hong Kong University of Science and Technology, Hong Kong, China.
Siow
Nina L
NL
Choi
Roy C Y
RC
Ting
Annie K L
AK
Kong
Ling W
LW
Tsim
Karl W K
KW
D000255:Adenosine Triphosphate / Q000008:administration & dosage* / Q000378:metabolism; D018171:Agrin / Q000008:administration & dosage* / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D002645:Chickens; D004357:Drug Synergism; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000187:drug effects / Q000378:metabolism; D009154:Mutation; D009469:Neuromuscular Junction / Q000187:drug effects / Q000378:metabolism; D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D051381:Rats; D011950:Receptors, Cholinergic / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058472:Receptors, Purinergic P2Y1; D015398:Signal Transduction; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
D018171:Agrin; C546384:P2ry1 protein, rat; D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D011950:Receptors, Cholinergic; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D000255:Adenosine Triphosphate; D020742:rhoA GTP-Binding Protein