unplugged/MuSK signaling coordinates pre- and postsynaptic development at the neuromuscular junction

Development of the neuromuscular junction (NMJ) is divided into two stages: an early stage when acetylcholine receptors (AChRs) accumulate into 'prepatterned' clusters at the center of muscle; and a later stage when the nerve contacts the muscle and induces neuromuscular synapses. At the later stage, nerve-derived agrin acts through unplugged/MuSK (muscle specific receptor kinase) to promote synapse formation. At the early stage, unplugged /MuSK induces AChR prepatterning independently of agrin and the nerve. This has raised the question how unplugged/MuSK is activated at the early stage. I have taken a genetic approach in zebrafish to identify a hitherto unknown ligand for unplugged/MuSK.;I show that morpholino-mediated knockdown of wnt11r results in pre- and postsynaptic defects at the early stage identical to those in unplugged mutants. I also show that wnt11r and unplugged/MuSK interact genetically and biochemically and that co-overexpression of wnt11r and unplugged /MuSK increases AChR prepatterning. Furthermore, I demonstrate that noncanonical disheveled signaling in muscle is required for unplugged /MuSK function. I propose that wnt ligands activate unplugged/MuSK to organize a central muscle zone to which the growth cones and AChR prepatterns are restricted through a mechanism reminiscent of the planar cell polarity pathway.;Using heat-shock inducible transgenes, I further demonstrate that the central muscle zone is dispensable for the formation of neuromuscular synapses, but essential for growth cone guidance. Thus, I propose that wnt-unplugged /MuSK polarizes a region in middle of muscle accessible for motor nerve, thereby determining the synaptic sites.;Finally, using the same transgenes, I show that despite its early expression in muscles, unplugged/MuSK activity is first required just prior to the appearance of AChR prepattern to simultaneously induce AChR accumulation and guide motor axons. I also demonstrate that the ubiquitous expression of unplugged/MuSK throughout the muscle membrane results in wildtype-like synaptogenesis. I propose that additional factor(s) restrict unplugged /MuSK signaling to the muscle center.;Based on these studies, I propose that unplugged/MuSK signaling employs distinct pathways during different stages to ensure the formation of precise synapses within the appropriate target field.