TY - JOUR T1 - Carrier of Wingless (Cow) Regulation of <em>Drosophila</em> Neuromuscular Junction Development JF - eneuro JO - eNeuro DO - 10.1523/ENEURO.0285-19.2020 SP - ENEURO.0285-19.2020 AU - Danielle L. Kopke AU - Shannon N. Leahy AU - Dominic J. Vita AU - Sofia C. Lima AU - Zachary L. Newman AU - Kendal Broadie Y1 - 2020/02/03 UR - http://www.eneuro.org/content/early/2020/02/03/ENEURO.0285-19.2020.abstract N2 - The first Wnt signaling ligand discovered, Drosophila Wingless (Wg; Wnt1 in mammals), plays critical roles in neuromuscular junction (NMJ) development, regulating synaptic architecture and function. Heparan sulfate proteoglycans (HSPGs), consisting of a core protein with heparan sulfate (HS) glycosaminoglycan (GAG) chains, bind to Wg ligands to control both extracellular distribution and intercellular signaling function. Drosophila HSPGs previously shown to regulate Wg trans-synaptic signaling at the NMJ include the glypican Dally-like Protein (Dlp) and perlecan Terribly Reduced Optic Lobes (Trol). Here, we investigate synaptogenic functions of the most recently described Drosophila HSPG, secreted Carrier of Wingless (Cow), which directly binds Wg in the extracellular space. At the glutamatergic NMJ, we find that Cow secreted from the presynaptic motor neuron acts to limit synaptic architecture and neurotransmission strength. In cow null mutants, we find increased synaptic bouton number and elevated excitatory current amplitudes, phenocopying presynaptic Wg overexpression. We show cow null mutants exhibit an increased number of glutamatergic synapses and increased synaptic vesicle (SV) fusion frequency based both on GCaMP imaging and electrophysiology recording. We find that membrane-tethered Wg prevents cow null defects in NMJ development, indicating that Cow mediates secreted Wg signaling. It was shown previously that the secreted Wg deacylase Notum restricts Wg signaling at the NMJ, and we show here that Cow and Notum work through the same pathway to limit synaptic development. We conclude Cow acts cooperatively with Notum to coordinate neuromuscular synapse structural and functional differentiation via negative regulation of Wg trans-synaptic signaling within the extracellular synaptomatrix.Significance Statement Wnt intercellular signaling is disrupted in numerous devastating neurological disorders, including Alzheimer’s disease. Therefore, an understanding of Wnt signaling regulation is important for the design and implementation of targeted treatments. As a disease model, the Drosophila glutamatergic NMJ system is large, accessible and genetically malleable, and thus well suited for discovering the molecular and cellular mechanisms of Wnt signaling regulation. Extracellular HSPGs are important players as regulators of Wnt intercellular signaling. Here, we show secreted HSPG Carrier of Wingless (Cow), which directly binds to the founding Wnt-1 ligand, regulates NMJ structure and function. The mammalian homolog of Cow, Testican-2, is highly expressed in the brain. Studying this HSPG in Drosophila should inform mechanisms of Wnt regulation in human brain. ER -