Extracellular matrix (ECM) proteins and their receptors have important roles during synapse assembly, development and plasticity, but very little is known about the underlying molecular mechanisms. In a genetic lethality screen for candidates interacting with neto, an obligatory auxiliary protein at the Drosophila NMJ, we found tenectin (tnc). Tnc is a mucin-like ECM protein expressed in motor neurons and striated muscles in 3rd instar larvae. tnc mutants are partially lethal and the escapers have severe climbing and locomotor defects, suggesting that Tnc is required for NMJ development. Using genetics, histology, super-resolution imaging, electron microscopy and electrophysiology approaches, we found that tnc mutants have collapsed synaptic boutons, but normal synaptic contacts and postsynaptic densities. Instead, Tnc is secreted into the synaptic cleft, and binds to pre- and postsynaptic βPS-containing integrin receptors. These Tnc-βPS bridges appear to stabilize the bouton shape partly by recruiting the membrane skeleton locally. Indeed, the distribution of membrane skeleton components such as α-Spectrin, Adducin, Ank2L are disrupted at Tnc-deprived NMJs. Furthermore, Tnc-coated beads recruit βPS and α-Spectrin to specific membrane domains in S2R+ cells.
Tnc is also involved in the regulation of synaptic vesicle release. Electrophysiology recordings show that tnc mutants NMJs have reduced mEJPs frequency, EJPs amplitude and quantal content, and exhibit higher paired-pulse ratio (PPR). Expression of Tnc in motor neurons, but not in muscles, can rescue the synaptic accumulation of βPS integrin,α-Spectrin and Adducin and partially rescue the behavior defects of tnc mutants. Together, our data demonstrate that pre- and postsynaptic secreted Tnc recruits integrin receptors at NMJs, and anchors the synaptic membrane skeleton to the ECM to ensure synaptic boutons architecture and function. Our findings extend the knowledge on how ECM proteins organize the synaptic membrane skeleton to regulate synaptic structure and function. .