SUMMARY We report a novel mechanism of ribonucleoprotein (RNP) nucleocytoplasmic export by nuclear envelope budding. During development of Drosophila synapses, a fragment of the Wnt-1 receptor, DFrizzled2, is imported into postsynaptic nuclei where it forms prominent foci. We now show these foci to be composed of large RNP granules harboring synaptic protein transcripts. These RNPs exit the nucleus via a budding mechanism akin to nuclear egress of Herpes-type viruses, a process previously thought to be exclusive to these viruses. During this mechanism, RNP granules bud into the space between the inner and the outer nuclear membranes (the perinuclear space), in a manner dependent on Lamin C, a nuclear protein linked to muscular dystrophies. Like herpes virus nuclear egress, this process requires protein kinase C, which is known to disrupt the lamin through phosphorylation. We suggest that nuclear budding is an endogenous nuclear export pathway for large RNP granules.

Activity-dependent modifications in synapse structure play a key role in synaptic development and plasticity, but the signaling mechanisms involved are poorly understood. We demonstrate that glutamatergic Drosophila neuromuscular junctions undergo rapid changes in synaptic structure and function in response to patterned stimulation. These changes, which depend on transcription and translation, include formation of motile presynaptic filopodia, elaboration of undifferentiated varicosities, and potentiation of spontaneous release frequency. Experiments indicate that a bidirectional Wnt/Wg signaling pathway underlies these changes. Evoked activity induces Wnt1/Wg release from synaptic boutons, which stimulates both a postsynaptic DFz2 nuclear import pathway, as well as a presynaptic pathway involving GSK-3 β/Shaggy. Our findings suggest that bidirectional Wg signaling operates downstream of synaptic activity to induce modifications in synaptic structure and function. We propose that activation of the postsynaptic Wg pathway is required for the assembly of the postsynaptic apparatus, while activation of the presynaptic Wg pathway regulates cytoskeletal dynamics.

Wnts play pivotal roles during development and in the mature nervous system. However, the mechanism by which Wnts traffic between cells has remained elusive. Here we demonstrate a novel mechanism of Wnt transmission through release of exosome-like vesicles containing the Wnt-binding protein Evenness Interrupted/Wntless/Sprinter (Evi/Wls/Srt). We show that at the Drosophila larval neuromuscular junction (NMJ), presynaptic vesicular release of Evi is required for the secretion of the Wnt, Wingless (Wg). We also show that Evi acts cell-autonomously in the postsynaptic Wnt-receiving cell to target dGRIP, a Wg-receptor-interacting protein, to postsynaptic sites. Upon Evi loss of function, dGRIP is not properly targeted to synaptic sites, interfering with postsynaptic Wnt signal transduction. These findings uncover a previously unknown cellular mechanism by which a secreted Wnt is transported across synapses by Evi-containing vesicles, and reveal novel trafficking functions of Evi in both the Wnt-producing and the Wnt-receiving cell.

Arc/Arg3.1 is required for synaptic plasticity and cognition, and mutations in this gene are linked to autism and schizophrenia. Arc bears a domain resembling retroviral/retrotransposon Gag-like proteins, which multimerize into a capsid that packages viral RNA. The significance of such a domain in a plasticity molecule is uncertain. Here, we report that the Drosophila Arc1 protein forms capsid-like structures that bind darc1 mRNA in neurons and is loaded into extracellular vesicles that are transferred from motorneurons to muscles. This loading and transfer depends on the darc1-mRNA 3' untranslated region, which contains retrotransposon-like sequences. Disrupting transfer blocks synaptic plasticity, suggesting that transfer of dArc1 complexed with its mRNA is required for this function. Notably, cultured cells also release extracellular vesicles containing the Gag region of the Copia retrotransposon complexed with its own mRNA. Taken together, our results point to a trans-synaptic mRNA transport mechanism involving retrovirus-like capsids and extracellular vesicles.

Neurexins have been proposed to function as major mediators of the coordinated pre- and postsynaptic apposition. However, key evidence for this role in vivo has been lacking, particularly due to gene redundancy. Here, we have obtained null mutations in the single Drosophila neurexin gene (dnrx). dnrx loss of function prevents the normal proliferation of synaptic boutons at glutamatergic neuromuscular junctions, while dnrx gain of function in neurons has the opposite effect. DNRX mostly localizes to the active zone of presynaptic terminals. Conspicuously, dnrx null mutants display striking defects in synaptic ultrastructure, with the presence of detachments between pre- and postsynaptic membranes, abnormally long active zones, and increased number of T bars. These abnormalities result in corresponding alterations in synaptic transmission with reduced quantal content. Together, our results provide compelling evidence for an in vivo role of neurexins in the modulation of synaptic architecture and adhesive interactions between pre- and postsynaptic compartments.