Latrophilin GPCRs direct synapse specificity by coincident binding of FLRTs and teneurins

INTRODUCTION In the brain, synaptic connections form neuronal communication networks, thereby constructing neural circuits. Synaptic connections are exquisitely specific and dynamic, but the underlying molecular mechanisms remain largely unexplored. In the hippocampus, Schaffer-collateral axons from the CA3 region form synapses on CA1-region pyramidal neurons exclusively on dendritic domains in the stratum (S.) oriens and S. radiatum of these neurons. In contrast, perforant-path axons from the entorhinal cortex form synapses on CA1-region pyramidal neurons exclusively on dendritic domains in the S. lacunosum-moleculare. However, it is unknown how this synaptic input specificity is achieved and what signaling mechanisms maintain the two classes of synapses. RATIONALE Synapse formation is thought to involve bidirectional signaling by transsynaptic cell adhesion molecules. Building on recent observations that the adhesion G protein–coupled receptor (GPCR) latrophilin-2 is essential for synapses in the S. lacunosum-moleculare of the CA1 region, we asked whether distinct latrophilins are localized to different dendritic domains of CA1-region neurons. Moreover, latrophilins are known to form transcellular interactions with two classes of cell adhesion molecules: teneurins and fibronectin leucine-rich repeat transmembrane proteins (FLRTs). Thus, we hypothesized that latrophilins may act in synapse formation via transsynaptic interactions with these adhesion molecules as ligands, and that such interactions may contribute to the specificity of synapse formation. RESULTS We produced genetic manipulations in transgenic mice to allow monitoring the localizations of endogenous latrophilin-2 and latrophilin-3 in vivo and to enable their conditional deletion. Using these manipulations, we found that latrophilin-2 and latrophilin-3 were specifically localized to postsynaptic spines in nonoverlapping dendritic domains of CA1-region pyramidal neurons. Latrophilin-2 was targeted only to excitatory synapses in the S. lacunosum-moleculare, whereas latrophilin-3 was targeted only to excitatory synapses in the S. oriens and S. radiatum, corresponding to distinct presynaptic inputs onto CA1-region pyramidal neurons. Deletion of latrophilin-3 selectively decreased Schaffer-collateral synapses in the S. radiatum and S. oriens, whereas deletion of latrophilin-2 selectively decreased entorhinal cortex–derived synapses in the S. lacunosum-moleculare of CA1 neurons. In vivo rescue experiments with latrophilin-3 mutants that selectively lack binding to only FLRTs or only teneurins revealed that both binding activities were required for input-specific synapse formation, as monitored by electrophysiology and retrograde rabies tracing. Thus, coincident binding of both latrophilin-3 ligands was necessary for synapse formation. Moreover, in vitro synapse formation assays showed that teneurin-2 or FLRT3 alone were unable to induce excitatory synapse formation, whereas together they potently did so. However, even in combination, FLRT3 and teneurin-2 induced excitatory synapses only when teneurin-2 was expressed as a splice variant that is competent to interact with latrophilins, indicating that simultaneous binding of both FLRT3 and teneurin-2 to latrophilins was necessary to induce synapse formation. CONCLUSION We suggest that latrophilin-2 and latrophilin-3 are postsynaptic adhesion GPCRs that are targeted in CA1 pyramidal neurons to nonoverlapping dendritic domains, where they promote excitatory synapse formation by specific and distinct presynaptic inputs. Because the function of latrophilin-3 in synapse formation requires simultaneous binding of two unrelated presynaptic ligands (FLRTs and teneurins), a coincidence signaling mechanism could account for the specificity of synaptic connections.