Kirrel3-mediated synapse formation is attenuated by disease-associated missense variants

Missense variants in Kirrel3 are repeatedly identified as risk factors for autism spectrum disorder and intellectual disability but it has not been reported if or how these variants disrupt Kirrel3 function. Previously, we studied Kirrel3 loss-of-function using knockout mice and showed that Kirrel3 is a synaptic adhesion molecule necessary to form one specific type of hippocampal synapse in vivo. Here, we developed a new gain-of-function assay for Kirrel3 and find that wild-type Kirrel3 induces synapse formation selectively between Kirrel3-expressing cells via homophilic, trans-cellular binding. We tested six disease-associated Kirrel3 missense variants and find that five attenuate this synaptogenic function. All variants tested traffic to the cell surface and localize to synapses similar to wild-type Kirrel3. Two tested variants lack homophilic trans-cellular binding, which likely accounts for their reduced synaptogenic function. Interestingly, we also identified variants that bind in trans but cannot induce synapses, indicating Kirrel3 trans-cellular binding is necessary but not sufficient for its synaptogenic function. Collectively, these results suggest Kirrel3 functions as a synaptogenic, cell-recognition molecule, and this function is attenuated by missense variants associated with autism spectrum disorder and intellectual disability. Thus, we provide critical insight to Kirrel3 function in typical brain development and the consequences of missense variants associated with autism spectrum disorder and intellectual disability. SIGNIFICANCE STATEMENTHere, we advance our understanding of mechanisms mediating target-specific synapse formation by providing evidence that Kirrel3 trans-cellular interactions mediate contact recognition and signaling to promote synapse development. Moreover, this is the first study to test the effects of disease-associated Kirrel3 missense variants on synapse formation, and thereby, provides a framework to understand the etiology of complex neurodevelopmental disorders arising from rare missense variants in synaptic genes.