AbstractPostsynaptic density-95/Discs large/Zonula occludens-1 (PDZ) domain-mediated protein interactions play pivotal roles in various molecular biological events, including protein localisation, assembly and signal transduction. Although the vertebrate regulator of planar cell polarity Van Gogh-like 2 (Vangl2) was recently described as a postsynaptic molecule with a PDZ-binding motif, the role of its PDZ interaction at the synapse is unknown. In this report, we demonstrate that the PDZ interaction was dispensable for the normal cluster formation of Vangl2 and not absolutely required for the synapse-associated localisation of Vangl2 in cultured hippocampal neurons. We further showed that the synaptic localisation of Vangl2 was categorised into two types: overlapping co-localisation with postsynaptic density (PSD)-95 or highly correlated but complementary pattern of association with PSD-95. Only the former was significantly sensitive to deletion of the PDZ-binding motif. In addition, the PDZ interaction enhanced the protein interactions between PSD-95 and Prickle2, which is another planar cell polarity factor that is localised at the postsynaptic density. Taken together with our recent report that the density of PSD-95 clusters was reduced in Vangl2-silenced neurons, these results suggest that Vangl2 determines the complex formation and clustering of postsynaptic molecules for synaptogenesis in mammalian brains.