Abstract
Copy number variants and point mutations of NEPH2 (also called KIRREL3) gene encoding an immunoglobulin (Ig) superfamily adhesion molecule have been linked to autism spectrum disorders, intellectual disability and neurocognitive delay associated with Jacobsen syndrome, but the physiological roles of Neph2 in the mammalian brain remain largely unknown. Neph2 is highly expressed in the dentate granule (DG) neurons of the hippocampus and is localized in both dendrites and axons. It was recently shown that Neph2 is required for the formation of mossy fiber filopodia, the axon terminal structure of DG neurons forming synapses with GABAergic neurons of CA3. In contrast, however, it is unknown whether Neph2 also has any roles in the postsynaptic compartments of DG neurons. We here report that, through its C-terminal PDZ domain-binding motif, Neph2 directly interacts with postsynaptic density (PSD)-95, an abundant excitatory postsynaptic scaffolding protein. Moreover, Neph2 protein is detected in the brain PSD fraction and interacts with PSD-95 in synaptosomal lysates. Functionally, loss of Neph2 in mice leads to age-specific defects in the synaptic connectivity of DG neurons. Specifically, Neph2−/− mice show significantly increased spontaneous excitatory synaptic events in DG neurons at postnatal week 2 when the endogenous Neph2 protein expression peaks, but show normal excitatory synaptic transmission at postnatal week 3. The evoked excitatory synaptic transmission and synaptic plasticity of medial perforant pathway (MPP)-DG synapses are also normal in Neph2−/− mice at postnatal week 3, further confirming the age-specific synaptic defects. Together, our results provide some evidence for the postsynaptic function of Neph2 in DG neurons during the early postnatal period, which might be implicated in neurodevelopmental and cognitive disorders caused by NEPH2 mutations.
Highlights
When coexpressed in heterologous cells, Neph2 and postsynaptic density (PSD)-95 were colocalized in discrete intracellular clusters in PDZ interaction-dependent manner, as shown by the loss of coclustering in the mutant Neph2 where the C-terminal PDZ-binding motif was blocked by FLAG tagging (Figure 1E)
These results indicate that Neph2 directly interacts with PSD-95 through its C-terminal PDZ-binding motif
We show some evidence indicating the postsynaptic localization of Neph2 proteins in the mammalian brain
Summary
Synaptic adhesion molecules play diverse roles in synaptic development and function, including synapse specificity, formation, maturation and plasticity (Shen and Scheiffele, 2010; Tallafuss et al, 2010; Williams et al, 2010; Nam et al, 2011; Siddiqui and Craig, 2011; Yuzaki, 2011; Missler et al, 2012; Valnegri et al, 2012; Takahashi and Craig, 2013; Um and Ko, 2013; Yogev and Shen, 2014; de Wit and Ghosh, 2014, 2016; Han et al, 2016). CASK is a presynaptic scaffolding protein implicated in X-linked brain malformation and intellectual disability (Hsueh, 2006; Najm et al, 2008) suggesting the potential roles of the Neph protein complex in synaptic development and function (Bhalla et al, 2008)
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