Neurexin-3 subsynaptic densities are spatially distinct from Neurexin-1 and essential for excitatory synapse nanoscale organization in the hippocampus

Abstract

Proteins critical for synaptic transmission are non-uniformly distributed and assembled into regions of high density called subsynaptic densities (SSDs) that transsynaptically align in nanocolumns. Neurexin-1 and neurexin-3 are essential presynaptic adhesion molecules that non-redundantly control NMDAR- and AMPAR-mediated synaptic transmission, respectively, via transsynaptic interactions with distinct postsynaptic ligands. Despite their functional relevance, fundamental questions regarding the nanoscale properties of individual neurexins, their influence on the subsynaptic organization of excitatory synapses and the mechanisms controlling how individual neurexins engage in precise transsynaptic interactions are unknown. Using Double Helix 3D dSTORM and neurexin mouse models, we identify neurexin-3 as a critical presynaptic adhesion molecule that regulates excitatory synapse nano-organization in hippocampus. Furthermore, endogenous neurexin-1 and neurexin-3 form discrete and non-overlapping SSDs that are enriched opposite their postsynaptic ligands. Thus, the nanoscale organization of neurexin-1 and neurexin-3 may explain how individual neurexins signal in parallel to govern different synaptic properties.

Regulation of hippocampal mossy fiber-CA3 synapse function by a Bcl11b/C1ql2/Nrxn3(25b+) pathway

“…Neurexin mRNAs are subjected to extensive alternative splicing that leads to the expression of thousands of isoforms with differential expression patterns (Treutlein et al, 2014; Ullrich et al, 1995) that act in a type-specific manner on synaptic functions (Dai et al, 2019; Schreiner et al, 2014; Traunmuller et al, 2016). Nrxn3 splice variants have been shown to regulate the function and plasticity of glutamatergic and GABAergic synapses through various mechanisms (Aoto et al, 2013; Dai et al, 2019; Lloyd et al, 2023; Trotter et al, 2023). The Nrxn3 splice site SS5 is a major contributor to the high number of Nrxn3 isoforms (Schreiner et al, 2014).…”

“…Compared to controls, Bcl11b mutants injected with the control AAV displayed a strong reduction of LTP at 20-30 and 30-40 min after induction (Figure 3a-c; 0-10 min: Control+EGFP: 90. 4±7.2,Bcl11b cKO+EGFP: 106.1±10.8,[10][11][12][13][14][15][16][17][18][19][20]Bcl11b cKO+EGFP: 39.5±4.6,[20][21][22][23][24][25][26][27][28][29][30]Bcl11b cKO+EGFP: 24.8±3.2,[30][31][32][33][34][35][36][37][38][39][40]Bcl11b cKO+EGFP: 20.3±3.7,mean±SEM), consistent with our previous data (De Bruyckere et al, 2018). The loss of LTP was completely reversed upon the re-expression of C1ql2, with the rescue mice exhibiting comparable LTP to controls at all time intervals (Figure 3a-c; 0-10 min: Control+EGFP: 90.4±7.2, Bcl11b cKO+EGFP-2A-C1ql2: 86.…”

“…The loss of LTP was completely reversed upon the re-expression of C1ql2, with the rescue mice exhibiting comparable LTP to controls at all time intervals (Figure 3a-c; 0-10 min: Control+EGFP: 90.4±7.2, Bcl11b cKO+EGFP-2A-C1ql2: 86. 5±7.4,[10][11][12][13][14][15][16][17][18][19][20][20][21][22][23][24][25][26][27][28][29][30][30][31][32][33][34][35][36][37][38][39][40]mean±SEM). Importantly, this rescue effect was specific to C1ql2 as the overexpression of C1ql3 failed to reverse the Bcl11b cKO phenotype (Figure 3a-c MF-LTP is known to be mediated by the second messenger cAMP, which is produced by adenylyl cyclase (AC) in response to Ca 2+ influx through voltage-gated Ca 2+ channels (Li et al, 2007) and kainate receptors (KAR) (Lauri et al, 2001;Schmitz et al, 2003).…”

Neurexins control the strength and precise timing of glycinergic inhibition in the auditory brainstem

“…This finding is consistent with many studies in glutamatergic and GABAergic neurons, which converge to show that multiple genes, isoforms, and splicing sites of neurexins normally co-express in individual neurons and the expression profiles are highly cell-type specific and brain region specific (Fuccillo et al ., 2015; Schreiner et al, 2014; Treutlein et al, 2014; Uchigashima et al ., 2019). Distinct neurexins may interact separately and/or redundantly with diverse signaling molecules to form specific molecular codes for shaping neuronal connectivity and functional properties (Aoto et al, 2015; Aoto et al, 2013; Dai et al, 2019; Lin et al, 2023; Lloyd et al, 2023; Sudhof, 2017). The observed differential expression of individual neurexins suggests the possibility that specific isoforms may exert dominant regulatory roles in neurotransmission in different synapses.…”

Large donor CRISPR for whole-CDS replacement of cell adhesion molecule LRRTM2

“…The use of knockdown-rescue can reduce overexpression phenotypes 4,8 , but expression via exogenous promoters can nevertheless cause diverse effects that are difficult to adequately monitor. While some studies have utilized antibody detection of endogenous LRRTM2, the low signal-to-noise ratio and lack of possibility to manipulate the protein sequence limits their scope 10,11 . Knock-in mouse models could bridge this gap; however LRRTM2 has numerous binding partners, and making a separate mouse model for each mutation is expensive and inefficient.…”