Figure 8.

Development of neuroligin-2-based SynView2. A, Schematic drawing of the NL2-GFP11^S2 construct used for SynView2. Numbers represent amino acid residues of rat neuroligin-2 (accession no. Q62888). SP, signal peptide; EHD, Esterase homology domain; TM, transmembrane region; and CT, cytoplasmic tail. B, HEK293 cells expressing Nrx1β-GFP1–10^S2 were mixed with cells expressing NL2-GFP11^S2 and development of the reconstituted GFP signal at the border of cell membrane contacts was monitored by fluorescence microscopy. Scale bar, 10 μm. C, Image of a hippocampal neuron sparsely transfected with NL2-GFP11^S2 surrounded by neurons expressing lentivirally transduced Nrx1β-GFP1–10^S2. Punctate SynView2 GFP fluorescence (green) was combined with immunofluorescence staining to NL2-GFP11^S2 (using an anti-HA antibody; red) and synapsin (blue). D, High-magnification images of the boxed dendritic area in C illustrate individual and composite images of the Synview2 signal and NL2-GFP11^S2 and synapsin-positive puncta. Scale bar, 10 μm. E, F, Quantitative analysis of the fraction of SynView2-positive (E) and NL1-GFP11^S2-positive puncta (F) that are adjacent to synapsin-positive puncta. Data represent means ± SEM; n = 3 independent experiments. Data show representative experiments independently repeated at least 3 times.