Figure 7

Menin knockdown reduces the presynaptic clustering of α7 subunit-containing nAChRs. (A,B) Super resolution images of C-menin, α-BTX, and SYT puncta in GFP+ synaptic ROIs from NTC shRNA (A) and MEN1 shRNA (B) lentivirus transduced hippocampal cultures at DIV 7 (n ≥ 10 images, 2 independent samples each, representative images). Cells were labeled with C-terminal menin and SYT antibodies (i–v; NTC: n = 16, MEN1: n = 14), fluorophore-tagged α-BTX and a SYT antibody (vi–x; NTC: n = 16, MEN1: n = 15), or α-BTX and a C-terminal menin antibody (xi–xv; NTC: n = 11, MEN1: n = 10). (i,vi,xi) shows GFP fluorescence, (ii,xiii) shows α-C-terminal menin fluorescence, (vii,xii) shows α-BTX fluorescence, (iii,viii) shows α-SYT fluorescence, (iv,ix,xiv) shows SYT/C-menin/α-BTX merged channels, (v,x,xv) shows channels merged with GFP. Scale bar, 2 μm. (C) Summary data, mean number of α-C-terminal menin, α-BTX, and α-SYT puncta in super resolution images. (D) Summary data, incidence of colocalization of SYT/C-menin/α-BTX puncta in super resolution images. Asterisks, statistical significance (independent t-test or Mann-Whitney U test); *P < 0.05. **P < 0.01. ***P < 0.001. See also Table S6.