Fig. 6: Multi-layered proteomics and ultrastructural analyses reveal critical roles of CDH11 in synaptic vesicle recycling.
(A) Gene Ontology (GO) analysis for Biological Process (BP) and Cellular Component (CC) of downregulated proteins in the ACC of EcKO vs. WT mice. GO terms with a false discovery rate (FDR) of < 0.05. (B) Heatmap of differentially expressed proteins in the ACC of EcKO vs. WT mice. (C and D) Western blots (C) and relative expression levels (D) of CDH11, VTI1A, and MAGI2 in ACC (n = 6 WT/6 EcKO for CDH11 and VTI1A; n = 7 WT/7 EcKO for MAGI2). (E) GO analysis for BP and CC of downregulated proteins in cortical synaptosomes of EcKO vs. WT mice. GO terms with an FDR < 0.05. (F) Heatmap of differentially expressed proteins in cortical synaptosomes of EcKO vs. WT mice. (G and H) Western blots (G) and the relative expression levels (H) of CDH11, UNC13A, SYT1, STX1B (n = 7 mice/group) in cortical synaptosome preparations. (I and J) GO analysis (I) and volcano plot (J) of proteins immunoprecipitated by CDH11 from P7 cortical tissue lysates. (K) Schematic illustration of CDH11 co-immunoprecipitation. (L) Western blots of co-immunoprecipitated STX17 with CDH11. See also Figures S6-S8. (M and N) Representative images of excitatory (M) and inhibitory (N) synapses captured by transmission electron microscopy. (O–Q and R–T) Quantification of excitatory (O–Q) and inhibitory (R–T) synaptic ultrastructure: number of presynaptic vesicles (O and R), synaptic cleft width (P and S), and distance of synaptic vesicles to the presynaptic membrane (Q and T) (n = 3 mice/group). Data are presented as mean ± S.E.M, with individual values represented as dots. A P value of ≤ 0.05, determined by either a one-tailed ($P) or two-tailed Student’s t-test, or a one-tailed (n$P) or two-tailed (nP) nonparametric Mann-Whitney test, as appropriate, was considered significant.
