Fig. 4: Identifying the protein targets bridging autophagy to aberrant spines. | Molecular Psychiatry

Fig. 4: Identifying the protein targets bridging autophagy to aberrant spines.

From: Autophagy controls the hippocampal postsynaptic organization and affects cognition in a mouse model of Fragile X syndrome

Fig. 4

Hippocampus were isolated from Fmr1 KO vs. WT mice with different treatments and protein lysis was subjected to proteomics. A Volcano plot for proteins significantly (p < 0.05) up- (red) or down-regulated (blue) in Fmr1 KO vs. WT mice. B Volcano plot for proteins significantly (p < 0.05) up- (red) or down-regulated (blue) in Fmr1 KO mice injected with Rilmenidine (Ril) vs. saline (Veh). C Volcano plot for proteins significantly (p < 0.05) up- (red) or down-regulated (blue) in WT mice injected with CQ (autophagy inhibitor) vs. Veh. D Venn diagram showing overlapped 42 proteins. Protein names are labeled in A, B, and C. Overlap between proteins increased in Fmr1 KO vs. WT and proteins increased in WT by CQ vs. Veh: p < 10−16. Overlap between proteins decreased by Rilmenidine in Fmr1 KO and proteins increased in WT by CQ vs. Veh: p = 1.4 × 10−9. Overlap between proteins increased in Fmr1 KO vs. WT and proteins decreased by Rilmenidine in Fmr1 KO: p < 10−16. E GO biological processes analysis (Mus musculus database of brain-expressed genes as the background, Dataset S8) of 42 overlapped proteins showing top nervous system-related processes (p < 0.05). F SFARI Gene database shows 7 of these 42 overlapped proteins are associated with reported autism cases. G, H Primary neurons were cultured from the hippocampus of WT and Fmr1 KO mice and treated with labeled drugs: WT (WT neurons with DMSO); Fmr1 KO (Fmr1 KO neurons with DMSO); Fmr1 KO + Ril (Rilmenidine, 10 µM for 6 hr) and WT + lyso Inhi (lysosomal inhibitors: 10 mM NH4Cl with 50 μM leupeptin for 6 hr). Lysates were extracted and immunoprecipitated with an antibody to ubiquitin. Whole cell lysates (Input) and immunoprecipitants (IP) were immunoblotted (IB) for eIF4G1 and PSD-95. I, J Summary data for ubiquitinated eIF4G1 and PSD-95 in precipitates (normalized to WT + DMSO). n = 4 mice (5-week-old) in each group of A-C. Significance were calculated by ANOVA followed by a Tukey’s test. *p < 0.05, **p < 0.01. β-actin was used as a loading control. Each circle in I and J represents data from an independent culture (n = 4).

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