Fig. 1

4EHP expression in the brain. a–c Developmental expression of 4EHP, GIGYF2, and GAPDH in the cortex, hippocampus, and cerebellum, respectively, as measured by western blot. Quantification of a, b, and c (lower panel, n = 3 per group, normalized to the average of all age points for each protein per membrane). d 4EHP expression in a synaptosome preparation (left panel). PSD95 was enriched in the synaptosome (Syn) as opposed to the cytosol (Cyto), demonstrating proper synaptosome preparation. GAPDH and β-actin were used as loading controls. Quantification of d (right panel, n = 3). e Primary neurons were derived from the hippocampus of wild-type mice and cultured for 14 d. Immunofluorescent analysis confirmed 4EHP expression in the synapse by colocalization with PSD95 (merge). The scale bar represents 20 µm in the upper panel of images and 5 µm in the lower panel of images. The lower panel of images correspond to 4 × zoom of the upper panel of images defined by the white box. Analysis of cell-type-specific expression of 4EHP by colocalization with f Empty Spiracles Homeobox 1 (EMX1, defining excitatory neurons), g parvalbumin (PV, defining a subset of inhibitory neurons), h somatostatin (SST, defining another subset of inhibitory neurons), and I laminin (LAMA1, defining endothelial cells) in the hippocampus of wild-type mice. 4EHP expression is colored in red, the cell type marker in green, and Hoechst-stained nucleus in blue. Arrows indicate a positive signal for the cell type maker. Scale bar represents 20 µm