Fig. 1.

Global proteomicsof theaging mice brain. A, experimental workflow. Brains from adult, mid-age and old mice were harvested and homogenized as described in methods. Proteins were then digested using four different enzymes (AspN, Chymotrypsin, GluC and Trypsin) in parallel, and phosphopeptides were enriched prior to LC-MS/MS. The raw LC–MS/MS data was then processed using the MaxQuant (www.maxquant.org) and Perseus (www.maxquant.org/perseus) platforms. B, Violin plots depicting the overall distribution of Log₂(LFQ) values in the global dataset. Yellow hue corresponds to adult mice, cyan mid-age and red corresponds to old mice. C, PCA plot of all brain samples analyzed in the global proteomics. Age groups of the mice are indicated by the same hues as described in (B). D, Volcano plot analysis of all quantified proteins. Significantly regulated (q < 0.05 & logFC >|0.5|) proteins are color coded, and top regulated proteins are labeled. E, Heatmap representation of all significantly changing (q < 0.05) proteins in our analysis. Hue represents the Z-score normalized Log2(LFQ) values. Red hue indicates upregulated proteins and blue represents downregulated proteins. Proteins were clustered into 5 clusters by k-means. F, Z-scored Log₂(LFQ) values from proteins from each k-mean cluster, across replicate mice from adult, mid-age, and old groups. G, top four Reactome Gene Set terms enriched for the significantly changing proteins found each k-means cluster represented in Figure 1, E and F.