Fig. 3: Aged neurons have chronic stress granules that repress translation.
From: Neuronal aging causes mislocalization of splicing proteins and unchecked cellular stress
a, Volcano plots of G3BP1 pulldown and MS of all detected proteins in transdifferentiated neurons versus unstressed (top) or stressed (bottom) iPSC-derived neurons (n = 2 replicates). Detected proteins included in the following KEGG terms were highlighted: spliceosome (dark blue, hsa03040), stress granules (orange, hsa03019 + 10146), HSP90 chaperones (light blue, no KEGG pathway) and RNA transport (green, hsa03013). P values were calculated using a two-tailed Welch’s t test. b, Confocal fluorescence images of stress granule markers in transdifferentiated and iPSC-derived (iPSC-diff) neurons. The yellow boxes denote the inset region, and the yellow dashed lanes indicate the intensity profiles that were normalized and plotted in the last column. White scale bar = 10 μm; yellow scale bar = 2 μm. c, Top and middle, western blots of total and phosphorylated eIF2α for the indicated samples. Bottom, quantification of average phospho-eIF2α-to-total eIF2α expression for n = 3 blots, where the centerline denotes median and the error bands denote the range of data values; statistics were calculated using a two-tailed Welch’s t test comparing the indicated samples to the unstressed iPSC-derived neurons. d, Fraction of G3BP1-bound RNA windows detected by eCLIP (n = 2 replicates) within each of the indicated transcript elements for transdifferentiated and iPSC-derived neurons. e, Same as d, but for Caprin1-bound RNAs. f, Fraction of G3BP1-bound RNA windows detected in unstressed iPSC-derived neurons and/or transdifferentiated neurons. g, Same as f, but for Caprin1-bound RNAs. h, Volcano plot of translation efficiency of all detected transcripts using Ribo-seq in transdifferentiated and iPSC-derived neurons. CDS, coding sequence; TE, translation efficiency.
