Fig. 4: Fuzzy clustering reveals translation dynamics as a potential driver of neuron aging.
a, Fuzzy clustering on z-score-normalized gene expression values (y axis) over the predicted aging course of the 128 CeNGEN neurons identified four clusters. The 128 neurons were merged into five age-prediction bins: (1) 97–110 h, (2) 110–120 h, (3) 120–130 h, (4) 130–140 h and (5) 140–180 h (x axis). The number of genes within each cluster are annotated. Dashed black lines show the central trend. b, Pathway overrepresentation analysis of the four clusters shows age-related pathways. The x axis shows pathway terms, and the y axis indicates clusters. Circle size reflects the enrichment score, defined as the log2 ratio of observed-to-expected genes per pathway (log2 enrichment ratio). Circle color represents statistical significance as –log10(false discovery rate (FDR)). Only representative nonredundant pathways are shown for visualization (Methods). c, Fraction plots displaying the fraction of nematodes at day 1 of adulthood expressing GFP in different neurons categorized as ‘healthy’, ‘mildly damaged’ and ‘severely damaged’ that were treated with 2 mM CHX for 24 h. Three to four cohorts were analyzed, consisting of 10–30 individual nematodes. Data shown are the mean ± s.d. Ordinal regression (CLM) was used to test for significant differences. d, Representative fluorescence microscopy images (z-stack maximum projections) of ASE neurons in 1-day-old adult nematodes of strain NY2067 after 24-h treatment with 2 mM CHX or dimethylsulfoxide (DMSO) control. Gray arrows indicate morphologic aberrations along the neurites. Scale bar, 20 µm. e, Fraction plots displaying the fraction of nematodes expressing GFP in ASE neurons categorized as ‘healthy’, ‘mildly damaged’ and ‘severely damaged’ that were treated with 2 mM CHX for 24 h at day 1 of adulthood. Four cohorts were analyzed, consisting of 10–30 individual nematodes. Data shown are the mean ± s.d. Ordinal regression (CLM) was used to test for significant differences. f, Swarm plot showing the chemotaxis index of nematodes conditioned at 0 mM (light blue) or 20 mM (dark blue) NaCl. The ASE neuronal marker strain (NY2067, mediating salt aversive conditioning) was incubated with 2 mM CHX or DMSO control for 24 h at day 1 of adulthood. Data shown are the mean ± s.d. from eight independent cohorts of 20–100 nematodes. A one-tailed t-test was used to test for significant differences.
