Fig. 5: Time-resolved SICFA reveals the sequential effects of 5-FU on cellular biochemical pathways.

a Schematic representation of the metabolic pathways of 5-FU and their impact on DNA and RNA damage. Proteins with significant changes in stability and abundance are highlighted in red. b Experimental workflow of the protein stability and protein abundance analysis. K562 cells were treated with 5-FU or DMSO at different time points (30 min, 4 h, 12 h, and 24 h), followed by cell lysis, protein extraction, digestion, and LC-MS/MS analysis. c Volcano plots showing changes in protein stability (SICFA) after 5-FU treatment in K562 cells at different time points (30 min, 4 h, 12 h, 24 h). The p values from two-sided empirical Bayes t tests without adjustment, with n = 4 cell replicates. Known targets are marked in red, and potential targets are shown in green. The dashed line corresponds to a significance threshold of −log₁₀(p value) = 4. Source data are provided as a Source Data file. d Venn diagram showing overlap between proteins with significant changes in stability (SICFA) and those with altered abundance. e Line chart showing the number of proteins with significant changes in stability and abundance over time. Cellular responses are categorized into four stages: (i) molecular initiating events, (ii) early events, (iii) intermediate events, and (iv) outcome events. Source data are provided as a Source Data file. f Heatmaps showing bioinformatics enrichment analysis of biological processes associated with proteins with significant changes in stability (SIC) and abundance (Abu) at different time points, highlighting the time-resolved effects of 5-FU on cellular biochemical pathways. Colors indicate p-values (one-sided Fisher’s exact test, no adjustment), with white cells indicating a lack of enrichment of the term in the corresponding gene list.