Fig. 2: Discerning functionally relevant clues from the phosphoproteome.
From: Insulin signaling regulates longevity through protein phosphorylation in Caenorhabditis elegans
a Schematic diagram illustrating the design of iFPS, which employs multinomial logistic regression from machine learning to integrate six features and predict the functionally impactful phosphosites in C. elegans. iFPS inference of functional phosphosites, UKF upstream kinase family, PhC phosphorylation conservation, RSA relative surface accessibility, ASC acetylation site co-occurrence, SS secondary structure, IDM interacting domain and/or motif, RCR residue conservation ratio, CMGC/AGC/CDK/CAMK kinase groups, HMMER software that predicts functional domain, 3did database of three-dimensional interacting domains, Phos phosphorylation, Ac acetylation. b Prediction power (AUC, area under the curve) of different models. One hundred and one known functional phosphosites and 605 randomly phosphosites from dbPAF served as the training data set. Tenfold cross-validations were performed. The iFPS model, which integrated six features, was superior to models constructed from individual feature. c Phosphorylation changes in the long-lived daf-2 mutant compared to the WT control (see “Methods”). Phosphoisoforms quantified at least three times in both daf-2 and WT control were subjected to statistical comparison. The log2[median of (14N/15N)daf-2)/median of (14N/15N)control] values and the log10(p value, Wilcoxon rank-sum test) of phosphoisoforms were plotted on the x axis and y axis, respectively. Dots (n = 2365 phosphoisoforms) met the criteria of [log2(fold change) beyond 1.5× SD and one-tailed p value < 0.05] or [log2(fold change) beyond 1.0× SD and two-tailed p value < 0.05] are colored in red (hyper-phosphorylated) or green (hypo-phosphorylated). AKT-1 pT492, EIF-2α pS49, and CDK-1 pT179 are functionally validated in this study. See statistics in Source data. d Prioritizing the daf-2-regulated phosphosites by iFPS ranking. The daf-2-regulated phosphosites that ranked among the top 5% highest scoring iFPS phosphosites are shown. Proteins were grouped by KEGG ontology or function annotations recorded in WormBase release WS275. Royal blue marks the lifespan-regulating proteins. Red colors the upregulated phosphoisoforms. Green colors the downregulated phosphoisoforms. The daf-2/WT values are the relative phosphorylation levels of phosphoisoforms in daf-2 compared to WT.
