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Fig. 4 | Genome Biology

Fig. 4

From: A rare codon-based translational program of cell proliferation

Fig. 4

Enhanced translation capacity in proliferating cells. a Normalized enrichment score (ES) for translation- and tRNA-related Gene Ontology terms, derived from gene set enrichment analysis (GSEA) comparing gene expression data from cells grown in 10% relative to 1% FCS. For all the pathways depicted, P < 0.05. P values were calculated by comparing the empirical ES of a gene set to a null distribution of ESs derived from permuting the gene set and then adjusted for multiple hypotheses testing. b Representative result of HPG incorporation measurement (a.u., arbitrary units) by flow cytometry in the wild-type NIH-3T3 cells grown in either 1% or 10% FCS (left). Quantification of the mean (± s.e.m.) ratio of HPG incorporation in 10% compared to 1% FCS for the same cells (n = 5) (right). For control, we also incubated wild-type NIH-3T3 cells with the translation inhibitor cycloheximide (CHX). Shown are the P values determined by the non-parametric Mann-Whitney U test. c Scatter plot of tRNA gene expression levels (log10, library size-normalized counts) in cells grown in either 10% or 1% FCS. Shown are the mean expression values for each tRNA gene (n = 3). The dashed line indicates equal relative abundances in the two conditions. No tRNA genes were significantly upregulated or downregulated between the two conditions (false discovery rate, FDR < 0.01). d Mean (± s.e.m.) mRNA and protein fold changes of tRNA deaminases for cells grown in 10% relative to 1% FCS (n = 3). e Mean (± s.e.m.) percentage of A34I modification in different tRNAs, as inferred from tRNA sequencing data, for cells grown in 10% relative to 1% FCS (n = 3). Shown are the P values determined by the non-parametric Mann-Whitney U test. The amino acid encoded by each codon is shown with a three-letter code: Arg, arginine; Ile, isoleucine; Leu, leucine; Pro, proline; Ser, serine; Thr, threonine; Val, valine. f Representative result of charged/uncharged tRNA-Lys-UUU abundances in cells grown in either 10% or 1% FCS using northern blot analysis. Total RNA was also treated with alkaline buffer to deaminoacylate tRNA-Lys-UUU. g Mean (± s.e.m.) ratio of charged to total tRNA-Lys-UUU in cells grown in either 10% or 1% FCS using northern blot analysis (n = 3). Shown is the P value determined by t test

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