XUT levels remain unaffected upon translation initiation inhibition. (A) Working model showing the presumed effect of CHX-mediated inhibition of translation elongation (left) and of stress-induced inhibition of translation initiation (glucose starvation, right) on Xrn1-dependent degradation of XUTs (red). The red arrow on the XUT represents a smORF. (B) Total RNA-seq was performed in WT (YAM1), xrn1Δ (YAM6), and upf1Δ (YAM202) grown in complete synthetic medium (CSM). WT cells grown in the same conditions and then submitted to a CHX treatment or glucose starvation (−Glu) were also included. Densities (tag/nt) were computed for the 1470 XUTs significantly up-regulated in the xrn1 mutant grown in CSM (see Supplemental Fig. S3A), which were then separated according to their sensitivity to NMD (see Supplemental Fig. S3B). The sensitivity of each of these XUTs to CHX and glucose starvation is presented as a heatmap of the FC (log2 scale). As an indication, the sensitivity of these XUTs to Xrn1 (xrn1Δ/WT) and NMD (upf1Δ/WT) is also presented. (C) Box-plot showing the RNA-seq signals (densities, tag/nt, log2 scale) for the same set of XUTs as in B, in WT (YAM1), upf1Δ (YAM202), and xrn1Δ (YAM6) cells grown in CSM with glucose (control; black) or undergoing glucose starvation for 16 min (−Glucose; gray). An aliquot of WT cells was then treated with CHX for 15 min. (***) P-value < 0.001; (ns) not significant upon two-sided Wilcoxon rank-sum test (adjusted for multiple testing with the Benjamini–Hochberg procedure).
