Figure 1. Dynamic measurements of protein synthesis with a translocation reporter.

(a) A FP is fused to a SynZip (SZ2), expressed under the control of a constitutive promoter and can freely diffuse between the cytoplasm and the nucleus (dPSTR OFF, top). The induction of the promoter of interest drives the expression of the second peptide of the reporter, composed of two NLSs fused to a compatible SynZip (SZ1). The strong interaction between the SynZip peptides leads to the enrichment of the FP in the nucleus (dPSTR ON, bottom). (b) Microscopy images of cells with histone Hta2 tagged with CFP and carrying the pSTL1-dPSTR^R submitted to a 0.2 M NaCl stress. The inducible peptide is fused to a Venus FP. Scale bar, 5 μm. (c,d) Quantifications of the nuclear enrichment in the dPSTR^R (c) and the Venus (d) channels for cells stressed with 0 (orange, N_C=285), 0.1 (cyan, N_C=266), 0.2 (blue, N_C=294) or 0.4 M NaCl (red, N_C=265). Nuclear enrichment is measured as the difference between nuclear and cytoplasmic fluorescence for each single cell. For all similar graphs throughout the paper, the solid lines represent the population average and the error bars are the s.e.m. N_C represents the number of single cells measured. (e) Histograms of the time needed for each single cell to reach half of its expression output for either the dPSTR^R (solid lines) or the Venus (dashed lines). The expression output represents the maximal amplitude of the nuclear enrichment (see Methods). (f) Single cell correlation of the expression output measured by either the pSTL1-dPSTR^R or the pSTL1-Venus assay, for control cells (orange) or cells induced with 0.2 M NaCl (blue). The dashed lines represent the expression thresholds, above which cells are considered as expressing. All the figures of the paper represent one representative experiment of at least three biological replicates.