Fig. 5. NPC association alleviates R-loop toxicity.
a Serial dilutions of the indicated strains were grown at the indicated temperatures (25 °C, 30 °C, 37 °C) on rich medium (YPD). b Principle of the tethering assay. The intronless version of the YAT1 transgene, under the control of the GAL1-10 promoter, is flanked by direct leu2 repeats to allow quantification of R-loop-dependent recombination events. The reporter is integrated at the chromosome II GAL locus, which also contains an array of Lac operator (LacO) repeats for microscopy visualization and LexA-binding sites. Expression of the LexA-Nup60 fusion ensures the permanent tethering of the locus to NPCs. c Recombination frequencies were calculated for the indicated strains as described in Methods (fraction of Leu+ prototrophs, ×10−4; n = 31 for wt LexA, n = 32 for tho LexA, n = 33 for wt LexA-Nup60, n = 35 for tho LexA-Nup60; n refers to biologically independent cultures). Glucose treatment was achieved for 1 h following 6 h growth in glycerol-lactate medium. Box-plots are defined as above (Fig. 1d–f). Statistical test: two-sided Mann-Whitney-Wilcoxon test; (a), p = 2.57 × 10−2; (b), p = 1.2 × 10−3; (c), p = 1.6 × 10−3. Source data are provided as a Source Data file.