Fig. 4.

HU induction of RSZ expression in various checkpoint mutants. (A) α-factor-arrested mec1Δ sml1Δ cells were released into YPD medium containing varying concentrations of HU at 30°C. Samples were collected at the indicated times and subjected to different analyses. (i) Status of ChrIII. (ii) Direct comparison between the distribution of HU induced DSBs in mec1Δ sml1Δ cells and that induced by thermal inactivation of Mec1. Roman numerals correspond to the six RSZs in ChrIII (Fig. 1B). (iii) The number of CFUs. Dotted lines denote the time at which 50% of the culture becomes commitment to inviability (T₅₀). (iv) FACS profiles. (B) Effects of HU on the indicated checkpoint mutants. (C) A model for dNTP regulation of RSZ expression. The extent of dNTP depletion regulates RSZ expression in mec1 or rad53K277A mutants. The overall dNTP levels in each strain are determined by the status of MEC1, SML1 and the concentrations of HU. When dNTP concentrations fall below the viable threshold (asterisks), mec1 or rad53 cells lose viability. If the extent of dNTP depletion is modest (the area marked by the grey box), RSZ expression is observed (grey circles). When the dNTP concentration falls below this level, the cells become prematurely committed to death before RSZ expression (black circles).