Macromolecular syntheses in the cell cycle mutant cdc25 of budding yeast

Abstract

A major control point of the cell cycle in Saccharomyces cerevisiae is a G1 event called 'start'. At start a yeast cell integrates external and internal signals and decides to progress toward mitosis or to choose alternative pathways such as sporulation, conjugation etc.cdc25 is a class I1 temperature-sensitive start mutant that blocks at restrictive temperature in GI as round unbudded cells.The arrest of the cell cycle appears to be independent of the carbon and nitrogen sources, and the cell wall of cdc25-arrested cells shows changes similar to those found in cells undergoing entry in to the stationary phase.After a shift to 36 "C the increase in cell number of cdc25 cultures is gradually inhibited. The nuclear division cycle appears to be inhibited immediately after the shift and the percentage of budded cells decreases, while cytoplasmic growth, monitored either as increase of adsorbance at 450 nm or as protein accumulation, continues for many hours leading to a progressive increase of mean cell volume and mean protein content per cell.The stable RNA accumulation instead is immediately inhibited and this is partially due to a 50 inhibition of ribosomal RNA synthesis, while the rate of synthesis of ds-killer RNA is relatively unaffected.These data suggest that the CDC25 gene product could be a part of a mechanism that leads yeast cells to choose between the progression towards DNA replication and cell division or to enter into the stationary phase. This mechanism appears to turn off both rRNA accumulation and cell-cycle progression and to activate differentiative pathways in response to environmental restriction.The molecular basis of the coordination between growth and the nuclear division cycle is far from being understood. For budding yeast Saccharomyces cerevisiae a major control point has been found in G1 and it has been called 'start' [1,2]. At start, a yeast cell integrates much biochemical and environmental information and decides, depending on its genetic background, on a number of alternative pathways : sporulation, conjugation, entrance in stationary phase, or progress towards mitosis [I -41. The traverse of start is required for the beginning of the nuclear division cycle, and in growing cells, a few minutes after the completion of this event, both initiation of DNA synthesis and bud emergence begin [5]. However, many recent papers [3,4,6] have stressed that start is more an area than a point, since different environmental factors block yeast cells at start in physiologically distinct states. Accordingly, the start cdc mutants, i.e. temperature-sensitive mutants that block at a restrictive temperature at or near start, [7] have been divided into class I and class 11 mutants according to their terminal phenotype [6].Class I mutants continue to grow, while bud emergence and DNA replication are blocked, thus resembling alphafactor-treated cells [6]. Moreover, they are able to conjugate at restrictive temperature but not to sporulate if diploid. Class I1 mutants do not grow at restrictive temperature ...

Cell size modulation by CDC25 and RAS2 genes in Saccharomyces cerevisiae.

“…The NA1-D2 Ts strain underwent an arrest with biphasic kinetics in the unbudded Gl phase of the cycle ( Fig. 2A to C) as previously shown for this start mutant by using a synthetic medium (29,30). On the other hand, the VNA1-D2 strain gave a response to the temperature shift which was identical to that of the wild type, demonstrating that the RAS2v"'-l9 mutation can efficiently suppress the impaired CDC25 function in the Ts mutants.…”

“…When this system is genetically altered by ((1(25-1 and RAS2V`l9 mutations, two main effects are observed under very different conditions: a constitutive repression or derepression of cell growth and metabolism that is no longer dependent on the actual nutritional conditions of cells (30,46) and an altered modulation of critical cell size during the exponential phase of growth, as demonstrated in this paper. Both phenotypes are probably caused by altered nutritional information given to cells by the CDC25-RAS2-modified system.…”

“…resemble a-factor-treated cells at the restrictive temperature and continue cytoplasmic growth. In class II start mutants (cdc25, (d(33, cdc35), a growth lesion is also apparent; these mutants arrest the cell cycle, mimicking stationary-phase cells (15,30).…”

“…In fact, all the mutations that inactivate this mechanism, thereby reducing the ability to increase intracellular cAMP levels ((dc25, (dc135, rasl. ras2), arrest growth in a manner that mimics nutrient starvation (17,22,30). If the inactivation is only partial, as in some ras2 mutants, it causes a so-called repressed phenotype characterized by high heat shock resistance, hyperaccumulation of glycogen, and inability to grow on nonfermentable carbon sources (44).…”

Genetic assessment of stationary phase for cells of the yeast Saccharomyces cerevisiae

“…Mutant phenotypes reveal the role of cAMPmediated signal transduction in nutrient sensing (21,43). Conditional mutations in either CDC25 or CYR] that decrease the activity of the cAMP-mediated pathway (6) bring about regulated arrest of cell proliferation, presumably because of specious signaling of nutrient status (17,24,25). Furthermore, cells blocked in proliferation by these cdc25 or cyri mutations exhibit properties of stationary-phase cells, even in nutrient-replete medium (17,24,25).…”