Abstract
This laboratory study aimed to investigate the effect of α-factor on cell division processes in Saccharomyces cerevisiae. The investigation involved three strains of baker’s yeast: the wild type and the mutant variants Far1 and Ste2. In the presence of the α-factor, signal transduction activation was expected to increase the number of diploid cells in the WT sample and decrease in the mutants.
The results confirmed the hypotheses for WT and Ste2, but were not unequivocal for Far1, which increased the number of unbudded cells when stimulated but reduced the number of NOT shmooing cells. The findings suggest the feasibility of α-stimulating signal transduction for wild-type yeast and also confirm the efficacy of the Ste2 mutation in reducing the proportion of sexually reproducing cells.
Introduction
Signal transduction is the ability of cells of the same or different types to exchange information with each other and regulate processes. The information in this cellular exchange is biochemical reactions that occur on a receptor/molecule type basis, allowing cells to understand their environment and how they should respond¹. One of the features of baker’s yeast (S. cerevisiae) is the ability to change cell division patterns depending on signal transduction. Thus, in the absence of α-yeast, a yeast divides asexually, characterized as budding; however, in the presence of the second type of yeast, a yeast enters the signal transduction stage and divides sexually.
Transduction occurs through the production of molecules on the α-side of the yeast (α-factor), which are taken up by the a-yeast receptors, but the opposite is also true. The binding of α-factor to the receptors leads to the activation of a kinase complex that activates the protein kinase Fus3, which phosphorylates the proteins Far1 (cell stopping in the G1phase of the cell cycle) and Ste2 (activation of genes involved in mating)2. These changes prevent the budding process, allowing two yeast cells of different types to form a diploid zygote.
The effect of α-factor presence on the fission patterns of baker’s yeast, both wild-type and mutant forms, was investigated. Samples of S. cerevisiae wild-type (WT), with a mutation in the Far1 and the Ste2 genes, were either exposed to α-factor to study sexual fission abilities or used as a control group.
Appropriate hypotheses were formulated for each of the yeast variants used. First, WT stimulation would increase the proportion of diploid zygotes and reduce the number of budding cells by stimulating signal transduction. Second, a mutation in Far1 in the presence of the α-factor should have reduced the proportion of shmooing yeast because it prevented activation of the budding arrest factor. Similarly, a mutation in the Ste2 gene should have led to a decrease in the proportion of shmooing yeast because fewer yeast would have been able to mate.
Materials and Methods
For each of the three samples (WT, Far1, and Ste2), microscopic examinations were performed at 40X magnification. 20 mL of yeast from stimulated or control conditions were carefully transferred to a slide, and then the numbers of budding and shmooing cells were counted using a light microscope.
Results
Figure 1 shows the results of the visual representation of the slides studied with the microscope. Table 1 presents information on cell counts in the budding state, yeast shmooing, and the proportion of cells that did not undergo budding. As shown, the proportion of unbudded cells increased for WT under α-stimulation conditions, increased for Far1 under the same conditions, but decreased for Ste2. In contrast, the number of budding cells was zero in the WT-stimulated condition, decreased in Far1+, and increased in Ste2+. The number of non-shmooing yeast cells, as shown in Fig. 2, increased only for WT+ and decreased for Far1+ and Ste2+. However, shmooing cells were detected only in WT+ and Far1+ strains.
Table 1. Results of counts of the number of cells in different states of division and the proportion not budding.
Discussion
The present study focused on how α-factor affects the cell division process of Saccharomyces cerevisiae in three types of baker’s yeast. It was hypothesized that stimulation of signal transduction would increase the proportion of diploid cells for WT but decrease it for mutant forms (Far1 and Ste2) because the steps of sexual division regulation must be disrupted. Signal transduction under α-stimulation was successful for wild-type yeast, as indicated by an increased proportion of shmooing compared to the control line.
For the Far1-stimulated mutant, the proportion of unbudded cells also grew, which rejected the hypothesis and indicated that the cell cycle arrest mechanism persisted: likely, the mutation produced was ineffective. Nevertheless, compared to WT, the increase in this fraction was not so large, indicating a weakening of activation by the mutation. This is consistent with the fact that the number of non-shmooing yeast was reduced for Far1+ compared to the control line, in contrast to WT+.
Notably, for the Ste2 mutant, the proportion of unbudded cells decreased, indicating the successful inhibition of mating gene expression and an increase in budding processes. In other words, for WT, it was confirmed that the presence of α-factor induces the process of sexual reproduction in Saccharomyces cerevisiae. For the Ste2 mutant, inactivation of mating genes was observed, whereas for the Far1 mutant, the effect was less unambiguous.
The experiment had several limitations, some of which may be addressed in future research. In particular, the number of repeated trials was minimized, which could have led to distorted counts and inaccuracies. Additionally, cell counts were not standardized and were subject to the subjectivity of measurement for different yeast types. Addressing these limitations, combined with studying other forms of mutation in yeast and the influence of external factors (such as temperature and pH), could be an extension of the current study.
Reference List
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