Project description:DDI2 and DDI3 (DDI2/3) are duplicated genes in Saccharomyces cerevisiae that exhibit strong induction by a transcription factor Fzf1 in response to chemical treatments like cyanamide (CY) and methyl methanesulfonate (MMS). Although, like DDI2/3, SSU1, YHB1 and YNR064C also contain an Fzf1-binding consensus sequence CS2 and are coordinately regulated by Fzf1, these genes are only modestly induced by CY and MMS. To identify additional cis-acting elements in the DDI2/3 promoter, we made DDI2/3 promoter deletions in a reporter system and identified upstream repressing sequences (URS) spanning 480 nucleotides. To test a hypothesis that the chromatin structure constitutes the URS, we utilized a yeast strain capable of histone H3/H4 depletion by shifting carbon sources. Following histone depletion, DDI2/3 were strongly induced in an Fzf1 dependent manner, while YHB1 was repressed. Interestingly, under histone depletion conditions, CY or MMS treatment further increased expression of all Fzf1-regulated genes to comparable levels in an Fzf1 dependent manner. A genome-wide MNase-seq analysis showed that CY treatment reduced the nucleosome occupancy at the mapped DDI2/3 URS region in wild-type cells, but not in in fzf1Δ cells. These findings collectively indicate that Fzf1 plays dual roles in regulating the DDI2/3 response to CY. Firstly, it binds CS2 and serves as a transcription activator. Secondly, it is required for the chromatin remodeling at URS. This two-tier regulation at the DDI2/3 promoter helps to explain why DDI2/3 achieve much higher fold induction by CY and MMS than other Fzf1-regulated genes, suggesting Fzf1 to be a candidate pioneer transcription factor.
Project description:The cellular response to treatment with DNA-damaging substances at low concentrations which are genotoxic but do not have a strong cytotoxic effect are of special interest. In addition, environmental variations that influence growth conditions, e.g. different media, and individual fitness, e.g. different strains, are likely to influence and modulate the adverse effects of individual DNA damaging substances. At sub-cytotoxic levels, DNA damaging substances play an important role in the accumulation of genomic mutations. In longer living organisms, like humans and other mammals, exposure to DNA damaging substances over extended period of time is a critical factor that contributes to the development of various diseases and in particular of tumors. The aim of our work was to study how strain background and growth conditions influence respond to DNA damage caused by low doses of MMS and which part of these changes is responsible for their sensitivity to toxic conditions. We analyzed sensitivity of two yeast strains FF18984 and BY4742 to MMS in media with limited and full nutrient availability. Keywords: Yeast, S.cerevisiae, MMS, stress response, DNA damage
Project description:The N-terminal tail of histone H2A shows evolutionary changes that parallel genome size and aid chromatin compaction. As genome size increases, so does the number of arginines. In contrast, serines corellate with small genomes. Examples for such changes are arginine in position 11 and serine in position 15. To test if these residues affect mRNA levels, we analysed gene expression profiles of S.cerevisiae strains containing either WT or mutant H2A.