Phenotypic analysis of gene deletant strains for sensitivity to oxidative stress

Abstract

Ascertaining the impact of inhibitors on the growth phenotype of yeast mutants can be useful in elucidating the function of genes within the cell. Microtitre plates and robotics have been used to screen over 600 deletions from EUROSCARF, constructed in an FY1679 strain background, for sensitivity to various oxidants. These included the inorganic hydroperoxide, H 2 O 2 , an organic peroxide (cumene hydroperoxide) and a lipid hydroperoxide (linoleic acid hydroperoxide). These produce within the cell several different reactive oxygen species that can cause damage to DNA, proteins and lipids. Approximately 14% of deletants displayed sensitivity to at least one of the oxidants and there was also a distribution of deletants that showed sensitivity to all or different combinations of the oxidants. Deletants included genes encoding proteins involved in stress responses, heavy metal homeostasis and putative cell wall proteins. Although global mechanisms have been identified that provide general stress responses, these results imply that there are also distinct mechanisms involved in the protection of the cell against specific damage caused by different oxidants. Further analysis of these genes may reveal unknown mechanisms protecting the cell against reactive oxygen species.

Decrease of H2O2 Plasma Membrane Permeability during Adaptation to H2O2 in Saccharomyces cerevisiae

“…Therefore, the cell wall does not limit H 2 O 2 diffusion into Sc cells, as could be expected because the cell wall has pores that are permeable to low molecular weight molecules (26). Our observation is also consistent with the fact that Sc cell susceptibility to H 2 O 2 is not altered by deleting genes ECM25, ECM33, and YOR275c, which are involved in cell wall integrity (27).…”

The Rho1 GTPase Acts Together With a Vacuolar Glutathione S-Conjugate Transporter to Protect Yeast Cells From Oxidative Stress

“…Gray, University of Glasgow, Glasgow, UK) were tested similarly, except plated on the YPD plates. The laboratory wild-type strains exhibited varying degrees of sensitivity to H 2 O 2 depending on the background: BY4741 was more sensitive to H 2 O 2 compared with other wild-type strains tested ( Figure 9, B and C and Figure S1), as previously reported (Higgins et al 2002;Singh et al 2008). The sensitivity of the ycf1D and rho1-5 mutants to cadmium was tested by making fivefold serial dilutions starting from OD 600 = 2, followed by plating on SC containing 30 mM CdCl 2 .…”

Lipid Hydroperoxides Activate the Mitogen-activated Protein Kinase Mpk1p in Saccharomyces cerevisiae

“…Indeed, deletion of some cell wall genes has been noted to result in sensitivity to LoaOOH (42). However, Mpk1p must also play an active role in the protection against lipid hydroperoxides, since it is activated in the wildtype strain upon exposure to LoaOOH and is required for proper regulation of at least a subset of LoaOOH-responsive genes.…”