The sensitivity of the yeast, Saccharomyces cerevisiae, to acetic acid is influenced by DOM34 and RPL36A

The presence of acetic acid during industrial alcohol fermentation reduces the yield of fermentation by imposing additional stress on the yeast cells. The biology of cellular responses to stress has been a subject of vigorous investigations. Although much has been learned, details of some of these responses remain poorly understood. Members of heat shock chaperone HSP proteins have been linked to acetic acid and heat shock stress responses in yeast. Both acetic acid and heat shock have been identified to trigger different cellular responses including reduction of global protein synthesis and induction of programmed cell death. Yeast HSC82 and HSP82 code for two important heat shock proteins that together account for 1-2% of total cellular proteins. Both proteins have been linked to responses to acetic acid and heat shock. In contrast to the overall rate of protein synthesis which is reduced, the expression of HSC82 and HSP82 is induced in response to acetic acid stress. In the current study we identified two yeast genes DOM34 and RPL36A that are linked to acetic acid and heat shock sensitivity. We investigated the influence of these genes on the expression of HSP proteins. Our observations suggest that Dom34 and RPL36A influence translation in a CAP-independent manner.This work was funded by the Natural Sciences and Engineering Research Council of Canada, NSERC.info:eu-repo/semantics/publishedVersio

Mode of action of nisin on Escherichia coli

Nisin is a class I polycyclic bacteriocin produced by the bacterium Lactococcus lactis, which is used extensively as a food additive to inhibit the growth of foodborne Gram-positive bacteria. Nisin also inhibits growth of Gram-negative bacteria when combined with membrane-disrupting chelators such as citric acid. To gain insight into nisin’s mode of action, we analyzed chemical–genetic interactions and identified nisin-sensitive Escherichia coli strains in the Keio library of knockout mutants. The most sensitive mutants fell into two main groups. The first group accords with the previously proposed mode of action based on studies with Gram-positive bacteria, whereby nisin interacts with factors involved in cell wall, membrane, envelope biogenesis. We identified an additional, novel mode of action for nisin based on the second group of sensitive mutants that involves cell cycle and DNA replication, recombination, and repair. Further analyses supported these two distinct modes of action.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Disruption of protein synthesis as antifungal mode of action by chitosan

The antimicrobial activity of chitosan has been acknowledged for more than 30. years and yet its mode-of-action remains ambiguous. We analyzed chemical-genetic interactions of low-molecular weight chitosan using a collection of ≈ 4600 S. cerevisiae deletion mutants and found that 31% of the 107 mutants most sensitive to chitosan had deletions of genes related primarily to functions involving protein synthesis. Disruption of protein synthesis by chitosan was substantiated by an in vivo β-galactosidase expression assay suggesting that this is a primary mode of antifungal action. Analysis of the yeast gene deletion array and secondary assays also indicate that chitosan has a minor membrane disruption effect - a leading model of chitosan antimicrobial activity

Zinc oxide and silver nanoparticles toxicity in the baker's yeast, Saccharomyces cerevisiae.

Engineered nanomaterials (ENMs) are increasingly incorporated into a variety of commercial applications and consumer products; however, ENMs may possess cytotoxic properties due to their small size. This study assessed the effects of two commonly used ENMs, zinc oxide nanoparticles (ZnONPs) and silver nanoparticles (AgNPs), in the model eukaryote Saccharomyces cerevisiae. A collection of ≈4600 S. cerevisiae deletion mutant strains was used to deduce the genes, whose absence makes S. cerevisiae more prone to the cytotoxic effects of ZnONPs or AgNPs. We demonstrate that S. cerevisiae strains that lack genes involved in transmembrane and membrane transport, cellular ion homeostasis, and cell wall organization or biogenesis exhibited the highest sensitivity to ZnONPs. In contrast, strains that lack genes involved in transcription and RNA processing, cellular respiration, and endocytosis and vesicular transport exhibited the highest sensitivity to AgNPs. Secondary assays confirmed that ZnONPs affected cell wall function and integrity, whereas AgNPs exposure decreased transcription, reduced endocytosis, and led to a dysfunctional electron transport system. This study supports the use of S. cerevisiae Gene Deletion Array as an effective high-throughput technique to determine cellular targets of ENM toxicity

Mode of action of nisin on escherichia coli

Nisin is a class I polycyclic bacteriocin produced by the bacterium Lactococcus lactis, which is used extensively as a food additive to inhibit the growth of foodborne Gram-positive bacteria. Nisin also inhibits growth of Gram-negative bacteria when combined with membrane-disrupting chelators such as citric acid. To gain insight into nisin’s mode of action, we analyzed chemical–genetic interactions and identified nisin-sensitive Escherichia coli strains in the Keio library of knockout mutants. The most sensitive mutants fell into two main groups. The first group accords with the previously proposed mode of action based on studies with Gram-positive bacteria, whereby nisin interacts with factors involved in cell wall, membrane, envelope biogenesis. We identified an additional, novel mode of action for nisin based on the second group of sensitive mutants that involves cell cycle and DNA replication, recombination, and repair. Further analyses supported these two distinct modes of action

Functional distribution of deletion mutants that are highly sensitive to ZnONPs and AgNPs.

<p>(A) Clustering of the 59 most sensitive deletion mutant strains to 1 mg/mL ZnONPs reveal that mutants lacking genes involved in membrane and transmembrane transport, ion homeostasis and transport, and cell organization of biogenesis encompass the significantly enriched groups. (B) Clustering of the 96 most sensitive deletion mutant strains to 0.095 mg/mL AgNPs indicate that mutants lacking genes involved in transcription, cellular respiration, and endocytosis and vesicular transport represent the significantly enriched groups.</p

Transcription rate, cellular respiration, and endocytosis in yeast cells exposed to AgNPs.

<p>(A) β-galactosidase reporter gene expression assay was used to estimate transcription rate in response to AgNPs or 6-azauracil (positive control). β-galactosidase activity in treatments is expressed relative to no-AgNPs control (negative control). (B) MTT reduction assay was used to estimate cellular respiration in response to AgNPs or sodium azide (positive control). (C) Brightfield, fluorescence, and merged images of negative control, AgNPs (80 μg/mL), and positive control (NaN₃) groups. The cells that internalized Lucifer Yellow (LY) are fluorescent. (D) The uptake of LY was used to estimate endocytosis in response to AgNPs. Percentage of fluorescent cells relative to control was determined by examining at least 6 different fields, each with >20 cells. Mean ± SEM are presented (n ≥ 3). Significant differences are indicated with letters.</p

ZnONPs compromise cell wall integrity.

<p>(A) Cells were exposed to various concentrations of ZnONPs, subjected to mild sonication, diluted and then spotted onto YPD and compared to non-sonicated counterparts. (B) Cell survival rate (% survival sonicated/non-sonicated cells) was then quantified. Mean ± SEM are presented (n ≥ 3). Significant differences are indicated with letters.</p

Size characterization of nanoparticles.

<p>(A) Dynamic light scattering (DLS) was used to assess the size of ZnONPs and AgNPs. Mean ± SEM are presented (n ≥ 3). (B) Transmission electron microscopy images of ZnONPs and AgNPs (Note: the scale bars are 200 and 20 nm for ZnONPs and AgNPs, respectively).</p

Cell membrane disruption by ZnONPs.

<p>(A) Carboxyfluorescein leakage from liposomes that were exposed to various concentrations of ZnONPs for 30 min. (B) Trypan blue exclusion assay in yeast cells following a 2 h exposure to various concentrations of ZnONPs. Mean ± SEM are presented (n ≥ 3). Significant differences are indicated with letters. (C) Membrane depolarization analysis in yeast cells that were subjected to: 0.01 mM citric acid (negative control), 20 μM CCCP (carbonyl cyanide 3-chlorophenylhydrazone; positive control), and 1 mg/mL ZnONPs. The histograms show the number (Count, Y-axis) of yeast cells in a sample with depolarized membranes (FL2-H interval = 10⁴–10⁵) and cells at resting potential (FL2-H = 10²–10⁴).</p