scholarly journals The SPI1 Gene, Encoding a Glycosylphosphatidylinositol-Anchored Cell Wall Protein, Plays a Prominent Role in the Development of Yeast Resistance to Lipophilic Weak-Acid Food Preservatives

2006 ◽  
Vol 72 (11) ◽  
pp. 7168-7175 ◽  
Author(s):  
T. Simões ◽  
N. P. Mira ◽  
A. R. Fernandes ◽  
Isabel Sá-Correia
Keyword(s):  
Cell Wall ◽  
Transcription Factor ◽  
Acetic Acid ◽  
Benzoic Acid ◽  
Acid Stress ◽  
Weak Acid ◽  
Cell Wall Protein ◽  
Weak Acids ◽  
Food Preservatives ◽  
Cell Wall Porosity

ABSTRACT The Saccharomyces cerevisiae SPI1 gene encodes a member of the glycosylphosphatidylinositol-anchored cell wall protein family. In this work we show results indicating that SPI1 expression protects the yeast cell from damage caused by weak acids used as food preservatives. This is documented by a less extended period of adaptation to growth in their presence and by a less inhibited specific growth rate for a parental strain compared with a mutant with SPI1 deleted. Maximal protection exerted by Spi1p against equivalent concentrations of the various weak acids tested was registered for the more lipophilic acids (octanoic acid, followed by benzoic acid) and was minimal for acetic acid. Weak-acid adaptation was found to involve the rapid activation of SPI1 transcription, which is dependent on the presence of the Msn2p transcription factor. Activation of SPI1 transcription upon acetic acid stress also requires Haa1p, whereas this recently described transcription factor has a negligible role in the adaptive response to benzoic acid. The expression of SPI1 was found to play a prominent role in the development of yeast resistance to 1,3-β-glucanase in benzoic acid-stressed cells, while its involvement in acetic acid-induced resistance to the cell wall-lytic enzyme is slighter. The results are consistent with the notion that Spi1p expression upon weak-acid stress leads to cell wall remodeling, especially for the more lipophilic acids, decreasing cell wall porosity. Decreased cell wall porosity, in turn, reduces access to the plasma membrane, reducing membrane damage, intracellular acidification, and viability loss.

scholarly journals Weak acid and alkali stress regulate phosphatidylinositol bisphosphate synthesis in Saccharomyces cerevisiae

2006 ◽  
Vol 395 (1) ◽  
pp. 73-80 ◽  
Author(s):  
Mehdi Mollapour ◽  
John P. Phelan ◽  
Stefan H. Millson ◽  
Peter W. Piper ◽  
Frank T. Cooke
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Stress Responses ◽  
Acid Stress ◽  
Weak Acid ◽  
Weak Acids ◽  
Extracellular Medium ◽  
Food Preservatives ◽  
Active Efflux ◽  
Signalling Molecules ◽  
Spoilage Yeasts

Weak organic acids are used as food preservatives to inhibit the growth of spoilage yeasts, including Saccharomyces cerevisiae. Long-term adaptation to weak acids requires the increased expression of the ATP-binding cassette transporter Pdr12p, which catalyses the active efflux of the weak acids from the cytosol; however, very little is known about the signalling events immediately following application of weak acid stress. We have investigated the effects of weak acids on two stress-responsive signalling molecules, PtdIns(3,5)P2 and PtdIns(4,5)P2, which in S. cerevisiae are synthesized by Fab1p and Mss4p respectively. At low extracellular pH, benzoic acid, sorbic acid and acetic acid all cause a transient reduction in PtdIns(3,5)P2 accumulation and a more persistent rise in PtdIns(4,5)P2 levels. The increase in PtdIns(4,5)P2 levels is accompanied by a reorganization of the actin cytoskeleton. However, changes in PtdInsP2 levels are independent of weak acid-induced Pdr12p expression. In contrast, changing the extracellular medium to alkaline pH provokes a prolonged and substantial rise in PtdIns(3,5)P2 levels. As PtdIns(3,5)P2 synthesis is required for correct vacuole acidification, it is possible that levels of this molecule are modulated to maintain intracellular pH homoeostasis in response to weak acid and alkali stresses. In conclusion, we have expanded the repertoire of stress responses that affect PtdInsP2 levels to include weak acid and alkali stresses.

scholarly journals Weak Acid Resistance A (WarA), a Novel Transcription Factor Required for Regulation of Weak-Acid Resistance and Spore-Spore Heterogeneity in Aspergillus niger

mSphere ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Ivey A. Geoghegan ◽  
Malcolm Stratford ◽  
Mike Bromley ◽  
David B. Archer ◽  
Simon V. Avery
Keyword(s):  
Transcription Factor ◽  
Aspergillus Niger ◽  
Sorbic Acid ◽  
Acid Resistance ◽  
Resistance Mechanisms ◽  
Weak Acid ◽  
Benzoic Acids ◽  
Weak Acids ◽  
Food Preservatives ◽  
Content Type

ABSTRACT Propionic, sorbic, and benzoic acids are organic weak acids that are widely used as food preservatives, where they play a critical role in preventing microbial growth. In this study, we uncovered new mechanisms of weak-acid resistance in molds. By screening a library of 401 transcription factor deletion strains in Aspergillus fumigatus for sorbic acid hypersensitivity, a previously uncharacterized transcription factor was identified and named weak acid resistance A (WarA). The orthologous gene in the spoilage mold Aspergillus niger was identified and deleted. WarA was required for resistance to a range of weak acids, including sorbic, propionic, and benzoic acids. A transcriptomic analysis was performed to characterize genes regulated by WarA during sorbic acid treatment in A. niger. Several genes were significantly upregulated in the wild type compared with a ΔwarA mutant, including genes encoding putative weak-acid detoxification enzymes and transporter proteins. Among these was An14g03570, a putative ABC-type transporter which we found to be required for weak-acid resistance in A. niger. We also show that An14g03570 is a functional homologue of the Saccharomyces cerevisiae protein Pdr12p and we therefore name it PdrA. Last, resistance to sorbic acid was found to be highly heterogeneous within genetically uniform populations of ungerminated A. niger conidia, and we demonstrate that pdrA is a determinant of this heteroresistance. This study has identified novel mechanisms of weak-acid resistance in A. niger which could help inform and improve future food spoilage prevention strategies. IMPORTANCE Weak acids are widely used as food preservatives, as they are very effective at preventing the growth of most species of bacteria and fungi. However, some species of molds can survive and grow in the concentrations of weak acid employed in food and drink products, thereby causing spoilage with resultant risks for food security and health. Current knowledge of weak-acid resistance mechanisms in these fungi is limited, especially in comparison to that in yeasts. We characterized gene functions in the spoilage mold species Aspergillus niger which are important for survival and growth in the presence of weak-acid preservatives. Such identification of weak-acid resistance mechanisms in spoilage molds will help in the design of new strategies to reduce food spoilage in the future.

scholarly journals Casein Kinase I Isoform Hrr25 Is a Negative Regulator of Haa1 in the Weak Acid Stress Response Pathway in Saccharomyces cerevisiae

2017 ◽  
Vol 83 (13) ◽  
Author(s):  
Morgan E. Collins ◽  
Joshua J. Black ◽  
Zhengchang Liu
Keyword(s):  
Acetic Acid ◽  
Target Genes ◽  
Acid Stress ◽  
Weak Acid ◽  
Casein Kinase ◽  
Negative Regulator ◽  
Weak Acids ◽  
Casein Kinase I ◽  
Content Type ◽  
Weak Acid Stress

ABSTRACT Haa1 is a transcription factor that adapts Saccharomyces cerevisiae cells to weak organic acid stresses by activating the expression of various genes. Many of these genes encode membrane proteins, such as TPO2 and YRO2. How Haa1 is activated by weak acids is not clear. Here, we show that casein kinase I isoform Hrr25 is an important negative regulator of Haa1. Haa1 is known to be multiply phosphorylated. We found that mutations in HRR25 lead to reduced Haa1 phosphorylation and increased expression of Haa1 target genes and that Hrr25 interacts with Haa1. The other three casein kinase I isoforms, Yck1, Yck2, and Yck3, do not seem to play critical roles in Haa1 regulation. Hrr25 has a 200-residue C-terminal region, including a proline- and glutamine-rich domain. Our data suggest that the C-terminal region of Hrr25 is required for normal inhibition of expression of Haa1 target genes TPO2 and YRO2 and is important for cell growth but is not required for cell morphogenesis. We propose that Hrr25 is an important regulator of cellular adaptation to weak acid stress by inhibiting Haa1 through phosphorylation. IMPORTANCE Our study has revealed the casein kinase I protein Hrr25 to be a negative regulator of Haa1, a transcription factor mediating the cellular response to stresses caused by weak acids. Many studies have focused on the target genes of Haa1 and their roles in weak acid stress responses, but little has been reported on the regulatory mechanism of Haa1. Weak acids, such as acetic acid, have long been used for food preservation by slowing down the growth of fungal species, including S. cerevisiae. In the biofuel industry, acetic acid in the lignocellulosic hydrolysates limits the production of ethanol, which is undesirable. By understanding how Haa1 is regulated, we can make advances in the field of food sciences to better preserve food and engineer acetic acid-resistant strains that will increase productivity in the biofuel industry.

scholarly journals Weak Acid Resistance A (WarA), a novel transcription factor required for regulation of weak-acid resistance and spore-spore heterogeneity in Aspergillus niger

2019 ◽  
Author(s):  
Ivey A. Geoghegan ◽  
Malcolm Stratford ◽  
Mike Bromley ◽  
David B. Archer ◽  
Simon V. Avery
Keyword(s):  
Transcription Factor ◽  
Aspergillus Niger ◽  
Sorbic Acid ◽  
Acid Resistance ◽  
Resistance Mechanisms ◽  
Weak Acid ◽  
Benzoic Acids ◽  
Food Spoilage ◽  
Weak Acids ◽  
Food Preservatives

ABSTRACTPropionic, sorbic and benzoic acids are organic weak acids that are widely used as food preservatives, where they play a critical role in preventing microbial growth. In this study, we uncovered new mechanisms of weak acid resistance in moulds. By screening a library of 401 transcription-factor deletion strains in Aspergillus fumigatus for sorbic acid hypersensitivity, a previously uncharacterised transcription factor was identified, and named as WarA (Weak Acid Resistance A). The orthologous gene in the spoilage mould Aspergillus niger was identified and deleted. WarA was required for resistance to a range of weak acids, including sorbic, propionic and benzoic acids. A transcriptomic analysis was performed to characterise genes regulated by WarA during sorbic acid treatment in A. niger. Several genes were significantly upregulated in the wild type compared with a ΔwarA mutant, including genes encoding putative weak acid detoxification enzymes and transporter proteins. Among these was An14g03570, a putative ABC-type transporter which we found to be required for weak acid resistance in A. niger. We also show that An14g03570 is a functional homologue of the Saccharomyces cerevisiae protein Pdr12p, and therefore named as PdrA. Lastly, resistance to sorbic acid was found to be highly heterogeneous within genetically-uniform populations of ungerminated A. niger conidia, and we demonstrate that pdrA is a determinant of this heteroresistance. This study has identified novel mechanisms of weak acid resistance in A. niger which could help to inform and improve future food spoilage prevention strategies.IMPORTANCEWeak acids are widely used as food preservatives, as they are very effective at preventing growth of most species of bacteria and fungi. However, some species of moulds can survive and grow in the concentrations of weak acid employed in food and drink products, thereby causing spoilage with resultant risks for food security and health. Current knowledge of weak acid resistance mechanisms in these fungi is limited, especially in comparison to that in yeasts. We characterised gene functions in the spoilage mould species Aspergillus niger which are important for survival and growth in the presence of weak acid preservatives. Such identification of weak acid resistance mechanisms in spoilage moulds will help to design new strategies to reduce food spoilage in the future.

Plasma membrane H+ and K+ transporters are involved in the weak-acid preservative response of disparate food spoilage yeasts

Microbiology ◽  
2005 ◽  
Vol 151 (6) ◽  
pp. 1995-2003 ◽  
Author(s):  
Neil Macpherson ◽  
Lana Shabala ◽  
Henrietta Rooney ◽  
Marcus G. Jarman ◽  
Julia M. Davies
Keyword(s):  
Plasma Membrane ◽  
Benzoic Acid ◽  
Acid Stress ◽  
Weak Acid ◽  
Uptake System ◽  
Food Spoilage ◽  
Adaptive Responses ◽  
Spoilage Yeasts ◽  
Food Spoilage Yeasts

The food spoilage yeasts Zygosaccharomyces bailii and Saccharomyces cerevisiae have been proposed to resist weak-acid preservative stress by different means; Z. bailii by limiting influx of preservative combined with its catabolism, S. cerevisiae by active extrusion of the preservative weak-acid anion and H+. Measurement of H+ extrusion by exponential-phase Z. bailii cells suggest that, in common with S. cerevisiae, this yeast uses a plasma membrane H+-ATPase to expel H+ when challenged by weak-acid preservative (benzoic acid). Simultaneous measurement of Z. bailii net H+ and K+ fluxes showed that net K+ influx accompanies net H+ efflux during acute benzoic acid stress. Such ionic coupling is known for S. cerevisiae in short-term preservative stress. Both yeasts significantly accumulated K+ on long-term exposure to benzoic acid. Analysis of S. cerevisiae K+ transporter mutants revealed that loss of the high affinity K+ uptake system Trk1 confers sensitivity to growth in preservative. The results suggest that cation accumulation is an important factor in adaptation to weak-acid preservatives by spoilage yeasts and that Z. bailii and S. cerevisiae share hitherto unsuspected adaptive responses at the level of plasma membrane ion transport.

scholarly journals Identification of a DNA-binding site for the transcription factor Haa1, required for Saccharomyces cerevisiae response to acetic acid stress

2011 ◽  
Vol 39 (16) ◽  
pp. 6896-6907 ◽  
Author(s):  
Nuno P. Mira ◽  
Sílvia F. Henriques ◽  
Greg Keller ◽  
Miguel C. Teixeira ◽  
Rute G. Matos ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcription Factor ◽  
Acetic Acid ◽  
Dna Binding ◽  
Binding Site ◽  
Acid Stress ◽  
Dna Binding Site

scholarly journals Intracellular pH Response to Weak Acid Stress in Individual Vegetative Bacillus subtilis Cells

2016 ◽  
Vol 82 (21) ◽  
pp. 6463-6471 ◽  
Author(s):  
Rachna Pandey ◽  
Norbert O. E. Vischer ◽  
Jan P. P. M. Smelt ◽  
Johan W. A. van Beilen ◽  
Alexander Ter Beek ◽  
...  
Keyword(s):  
Organic Acid ◽  
Single Cell ◽  
Acid Stress ◽  
Weak Acid ◽  
Food Preservation ◽  
Cell Level ◽  
Food Preservatives ◽  
Content Type ◽  
Weak Organic Acid ◽  
Weak Acid Stress

ABSTRACTIntracellular pH (pHi) critically affects bacterial cell physiology. Hence, a variety of food preservation strategies are aimed at perturbing pHihomeostasis. Unfortunately, accurate pHiquantification with existing methods is suboptimal, since measurements are averages across populations of cells, not taking into account interindividual heterogeneity. Yet, physiological heterogeneity in isogenic populations is well known to be responsible for differences in growth and division kinetics of cells in response to external stressors. To assess in this context the behavior of intracellular acidity, we have developed a robust method to quantify pHiat single-cell levels inBacillus subtilis. Bacilli spoil food, cause disease, and are well known for their ability to form highly stress-resistant spores. Using an improved version of the genetically encoded ratiometric pHluorin (IpHluorin), we have quantified pHiin individualB. subtiliscells, cultured at an external pH of 6.4, in the absence or presence of weak acid stresses. In the presence of 3 mM potassium sorbate, a decrease in pHiand an increase in the generation time of growing cells were observed. Similar effects were observed when cells were stressed with 25 mM potassium acetate. Time-resolved analysis of individual bacteria in growing colonies shows that after a transient pH decrease, long-term pH evolution is highly cell dependent. The heterogeneity at the single-cell level shows the existence of subpopulations that might be more resistant and contribute to population survival. Our approach contributes to an understanding of pHiregulation in individual bacteria and may help scrutinizing effects of existing and novel food preservation strategies.IMPORTANCEThis study shows how the physiological response to commonly used weak organic acid food preservatives, such as sorbic and acetic acids, can be measured at the single-cell level. These data are key to coupling often-observed single-cell heterogeneous growth behavior upon the addition of weak organic acid food preservatives. Generally, these data are gathered in the form of plate counting of samples incubated with the acids. Here, we visualize the underlying heterogeneity in cellular pH homeostasis, opening up avenues for mechanistic analyses of the heterogeneity in the weak acid stress response. Thus, microbial risk assessment can become more robust, widening the scope of use of these well-known weak organic acid food preservatives.

scholarly journals Yeast adaptive response to acetic acid stress involves structural alterations and increased stiffness of the cell wall

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ricardo A. Ribeiro ◽  
Miguel V. Vitorino ◽  
Cláudia P. Godinho ◽  
Nuno Bourbon-Melo ◽  
Tiago T. Robalo ◽  
...  
Keyword(s):  
Cell Wall ◽  
Acetic Acid ◽  
Yeast Cell ◽  
Adaptive Response ◽  
Time Course ◽  
Acid Stress ◽  
Cell Stiffness ◽  
Gene Encoding ◽  
Futile Cycle ◽  
Robust Adaptive

AbstractThis work describes a coordinate and comprehensive view on the time course of the alterations occurring at the level of the cell wall during adaptation of a yeast cell population to sudden exposure to a sub-lethal stress induced by acetic acid. Acetic acid is a major inhibitory compound in industrial bioprocesses and a widely used preservative in foods and beverages. Results indicate that yeast cell wall resistance to lyticase activity increases during acetic acid-induced growth latency, corresponding to yeast population adaptation to sudden exposure to this stress. This response correlates with: (i) increased cell stiffness, assessed by atomic force microscopy (AFM); (ii) increased content of cell wall β-glucans, assessed by fluorescence microscopy, and (iii) slight increase of the transcription level of the GAS1 gene encoding a β-1,3-glucanosyltransferase that leads to elongation of (1→3)-β-d-glucan chains. Collectively, results reinforce the notion that the adaptive yeast response to acetic acid stress involves a coordinate alteration of the cell wall at the biophysical and molecular levels. These alterations guarantee a robust adaptive response essential to limit the futile cycle associated to the re-entry of the toxic acid form after the active expulsion of acetate from the cell interior.

scholarly journals The pH-Responsive Transcription Factors YlRim101 and Mhy1 Regulate Alkaline pH-Induced Filamentation in the Dimorphic Yeast Yarrowia lipolytica

mSphere ◽  
2021 ◽  
Vol 6 (3) ◽  
Author(s):  
Tao Shu ◽  
Xin-Yu He ◽  
Jia-Wen Chen ◽  
Yi-Sheng Mao ◽  
Xiang-Dong Gao
Keyword(s):  
Cell Wall ◽  
Transcription Factor ◽  
Yarrowia Lipolytica ◽  
Regulatory Mechanism ◽  
Cell Wall Protein ◽  
Alkaline Ph ◽  
Dimorphic Fungi ◽  
Content Type ◽  
Dimorphic Yeast

ABSTRACT Environmental pH influences cell growth and differentiation. In the dimorphic yeast Yarrowia lipolytica, neutral-alkaline pH strongly induces the yeast-to-filament transition. However, the regulatory mechanism that governs alkaline pH-induced filamentation has been unclear. Here, we show that the pH-responsive transcription factor Y. lipolytica Rim101 (YlRim101) is a major regulator of alkaline-induced filamentation, since the deletion of YlRIM101 severely impaired filamentation at alkaline pH, whereas the constitutively active YlRIM1011-330 mutant mildly induced filamentation at acidic pH. YlRim101 controls the expression of the majority of alkaline-regulated cell wall protein genes. One of these, the cell surface glycosidase gene YlPHR1, plays a critical role in growth, cell wall function, and filamentation at alkaline pH. This finding suggests that YlRim101 promotes filamentation at alkaline pH via controlling the expression of these genes. We also show that, in addition to YlRim101, the Msn2/Msn4-like transcription factor Mhy1 is highly upregulated at alkaline pH and is essential for filamentation. However, unlike YlRim101, which specifically regulates alkaline-induced filamentation, Mhy1 regulates both alkaline- and glucose-induced filamentation, since the deletion of MHY1 abolished them both, whereas the overexpression of MHY1 induced strong filamentation irrespective of the pH or the presence of glucose. Finally, we show that YlRim101 and Mhy1 positively coregulate seven cell wall protein genes at alkaline pH, including YlPHR1 and five cell surface adhesin-like genes, three of which appear to promote filamentation. Together, these results reveal a conserved role of YlRim101 and a novel role of Mhy1 in the regulation of alkaline-induced filamentation in Y. lipolytica. IMPORTANCE The regulatory mechanism that governs pH-regulated filamentation is not clear in dimorphic fungi except in Candida albicans. Here, we investigated the regulation of alkaline pH-induced filamentation in Yarrowia lipolytica, a dimorphic yeast distantly related to C. albicans. Our results show that the transcription factor YlRim101 and the Msn2/Msn4-like transcription factor Mhy1 are the major regulators that promote filamentation at alkaline pH. They control the expression of a number of cell wall protein genes important for cell wall organization and filamentation. Our results suggest that the Rim101/PacC homologs play a conserved role in pH-regulated filamentation in dimorphic fungi.

Sign in / Sign up

Export Citation Format

Share Document