Accumulation Of Intracellular Trehalose Research Articles

Trehalose enhances the resistance of Lactiplantibacillus plantarum LIP-1 to spray- and freeze-drying treatments by regulating amino acid metabolism

The effect of adding trehalose in the culture medium on Lactiplantibacillus plantarum LIP-1 was investigated during spray- and freeze-drying. Although previous studies reported that the enhancement of bacterial resistance is due to intracellular trehalose accumulation, our study revealed that it was related to trehalose metabolism by the strain, and not accumulation. The underlying mechanisms were mainly related to the activation of l-glutamic acid and cysteine metabolic pathways during trehalose metabolism. First, murL, murD, and vanY in the l-glutamic acid metabolic pathway were significantly upregulated, increasing peptidoglycan synthesis and thus improving the cell wall integrity during drying process. Second, gshA in the cysteine metabolic pathway was upregulated, increasing intracellular glutathione content, enhancing antioxidant capacity, and thereby reducing the degree of cell membrane damage during drying process. Third, alkaline substances produced during l-glutamic acid and cysteine metabolisms increased the intracellular pH and reduced the degree of DNA damage caused by acid stress.

A universal mechanism on desiccation tolerance of Cronobacter based on intracellular trehalose accumulation regulated by EnvZ/OmpR

Cronobacter (seven species) can survive in dry powdered infant formula for a long time, but the thorough molecular mechanism of resistance to desiccation remains elusive. Here we examine the regulation mechanism of Cronobacter's tolerance to desiccation by the typical two-component system (TCS) EnvZ/OmpR. When exposed to desiccation conditions, Cronobacter showed higher survival than other pathogens, as well as significantly up-regulated expression of ompR and otsAB genes with markedly decreased survival of their mutants, suggesting their relationship with desiccation tolerance. OmpR directly binds to the promoter of trehalose biosynthesis operon otsBA, significantly enhancing their expression, and boosting the trehalose levels. The ompR-deletion in other six species further confirmed its positive regulation in desiccation tolerance. Our data present a hypothesis that EnvZ/OmpR increases intracellular trehalose levels against damage to the cells, which prompts Cronobacter to survive in desiccation conditions. This study reveals a universal molecular mechanism for desiccation resistance in Cronobacter species.

Deciphering cell wall sensors enabling the construction of robust P. pastoris for single-cell protein production

Single-cell protein (SCP) production in the methylotrophic yeast Pichia pastoris has the potential to achieve a sustainable protein supply. However, improving the methanol fermentation efficiency and reducing carbon loss has been a long-standing challenge with far-reaching scientific and practical implications. Here, comparative transcriptomics revealed that PAS_0305, a gene directly associated with cell wall thickness under methanol stress, can be used as a target for unlocking cell wall sensors. Intracellular trehalose accumulation confirmed that cell wall sensors were activated after knocking out PAS_0305, which resulted in increased cell wall permeability. Genome-wide signal perturbations were transduced through the HOG module and the CWI pathway, which was confirmed to connected by Pbs2-Mkk. As a consequence of CWI pathway activation, ΔPAS_0305 elicited a rescue response of cell wall remodeling by increasing the β-1,3-glucan content and decreasing the chitin/mannose content. Remarkably, perturbations in global stress signals led to a fine-tuning of the metabolic network of ΔPAS_0305, resulting in a superior phenotype with highest crude protein and methanol conversion rate of 67.21% and 0.46 gDCW/g. Further genome-scale metabolic models were constructed to validate the experimental results, confirming that unlocking cell wall sensors resulted in maximized flux from methanol towards SCP and effectively addressing the issue of carbon loss in methanol fermentation. This work sheds new light on the potential of manipulating cellular signaling pathways to optimize metabolic networks and achieve exceptional phenotypic characteristics, providing new strategies for constructing versatile cell factories in P. pastoris.

Exogenous trehalose application promotes survival by alleviating oxidative stress and affecting transcriptome in ethanol-stressed Wickerhamomyces anomalus

PurposeIncreased ethanol accumulation during ethanol fermentation generates stress in yeast cells, which finally reduces the fermentation performance and efficiency. Trehalose, a potential stress protectant, has been reported to regulate the response of yeast to diverse environmental stresses. This study aimed to explore how exogenous trehalose application affects the survival, transcriptome and antioxidant enzymes of Wickerhamomyces anomalus grown under ethanol stress conditions.Design/methodology/approachExogenous trehalose was applied to the growth condition of W. anomalus, and optical densitometric method was used to detect contents of intracellular trehalose and MDA and activities of CAT and SOD. The survival was evaluated using spot analysis. Differentially expressed genes (DEGs) were identified through transcriptomics analysis.FindingsThe results showed that ethanol stress induced the accumulation of intracellular trehalose, with further exogenous trehalose application improving the survival and alleviating oxidative stress in ethanol-stressed W. anomalus. Transcriptomic results showed that trehalose has pleiotropic regulating effects on ethanol-stressed W. anomalus since most DEGs annotated to energy metabolism, amino acid metabolism, translation, folding, sorting and transport were affected post trehalose addition. Therefore, it is found that trehalose protected W. anomalus against ethanol stress, and these findings provide interesting insights into the mechanistic role of trehalose in improving ethanol stress tolerance of W. anomalus.Originality/value(1) Protective effect of exogenous trehalose addition on the survival of ethanol-stressed W. anomalus was proved. (2) Exogenous trehalose addition could partly alleviate oxidative stress induced by ethanol stress and affect transcriptome in W. anomalus.

Higher thermal and ethanol tolerance of a yeast strain isolated from oral cavity

Efficient bioethanol producing microorganisms must be endowed with peculiar physiological and technological traits, such as, higher thermal and ethanol tolerance. This encompasses a strain thermotolerance and an ability to grow at elevated sugar and ethanol concentration or ability to sustain dehydration process such as freeze drying. In this study, we characterized a thermotolerant yeast strain isolated from the human oral cavity regarding the above mentioned parameters. Such an uncommon niche was considered as the great potential reservoir, to isolate strains endowed with metabolic aptitudes requested for ethanol production. In the process, we have defined the YTerm-1strain ability to sustain high sugar and ethanol concentration that are two technological constrains in bioethanol production. Finally, we highlighted that the strain YTerm-1 was able to accumulate, at a high level, trehalose and β-glucan, two compounds conferring the cells an increased resistance to freeze drying process and osmotic stress. Our results suggest that the Yterm-1 strain showed a better growth ability and higher ethanol yield as compared to the industrial strain. Other metabolic traits, such as resistance to dehydration stress, tolerance to ethanol, accumulation of intracellular trehalose or membrane β-glucan confer to that isolate all the characteristics requested in industrial production of ethanol.

Enhancing long-term storage and stability of engineered living materials through desiccant storage and trehalose treatment.

Engineered living materials (ELMs) have broad applications for enabling on-demand bioproduction of compounds ranging from small molecules to large proteins. However, most formulations and reports lack the capacity for storage beyond a few months. In this study, we develop an optimized procedure to maximize stress resilience of yeast-laden ELMs through the use of desiccant storage and 10% trehalose incubation before lyophilization. This approach led to over 1-year room temperature storage stability across a range of strain genotypes. In particular, we highlight the superiority of exogenously added trehalose over endogenous, engineered production in yielding robust preservation resilience that is independent of cell state. This simple, effective protocol enables sufficient accumulation of intracellular trehalose over a short period of contact time across a range of strain backgrounds without requiring the overexpression of a trehalose importer. A variety of microscopic analysis including µ-CT and confocal microscopy indicate that cells form spherical colonies within F127-BUM ELMs that have variable viability upon storage. The robustness of the overall procedure developed here highlights the potential for widespread deployment to enable on-demand, cold-chain independent bioproduction.

Metabolic characteristics of intracellular trehalose enrichment in salt-tolerant Zygosaccharomyces rouxii.

The salt-tolerant flavor yeast Zygosaccharomyces rouxii is an important food flavor microorganism, but its intracellular stress-resistant trehalose synthesis efficiency has been shown to be low, resulting in its weak high-temperature resistance. The intracellular and extracellular levels of carbohydrates, organic acids, and amino acids of Z. rouxii in a 20-L mechanically stirred ventilated fermenter were analyzed using metabolomics research methods. Our results showed that glucose supplementation could promote the growth of yeast cells, but high temperatures (> 35°C) significantly prevented cell growth. Under three different growth strategies, extracellular glucose was continuously utilized and intracellular glucose was continuously metabolized, but glucose overflow metabolism was inhibited by high temperature, which showed that the level of intracellular/extracellular ethanol was stable. High temperature stimulated significant intracellular trehalose accumulation (c20.5h = 80.78 mg/g Dry Cell Weight (DCW)) but not efflux, as well as xylitol accumulation (c20.5h = 185.97 mg/g DCW) but with efflux (c20.5h = 29.78 g/L). Moreover, heat resistance evaluation showed that xylitol and trehalose had heat-protective effects on Z. rouxii. In addition, a large amount of propionic acid and butyric acid accumulated inside and outside these cells, showing that the conversion of glucose to acid in yeast cells becomes the main pathway of glucose overflow metabolism in high temperatures. In addition, the increased demand of yeast cells for phenylalanine, threonine, and glycine at high temperatures suggested that these metabolites participated in the temperature adaptation of Z. rouxii in different ways. Valine and leucine/isoleucine [branched-chain amino acids (BCAAs)] were mainly affected by the addition of glucose, while glucose, sucrose, aspartic acid/asparagine, and glutamate/glutamine were not affected by this temperature regulation as a whole. This study could help deepen our understanding of the high-temperature adaptation mechanism of salt-tolerant Z. rouxii, and has theoretical significance for the application of highly tolerant yeast to food brewing.

Trehalose accumulation and radiation resistance due to prior heat stress in Saccharomyces cerevisiae

In this study, we examined the accumulation of trehalose, a stress-responsive substance, upon gamma-ray irradiation by evaluating the cause of trehalose accumulation and the development of gamma-ray resistance through intracellular trehalose accumulation. Saccharomyces cerevisiae cells cultured to the logarithmic growth phase were irradiated with gamma rays, and the intracellular trehalose content was measured. However, trehalose was not detectable. The yeast cells with trehalose accumulation caused by pre-treatment at 40°C were irradiated with gamma rays, and the resistance of these cells to gamma radiation was compared with that of cells without heat treatment. Trehalose accumulation resulted in gamma-ray resistance and suppressed the increase in reactive oxygen species, lipid peroxidation, and DNA double-strand break production in yeast cells. The tests were also performed with a trehalose-6-phosphate-synthase (TPS1)-deficient mutant strain (Δtps1) unable to synthesize trehalose, and the results revealed that TPS1 was involved in protection against oxidative stress.

Oligosaccharide-phenolic compound conjugates in soluble polysaccharides from rice straw alleviate ethanol fermentation stresses in Saccharomyces cerevisiae

In this study, it was found that soluble polysaccharides (SPs) from rice straw were conjugated with high levels of phenolic compounds. A gradient ethanol precipitation method was used to separate conjugates with different phenolic compound components. These conjugates were mainly composed of disaccharides or trisaccharides and phenolic acids that were mainly p-coumaric acid, ferulic acid, vanillic acid, and vanillin. SPs with more conjugates exhibited better effects on Saccharomyces cerevisiae and ethanol fermentation. The change in the proportion of conjugates in SPs significantly affected the accumulation of intracellular trehalose and glycerol content, indicating the effect of the conjugates on intracellular osmotic stress. Moreover, SPs with more conjugates generated lower intracellular hydrogen peroxide (H2O2) levels. Thus, oligosaccharide and phenolic compound conjugates in SPs promoted yeast growth and ethanol fermentation. For instance, OD value reached 9.4, which was 1.3 times higher than the control group, and ethanol content at 36 h was 36.5 g/L, which was 1.3 times higher than the control. Osmotic pressure alleviation and lowered intracellular H2O2 levels by conjugates are key mechanisms of the SPs during yeast growth and ethanol fermentation.

Macroscopic Modeling of Intracellular Trehalose Concentration in Saccharomyces cerevisiae Fed-batch Cultures

This paper describes the extension of a baker's yeast growth model to account for the intracellular trehalose storage and mobilization. Trehalose is a reserve carbohydrate that is accumulated and converted back to intracellular glucose when the yeast cells face certain stresses. This is modeled by a new macroscopic reaction, which is coupled to an existing macroscopic reaction scheme describing the coordinated uptake of glucose and ammonium by the yeast cells. The dynamics of the trehalose concentration is described by a delay differential equation as the available experimental data used to fit the model exhibits a time-delayed correlation between trehalose storage and glucose uptake as well as a time-delayed correlation between trehalose mobilization and ethanol respiration phases. The proposed extension contains 6 parameters of which 5 are estimated via nonlinear least squares identification. The proposed model predicts accurately the dynamics of trehalose storage and mobilization and can be used to optimize the intracellular trehalose accumulation in Saccharomyces cerevisiae, which is valuable for obtaining reinforced yeast cells, able to better withstand drying operations.

Temperature and presence of ethanol affect accumulation of intracellular trehalose in Lactobacillus plantarum WCFS1 upon pulsed electric field treatment

Pulsed electric field (PEF) treatment can be used to increase intracellular small molecule concentrations in bacteria, which can lead to enhanced robustness of these cells during further processing. In this study we investigated the effects of the PEF treatment temperature and the presence of 8% (v/v) ethanol in the PEF medium on cell survival, membrane fluidity and intracellular trehalose concentrations of Lactobacillus plantarum WCFS1. A moderate PEF treatment temperature of 21 °C resulted in a high cell survival combined with higher intracellular trehalose concentrations compared to a treatment at 10 and 35 °C. Interestingly, highest intracellular trehalose concentrations were observed upon supplementing the PEF medium with 8% ethanol, which resulted in more than a doubling in intracellular trehalose concentrations, while culture survival was retained. Overall, this study shows that treatment temperature and PEF medium optimization are important directions for improving molecule uptake upon PEF processing.

The MAPK Hog1 mediates the response to amphotericin B in Candida albicans

The HOG MAP kinase pathway plays a crucial role in the response to different stresses in the opportunistic pathogen Candida albicans. The polyene amphotericin B (AMB) has been reported to trigger oxidative stress in several pathogenic fungi, including C. albicans. In the present work, we have analyzed the role of the MAPK Hog1 in sensing and survival to AMB treatment. Mutants lacking Hog1 are more susceptible to AMB than their parental strains and Hog1 became phosphorylated in the presence of this polyene. A set of mutated versions of Hog1 revealed that both the kinase activity and phosphorylation of Hog1 are required to cope with AMB treatment. Flow cytometry analysis showed that AMB induced intracellular ROS accumulation in both parental and hog1 null mutant strains. In addition, AMB triggered a Hog1-independent synthesis of trehalose. The addition of rotenone to AMB-treated cells improved cell viability, decreased intracellular ROS and prevented intracellular trehalose accumulation, suggesting that AMB-induced ROS is associated to a functional electron transport chain but the presence of rotenone did not impair Hog1 phosphorylation in AMB-treated cells. Our results indicate that Hog1 is necessary during AMB treatment to increase its survival.

Accumulation of intracellular trehalose and lactose in Lactobacillus plantarum WCFS1 during pulsed electric field treatment and subsequent freeze and spray drying

Survival of bacteria during drying processes is required for products such as probiotics or starter cultures. In earlier studies high intracellular trehalose concentrations have been related to processing robustness during freezing and drying for several cell types. In this study we evaluated pulsed electric field (PEF) pre-treatment to increase intracellular trehalose in Lactobacillus plantarum WCFS1 and subsequent survival during freeze drying and storage in a trehalose matrix. Surprisingly, despite increased intracellular trehalose by PEF pre-treatment no enhanced survival was observed after drying and storage. Additional analysis of intracellular trehalose before and after freeze drying and spray drying revealed that during the drying process intracellular trehalose concentrations increased. Due to this increase the intracellular trehalose concentrations in the PEF pre-treated and control samples did not differ anymore. To explain what happens during the drying processes, results for trehalose were compared to similar experiments with lactose. A remarkable difference was observed between transfer of trehalose and lactose into cells during the freezing step before freeze drying, while both disaccharides gave similar cellular protection when used as freezing and drying matrix. Generated knowledge on transport of disaccharides into bacterial cells during freezing and drying can benefit processing of living bacterial formulations.

Adaptive evolution and selection of stress-resistant Saccharomyces cerevisiae for very high-gravity bioethanol fermentation

Abstract Background In industrial yeasts, selection and breeding for resistance to multiple stresses is a focus of current research. The objective of this study was to investigate the tolerance to multiple stresses of Saccharomyces cerevisiae obtained through an adaptive laboratory evolution strategy involving a repeated liquid nitrogen freeze–thaw process coupled with multi-stress shock selection. We also assessed the related resistance mechanisms and very high-gravity (VHG) bioethanol production of this strain. Results Elite S. cerevisiae strain YF10-5, exhibiting improved VHG fermentation capacity and stress resistance to osmotic pressure and ethanol, was isolated following ten consecutive rounds of liquid nitrogen freeze–thaw treatment followed by plate screening under osmotic and ethanol stress. The ethanol yield of YF10-5 was 16% higher than that of the parent strain during 35% (w/v) glucose fermentation. Furthermore, there was upregulation of three genes (HSP26, HSP30, and HSP104) encoding heat-shock proteins involved in the stress response, one gene (TPS1) involved in the synthesis of trehalose, and three genes (ADH1, HXK1, and PFK1) involved in ethanol metabolism and intracellular trehalose accumulation in YF10-5 yeast cells, indicating increased stress tolerance and fermentative capacity. YF10-5 also showed excellent fermentation performance during the simultaneous saccharification and fermentation of VHG sweet potato mash, producing 13.40% (w/v) ethanol, which corresponded to 93.95% of the theoretical ethanol yield. Conclusions A multiple-stress-tolerant yeast clone was obtained using adaptive evolution by a freeze–thaw method coupled with stress shock selection. The selected robust yeast strain exhibits potential for bioethanol production through VHG fermentation. How to cite: Zhang Q, Jin Y, Fang Y, et al. Adaptive evolution and selection of stress-resistant Saccharomyces cerevisiae for very high gravity bioethanol fermentation. Electron J Biotechnol 2019;41. https://doi.org/10.1016/j.ejbt.2019.06.003

Protective roles of trehalose in Pleurotus pulmonarius during heat stress response

High temperature is one of the major abiotic stresses that limit edible mushroom growth and development. The understanding of physiological alterations in response to heat stress and the corresponding mechanisms involved is vital for the breeding of heat-resistant edible mushroom strains. Although trehalose functions as a protectant against abiotic stresses in fungi, the putative role of trehalose in thermotolerance remains to be elucidated. In this study, we found heat stress inhibited the growth of two Pleurotus pulmonarius strains, heat-sensitive and less-sensitive, and the inhibition was more significant for the sensitive strain. Heat stress leads to the increase of lipid peroxidation and intracellular trehalose accumulation, with a higher level in the heat-sensitive strain, and this effect is independent of exogenous trehalose application. In addition, a lower concentration of exogenous trehalose application in sensitive strain than in less-sensitive strain was found to alleviate the inhibition of mycelium growth and further increase the intracellular trehalose concentration by heat stress. Thus, the protective effects of trehalose were more remarkable in the sensitive strain. The activities of intracellular trehalose metabolic enzymes, i.e., trehalose-6-phosphate synthase, trehalose phosphorylase and neutral trehalase, were determined, and our data indicated that the changes of these enzymes activities in the sensitive strain were more beneficial to accumulate trehalose than that in the less-sensitive strain.

Amino Acid Supplementations Enhance the Stress Resistance and Fermentation Performance of Lager Yeast During High Gravity Fermentation.

The effects of different wort gravity or ethanol concentration in initial wort on the fermentation performance of lager yeast and assimilation of free amino acids (FAAs) were studied. Results showed that compared with high wort gravity (24°P), high ethanol concentration (10%, v/v) decreased yeast growth, cell viability, and wort fermentability significantly. The assimilation of FAAs was changed dramatically by high ethanol toxicity, and positive correlations between the assimilation amounts of 10 FAAs (Asp, Ser, Gly, Arg, Tyr, Val, Met, Lys, Ile, and Leu) and fermentation performance (cell viability, fermentability, and ethanol production) were identified, especially for Arg and Lys exhibiting extremely significant positive correlations. Furthermore, confirmatory testing was carried out by supplementing 24°P worts with 10 FAAs of 0.5, 1, and 2 times of their standard concentrations, respectively. Results exhibited that 10 FAA supplementations improved physiological characteristics and fermentation performance of lager yeast significantly, especially for 1 times FAA supplementation increasing wort fermentability and ethanol yield by 6 and 17%, respectively, and upregulated the expression level of HSP12 and increased more intracellular trehalose accumulation in yeast cells, indicating that stronger protective function was stimulated in yeast cells. Therefore, it was suggested that these 10 FAAs could regulate yeast cells to adapt to high gravity environmental stresses.

Genomic Understanding of an Infectious Brain Disease from the Desert.

Rhinocladiella mackenziei accounts for the majority of fungal brain infections in the Middle East, and is restricted to the arid climate zone between Saudi Arabia and Pakistan. Neurotropic dissemination caused by this fungus has been reported in immunocompromised, but also immunocompetent individuals. If untreated, the infection is fatal. Outside of humans, the environmental niche of R. mackenziei is unknown, and the fungus has been only cultured from brain biopsies. In this paper, we describe the whole-genome resequencing of two R. mackenziei strains from patients in Saudi Arabia and Qatar. We assessed intraspecies variation and genetic signatures to uncover the genomic basis of the pathogenesis, and potential niche adaptations. We found that the duplicated genes (paralogs) are more susceptible to accumulating significant mutations. Comparative genomics with other filamentous ascomycetes revealed a diverse arsenal of genes likely engaged in pathogenicity, such as the degradation of aromatic compounds and iron acquisition. In addition, intracellular accumulation of trehalose and choline suggests possible adaptations to the conditions of an arid climate region. Specifically, protein family contractions were found, including short-chain dehydrogenase/reductase SDR, the cytochrome P450 (CYP) (E-class), and the G-protein β WD-40 repeat. Gene composition and metabolic potential indicate extremotolerance and hydrocarbon assimilation, suggesting a possible environmental habitat of oil-polluted desert soil.

Protective role of trehalose during radiation and heavy metal stress in Aureobasidium subglaciale F134

An isolated black yeast-like strain was obtained from radiation-polluted soil collected from Xinjiang province in northwest China. On the basis of ITS and LSU rDNA sequence analysis, in combination with the colony morphology and phenotypic properties, the isolated strain was revealed to represent a novel variety of Aureobasidium subglaciale, designated as A. subglaciale F134. Compared to other yeasts and bacteria, this isolate displayed superior resistance to gamma irradiation, UV light, and heavy metal ions. It was discovered that the resistance of the isolate was correlated with the stress protector trehalose. Through the overexpression of the trehalose-6-phosphate synthase gene tps1 and the deletion of acid trehalase gene ath1, the APT∆A double mutant exhibited a survival rate of 1% under 20 kGy of gamma-radiation, 2% survival rate at a UV dosage of 250 J/m2, and tolerance towards Pb2+ as high as 1500 mg/L, which was in agreement with the high accumulation of intracellular trehalose compared to the wild-type strain. Finally, the protective effects and the mechanism of trehalose accumulation in A. subglaciale F134 were investigated, revealing a significant activation of the expression of many of the stress tolerance genes, offering new perspectives on the adaptations of radioresistant microorganisms.

Improvement of Lead Tolerance of Saccharomyces cerevisiae by Random Mutagenesis of Transcription Regulator SPT3.

Bioremediation of heavy metal pollution with biomaterials such as bacteria and fungi usually suffer from limitations because of microbial sensitivity to high concentration of heavy metals. Herein, we adopted a novel random mutagenesis technique called RAISE to manipulate the transcription regulator SPT3 of Saccharomyces cerevisiae to improve cell lead tolerance. The best strain Mutant VI was selected from the random mutagenesis libraries on account of the growth performance, with higher specific growth rate than the control strain (0.068 vs. 0.040h-1) at lead concentration as high as 1.8g/L. Combined with the transcriptome analysis of S. cerevisiae, expressing the SPT3 protein was performed to make better sense of the global regulatory effects of SPT3. The data analysis revealed that 57 of S. cerevisiae genes were induced and 113 genes were suppressed, ranging from those for trehalose synthesis, carbon metabolism, and nucleotide synthesis to lead resistance. Especially, the accumulation of intracellular trehalose in S. cerevisiae under certain conditions of stress is considered important to lead resistance. The above results represented that SPT3 was acted as global transcription regulator in the exponential phase of strain and accordingly improved heavy metal tolerance in the heterologous host S. cerevisiae. The present study provides a route to complex phenotypes that are not readily accessible by traditional methods.

Arsenic inorganic compounds cause oxidative stress mediated by the transcription factor PHO4 in Candida albicans

Arsenic is a toxic metalloid widespread in nature. Recently, it has been demonstrated a main role of the transcription factor Pho4 in the acquisition of tolerance to arsenic-derived compounds, arsenite and arsenate in Candida albicans. Here, the effect of these compounds on this pathogenic yeast has been analyzed. In wild type cells, both arsenite and arsenate induced a marked increase in the endogenous production of Reactive Oxygen Species (ROS), together with the accumulation of intracellular trehalose and the activation of catalase, suggesting their role as generators of oxidative stress in this yeast. However, a pho4 null mutant showed a minor increase of intracellular ROS and a different kinetics of catalase activation upon exposure to arsenite and arsenate. Interestingly, the enzymatic activity of glutathione reductase and superoxide dismutase were exclusively triggered by arsenite but not by arsenate. pho4 mutant cells were also found to be sensitive to azide but significantly resistant to arsenate through a process dependent on an active electron transport chain and the alternative oxidase system. Therefore, arsenic-derived compounds induce a strong antioxidant response in C.albicans via different mechanisms.