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Systems Microbiology and Biomanufacturing https://doi.org/10.1007/s43393-021-00058-4

REVIEW

Overview of yeast environmental stress response pathways and the development of tolerant yeasts

Nai‑Xin Lin1,2 · Yan Xu1,2 · Xiao‑Wei Yu1,2

Received: 19 July 2021 / Revised: 1 September 2021 / Accepted: 2 September 2021 © Jiangnan University 2021

AbstractYeast is widely used for industrial production of various types of products, such as ethanol and enzymes. However, its fer‑mentation efficiency is strongly reduced by harmful environmental stresses. Specifically, harmful environmental stresses damage important cellular components, such as cell wall, cell membrane, proteins, etc. Then, these damages cause cellular metabolic disorders or even death. In the past decades, there has been a portfolio of studies on the environmental stress tol‑erance of yeasts, which mainly aimed at cell damages caused by different environmental stresses, different ways to improve yeast environmental stress tolerance or a tolerance mechanism for certain environmental stress. However, a comprehensive overview of how yeasts respond to environmental stresses is lacking, and the correlation of tolerance mechanism between different environmental stresses is unclear. In this review, we summarized the general damages induced by most of envi‑ronmental stresses, the existing major mechanisms of environmental stress tolerance from the perspective of key signalling pathways, and the common ways to improve the resistance to environmental stresses in yeast cells. The tolerance mechanisms of yeast cells to different environmental stresses are diverse, but sometimes they share the same signalling pathway. Cells use sensors on the cell surface to recognize environmental stresses and transmit signals to the nucleus to cause changes in gene expression. By summarizing the main signalling pathways, including MAPK pathway, cAMP/PKA pathway, YAP1/SKN7 pathway, it will provide a powerful reference for future efforts to promote yeast environmental stress tolerance and study yeast tolerance mechanisms.

Keywords Yeast · Environmental stress · Signalling pathway · Protein aggregation

Introduction

As an important cell factory, yeast has been widely used for industrial fermentation [1], but the fermentation efficiency of yeast is severely affected by the fermentation environ‑ment. During the fermentation process, yeast is exposed to various environmental stresses including high temperature, high osmolarity, acid, oxidative stress and so on. The dam‑age in yeast cells under environmental stresses is depicted in Fig. 1. Environmental stresses mainly cause damages to

cellular macromolecules, then induce protein denaturation, generate abnormal proteins, accumulate toxic proteins [2]. The accumulation of denatured proteins can compose pro‑tein aggregation, then induce endoplasmic reticulum (ER) stress to influence normal protein production. Furthermore, the integrity of cell wall can be damaged by environmental stresses. The environmental stresses also might directly or indirectly cause cell membrane disorder and degrade vacu‑ole membrane [3]. In addition, environmental stresses can impose DNA lesions and even induce lethal DNA damages [4, 5].

Environmental stresses cause damages to yeast cells in different ways. Specifically, osmolarity stress leads to yeast cells shrinking and forming improper cell volume then causes a retard of cellular processes in yeast cells [6–8]. Acidity stress unexpectedly improves cellular anions level, degrades intracellular vacuoles, inhibits cell growth, and influences the formation of product [9, 10]. Under oxidative stress, excessive ROS production induces the peroxidation

* Xiao‑Wei Yu yuxw@jiangnan.edu.cn

1 Lab of Brewing Microbiology and Applied Enzymology, School of Biotechnology, Jiangnan University, Wuxi 214122, China

2 Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China

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of several essential components, such as protein, lipid, and nucleic acids [11]. High ambient temperature, especially during summer or in tropical countries, causes protein to improperly aggregates and reduces cellular enzymes activity, and it will be costly to maintain normal fermentation [12].

The mechanisms that yeast cells respond to environmen‑tal stresses have been deeply explored and summarized in reviews. Specifically, responses of yeast cells to multiple environmental stresses were analysed from the perspective of transcription factors and functional compounds [13–15]. With regard to high temperature, how the cells respond to heat stress is a matter of great concern to researchers [16]. However, heat stress is also the main source of oxidative stress [17]. Oxidative stress mainly induces the accumula‑tion of harmful reactive oxygen species (ROS), so the source of ROS and cell response have received much attention [18]. The ability of yeast cells to adapt to a high osmolarity envi‑ronment is of great significance for alcoholic beverages and biofuels industries. The high osmolarity glycerol (HOG) pathway plays an important role in hyperosmotic stress tol‑erance [7, 19]. Yeast plays an important role in food indus‑try may have to overcome acid stress, the remodelling of cell wall and cell membrane structure, regulated by tran‑scription factors especially Haa1 [20]. The production of a fuel alcohol or brewing industry rely on yeast, so ethanol tolerance has received much attention. A comprehensive review of ethanol stress tolerance mechanism is given that

the cell membrane fluidity and protein folding, membrane lipid composition play important roles in resistance to etha‑nol [21–23]. How S. cerevisiae responds to toxic aldehyde compounds has been well summarized [24]. Even an in‑depth review of the methods using global regulators and cell surface properties to enhance stress tolerance is given by Kuroda et al. [25].

The response of yeasts to environmental stresses includes a complex system of signalling pathways. These signalling pathways allow the recognising of the external environmen‑tal changes and the signalling from the cell surface to the nucleus. Then, the environmental stress signals are trans‑formed into alterations in gene expression. The functional factors synthesized under the control of these signalling pathways allow damaged cells to recover and avoid further damage. Summarizing the signalling pathways of yeast cells in response to environmental stresses is conducive to a full understanding of the cellular environmental stress response mechanism, and facilitates the subsequent rational improve‑ment of the environmental stress tolerance of yeast cells.

At present, the mechanism of yeast environmental stress tolerance is still a long‑standing puzzle. It is known that the response of yeast cells to environmental stresses is con‑trolled by transcription regulators (TFs) [26]. Specifically, transcription factors Msn2/4 play dominating roles in vari‑able environmental stresses, such as osmotic stress, thermal stress, and high ethanol concentration [15]. Transcription

Fig. 1 The stress‑induced dam‑ages and stress response sche‑matic of yeasts. Environmental stresses cause cell wall damage, make plasma membrane dis‑order and aggregate denatured proteins, increase ROS level and lead to endoplasmic reticulum (ER) stress. In response to these damages, the ESR process is initiated, membrane stress sen‑sors first pass the stress signals to stimulate key signalling path‑ways and contributing factor biosynthesis pathways. With the cooperation of these functional pathways, yeasts try to remain stable

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factors Hsf1 and Yap1 specifically mediate heat shock response and oxidative stress response, respectively [26, 27]. Many researchers have used omics analysis to study the tol‑erance mechanism of yeasts that are resistant to environmen‑tal stresses [28, 29]. By comparing the expression patterns of environmental stress‑tolerant strains and environmental stress‑sensitive strains, functional signalling pathways were identified. These results indicate that the tolerance mecha‑nisms of different environmental stresses are diverse, but sometimes they share the same signalling pathway, which makes the mechanisms of environmental stress tolerance more complex. Therefore, this review aims to summarize key signalling pathways and attempts to figure out the main pathways that are resistant to various environmental stresses and the interactions between these pathways, which will provide a fundamental summary for future studies on yeast environmental stress tolerance.

Intracellular accumulation of denatured proteins and reactive oxygen species (ROS) that can be induced by most environmental stresses

Yeast cells can be provoked by environmental stresses dur‑ing fermentation, which can cause the formation of dena‑tured proteins. More seriously, misfolded proteins are prone to aggregate, leading to cell damage and affecting fermenta‑tion efficiency. Since all secreted and membrane proteins are synthesized and begin the process of maturation in the ER, the accumulation of misfolded proteins in the ER may induce ER stress. Then ER stress activates the unfolded protein response (UPR), endoplasmic reticulum‑associated degradation (ERAD) pathway and stimulates the assembly

of the pre‑autophagosome structure, and triggers autophagy [30–32]. Maintaining protein homeostasis involves the deg‑radation of harmful denatured proteins and the refolding of misfolded and damaged proteins [33] (Fig. 2). Induction of rapid function of multiple factors, including heat shock proteins (HSPs), ubiquitin–proteasome system (UPS), and autophagy act to maintain intracellular homeostasis in threatened yeast cells (Fig. 2). Different sizes of HSPs play different roles in yeast against environmental stress which have been well summarized in other reviews [34, 35]. Rapid clearance of denatured proteins can be achieved by those efficient functional factors. Because of the similar function, increased ubiquitin‑dependent degradation can substitute the vital requirement for HSP induction [36].

In general, environmental stresses may cause the accu‑mulation of excess ROS in cells (Fig. 1). Forrester et al. [37] well summarized the ROS production in compartments, such as the cytoplasm, mitochondria, peroxisome, and endo‑plasmic reticulum, in mammalian cells. The ROS produc‑tion sites summarized in that review are also applicable to yeast. Deregulated ROS generation results in DNA damage, even cause cells the induction of apoptosis in yeast cells. These two main consequences, denatured proteins, and ROS, caused by environmental stress tightly interact. ER stress is prolonged or acute, cell death may occur, accompanied by the generation of mitochondrial ROS [3]. In turn, ROS induces the production of misfolded proteins leading to ER stress [3].

Heat shock response (HSR) and oxidative stress response (OSR) are two main cellular responses against environmen‑tal stresses in yeasts. The HSR and OSR in S. cerevisiae have been well summarized in [38]. HSR mainly regulates the biosynthesis of HSPs to eliminate stress‑induced dena‑tured proteins. OSR mainly mediates the biosynthesis of

Fig. 2 Disposal of misfolded proteins and protein aggre‑gates. Intracellular proteins are affected by environmental stresses and become abnormal proteins and can be directly restored to native proteins by the help of chaperone proteins, such as heat shock proteins and trehalose. However, too many denatured proteins will form protein aggregates, which are toxic to yeast cells and need to be degraded opportunely to maintain cell homeostasis. This process is achieved mainly by trehalose, UPS, and autophagy

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antioxidants to scavenge cellular ROS and eliminate oxida‑tive stress. Especially, in our previous work, we found that high temperature induced OSR and HSR simultaneously in a heat‑sensitive Pichia pastoris strain [39].

Signalling pathways associated with environmental stress tolerance of yeast

An adverse fermentation environment may disturb yeast cells, thereby activating signalling pathways to maintain cell homeostasis. Signalling pathways including MAPK pathway [40], cAMP/PKA pathway [41], YAP1/SKN7 pathway [42], were depicted in Fig. 3. The MAPK pathways have three parts including the HOG pathway, the CWI pathway, and the Fus3/Kss1 pathway [43]. Other than the Fus3/Kss1 pathway, the HOG pathway and the CWI pathway are responsible for the response of yeast to environmental stresses [44]. The HOG pathway [45] rapidly regulates the synthesis of glyc‑erol to maintain cell osmotic pressure balance, while the CWI pathway responds to repair cell wall. The cAMP/PKA pathway, as an upstream regulatory pathway, can negatively regulate the transcript factor Msn2/4 in the environmental stress response (ESR) mechanism [46]. The YAP1/SKN7 pathway is primarily responsible for regulating intracellular oxidative stress responses.

Intriguingly, through intricate hierarchical regulatory net‑works, yeast can quickly respond to environmental changes (Fig. 3). These pathways dramatically interact with each other. Specifically, the sensors of the CWI pathway sense

environmental stresses and repress the cAMP/PKA pathway simultaneously [47]. Nevertheless, the cAMP/PKA pathway negatively regulates the YAP1/SKN7 pathway [48]. Rho1 as the upstream regulator of the CWI pathway can control YAP1/SKN7 pathway and then help yeast cells resist oxi‑dative stress [49]. Furthermore, Ypd1 in the HOG pathway can also regulate the YAP1/SKN7 pathway that Ypd1 trans‑fers its phosphate to aspartyl residues within the receiver domains Skn7, which activates Skn7 [50]. The functional factors involved in signalling pathways that respond to envi‑ronmental stresses were depicted in Table 1.

HOG pathway

Environmental stresses (e.g., high osmolarity), induce plasma membrane stretch and then activate the HOG path‑way [50]. The HOG pathway is composed of membrane‑associated sensors [59], cytoplasmic and nuclear effec‑tors [45] (Table 2). Furthermore, any single osmosensor is enough to improve osmoadaptation [59]. A comprehensive review of the HOG pathway is given in Tong et al. [43]. The Hog1 shuttles between the cytoplasm and nucleus to activate the transcription factors Msn2/4 [60], then acti‑vate the expression of glycerol phosphate dehydrogenase 1 Gpd1, leading to glycerol accumulation [61]. Then the accumulation of glycerol rapidly balances the intracellular and extracellular osmotic pressure. Conversely, knocking out the Hog1 gene causes yeast cells to be sensitive to NaCl, sorbitol, and high temperatures [62]. Cells often suffer from multiple environmental stresses at the same

Fig. 3 Cross‑talk relationship in major signalling pathways, including positive (the arrows) and negative (the T‑lines) regu‑lations. The different pathways were highlighted by different background colours

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time. For example, ethanol may cause high osmotic stress in yeast cells and enhances the production of ROS in mitochondria simultaneously [63]. In addition, the acti‑vation of the HOG pathway may induce other protection mechanisms. For instance, Hog1 plays an important role in acid stress by activating the antioxidant defence system [54]. Msn2/4 mediating environmental stress responses are crucial TFs downstream of the HOG pathway. The HOG pathway regulates the synthesis of glycerol, which plays an essential role in maintaining an osmotic equilibrium with the external environment.

By adjusting the percentage of membrane compounds, yeasts effectively improve environmental stress tolerance. Sterols are important membrane compounds of yeast cells. Under the hyperosmolarity condition, yeast cells consid‑erably lower their sterol by involving the HOG pathway [67]. The Hog1 activates the transcription repressor Mot3 and Rox1 to shut down transcription of sterol biosynthesis genes ERG2 and ERG11 under osmotic stress. In addition, by adjusting the proportion of the sterol compositions, the thermotolerance of yeasts may be enhanced [68].

In view of the role of the HOG pathway under osmotic stress, its main regulator Ypd1 is localized and expressed in the plasma membrane and nucleus to effectively improve the signal transduction efficiency of this pathway [69]. The activation of the HOG pathway under high osmotic stress is conducive to cell growth, while the activation of the HOG pathway under non‑stress conditions may cause growth defects on yeast cells [70]. Hence, a robust network structure of the Sln1‑Ypd1‑Ssk1 three‑component phospho‑relay with a large excess pool of Ypd1 avoids unexpected activation of the HOG pathway in S. cerevisiae [70].

CWI pathway

Morphological and structural properties of the cell wall are important factors affecting the yeast environmental stress tolerance. The expression of cell wall biogenesis and remod‑elling genes is mediated by the CWI pathway [51]. When the cell is challenged by environmental stresses, the CWI path‑way is activated [71]. Commonly, environmental stresses cause cell wall damage including high temperature, etha‑nol stress [51], and toxic chemical compounds. The envi‑ronmental stress signal is systematically conveyed to Rlm1 transcription factor, then the cell wall remodelling starts. Cell wall remodelling includes changes in the proportion of cell wall compositions and changes in cross‑linking in these compositions [71].

When the yeast cells are subjected to certain environmen‑tal stresses, their cell wall components also change. Upon ethanol stress, the expression of cell wall‑remodelling pro‑teins including β‑1,3‑glucan synthase, chitin trans‑glycos‑ylase, and O‑glycosylated cell wall protein is induced by the CWI pathway [51]. Schiavone et al. [72] found that heat shock can cause an increase in chitin content, a decrease in β‑1,3‑glucan content, and an increase in β‑1,6‑glucan content. Conversely, other researchers have found that the putative glycosylphosphatidylinositol‑linked aspartyl pro‑tease gene mutant of P. pastoris shows an increased osmotic tolerance, while the chitin content in the cell wall is reduced, the β‑1,3‑glucan content is increased, and the inner cell wall is thicker [73]. Interestingly, the phenomenon that the inner cell wall thickens happened in a lager yeast which exhib‑ited a multiple stress tolerance [74]. Fascinatingly, in our research on P. pastoris, it was also found that the environ‑mental stress tolerance was increased, and the inner cell wall became thicker [75]. Interestingly, the CWI pathway is not the only way to maintain cell wall integrity. The HOG pathway and the invasive growth pathway are also required to cope with the cell wall damage depending on the nature of the environmental stresses [51, 76]. It is noteworthy that Cdc19 encoding pyruvate kinase plays a crucial role in the viability of yeast at high temperatures [77]. A change in pyruvate kinase activity may influence glycolytic flux which triggers an increase in ATP metabolism. Afterward, the pro‑cess of cell wall remodelling can be accelerated. In addition,

Table 1 Pathways respond to environmental stresses

Pathways Environmental stress types Functional factor References

CWI pathway Cell wall perturbing stress, oxidative stress, high osmo‑larity, heat stress

Ubiquitin, glucan, chitin [51–53]

HOG pathway Osmotic stress, weak acid stress, DNA damage agent Glycerol, ubiquitin, lipids, fatty acids [54, 55]cAMP/PKA pathway Oxidative stress, nascent protein misfolding stress Thioredoxin, trehalose [56, 57]YAP1/SKN7 pathway Oxidative stress SOD1, SOD2, peroxiredoxin, glutathione [58]

Table 2 Functional protein in the HOG pathway

Protein Description References

Sln1 Osmosensor histidine kinase [64]Ypd1 Histidine phosphotransfer protein [65]Sho1 Tetraspanning membrane protein [59]Opy2 Membrane protein scaffold [66]Hkr1 Functionally redundant osmosensors [59]Msb2 Functionally redundant osmosensors [59]

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the yeast O‑Man glycoproteome plays a major role in main‑taining yeast cell wall integrity by activating the CWI path‑way [78, 79].

The Rho1/Pkc1 pathway is the upstream regulating path‑way of MAPK pathway [80], blocks polarized cell growth, and repairs the environmental stress‑induced wounds by activating the CWI pathways [81]. Rho1 targets effec‑tors including Pkc1, Skn7, and the glucan synthase which directly impact the biogenesis of cell wall [50]. Pkc1 includ‑ing phospholipid‑dependent protein kinases and serine/threonine‑specific protein kinases, responds to extracellular signals [80]. Loss of Pkc1 function may cause cell lysis at elevated temperatures due to a deficiency in cell wall con‑struction that both the inner and outer layers of cell wall are thinner [50].

To modulate protein homeostasis, ubiquitin–proteasome system (UPS) is one of the critical pathways for protein deg‑radation in eukaryotic cells [82]. Similarly, the phospho‑rylation–de‑phosphorylation of Ubc1 encoding ubiquitin‑conjugating enzymes was necessary for thermotolerance in the absence of either the HOG or CWI pathways [83], which means the HOG pathway and the CWI pathway have overlap functions with the UPS. In contradiction of anticipations, the UPS is not capable of compensating for a deficiency in autophagy [84]. Interestingly, both ubiquitin and HSPs are responsible for maintaining protein homeostasis. However, ubiquitin and HSPs have an obscure connection. As ubiq‑uitin expression increases, the demand for HSP decreases, resulting in restoration in HSP synthesis [36].

Overexpression of Pkc1 activated CWI pathway that expression levels of Slt2 and Rlm1 upregulated [85, 86]. Overexpression of a hyperactive allele of MKK1 can also induce the activation of CWI pathway by increasing the expression of Slt2 and Rlm1 [85, 87]. Rlm1 mediates posi‑tive autoregulatory transcriptional feedback by binding at the promoter of Slt2 and Rlm1 [85]. Therefore, overexpression of Rlm1 may mediate the activation of CWI pathway.

cAMP/PKA pathway

In yeast, the global regulators cAMP and PKA play central roles in metabolism regulation, environmental stress resist‑ance, and cell cycle progression [88]. It targets transcrip‑tion factors Msn2/4 [89], which govern vital stress response signalling pathways. Ras2p is a small GTP binding protein and homologous to the mammalian Ras protooncogenes [90]. Cdc25 encodes a membrane‑bound guanine nucleotide exchange factor, which activates Ras1 and Ras2, is a major regulator of cAMP signalling [91]. The cAMP negatively regulates the Msn2/4 [46]. When the level of intracellular cAMP is low, the Msn2/4 can be activated, and then yeast cells are able to fight against heat stress [46]. Adenylate cyclase Cyr1 synthesizes cAMP, then regulates the cAMP/

PKA pathway [88, 92]. It is worth mentioning that cell sen‑sor Wsc1 activates the CWI pathway and parallel inhibits the PKA signalling [71].

The PKA signalling pathway is of vital significance in response to environmental stresses. In yeast cells, thiore‑doxins partner with peroxiredoxin in H2O2 signalling [93]. Oxidized thioredoxins constrain the PKA pathway, in part through inhibiting the nuclear holding of the PKA cata‑lytic subunits, allowing Msn2 to converge in the nucleus [93]. Negative feedback regulation periodically renovates PKA activity, causing Msn2 to exit the nucleus [93]. Upon calcium stress, lacking the function of PKA resulted in an elevated expression of transcriptional regulator Prz1 which causes a CaCl2‑sensitive phenotype [94]. Inhibiting cAMP/PKA helps yeast resist environmental stresses. The major cytosolic peroxiredoxin, Tsa1 is required for both improv‑ing tolerance to H2O2 and prolonging lifespan under caloric restriction by inhibiting the cAMP/PKA pathway [95].

YAP1/SKN7 pathway

The YAP1/SKN7 pathway mainly involves the regulation of antioxidant genes to oxidative stress [42]. Moreover, Yap1 and Skn7 as significant ROS response signalling factors also have been proved to participate in DNA double‑strand break repair [27]. The YAP1/SKN7 pathway could be provoked by environmental stresses causing ER stress, but in the absence of oxygen, the damages will be repaired [96]. When Yap1 gene was knockout, cells failed to survive under oxidative stress [96]. Overexpression of Yap1 enhances the resistance of yeast cells to diverse toxic compounds [97]. Moreover, overexpression of Yap1 enhanced resistance to lignocellu‑lose‑derived fermentation inhibitors in S. cerevisiae [98]. Most YAP1‑regulated genes are classified in the numer‑ous functional categories of redox metabolism, amino acid metabolism, stress response, DNA repair, and co‑regulate with chaperones [99, 100].

As an important regulatory factor, Skn7 regulates multi‑functional responses in fungi and other organisms, and assists cells to resist oxidative stress and maintain cell wall integrity in yeast [101]. Evidence proved that the mutants lacking Skn7 were sensitive to H2O2 [63]. Besides, Skn7 is also governed by Rho1, sensor Sln1, and response factor Ypd1 [102], which then plays an important role in cell wall biogenesis when yeast cells undergo hypo‑osmolarity stress [50].

Environmental stresses always induce the generation of cellular ROS and then cause oxidative stress on cells. The antioxidant defence system plays a very important role in cell response to environmental stresses. Antioxidant GSH is the most abundant thiol and serves as a protectant in eukaryotic cells in redox unbalanced conditions [103]. Yap gene family members mediate numerous genes in a

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wide range of functional categories [100]. Interestingly, the concentration of GSH is found to be higher in Yap1 over‑expression yeast [104], revealing that GSH is one of the downstream factors of Yap gene family members. In addition, peroxiredoxin is one of the activators of the oxidant‑sensing transcription factor Yap1 and other stress‑responsive transcription factors, such as Msn2 and Skn7 [105, 106]. The maintenance of the thioredoxin system is of great significance that too many oxidized thioredoxins will cause UPR and then diminish cell growth rate [96].

SODs are responsible for reducing the cellular ROS level. Specifically, when superoxide anions (O2

·−) are generated as coproducts of the mitochondrial electron transport chain, they are changed into hydrogen perox‑ide (H2O2) by SODs [11]. Overexpression of SOD1 in P. pastoris reduced the ROS level caused by methanol expo‑sure [107]. H2O2 is then transformed into water and oxy‑gen by catalase, glutathione peroxidase, and thioredoxin peroxidase, to prevent the formation of oxidant hydroxyl radicals (∙OH) [54]. Previously, the Δsod1 mutant lacking cytosolic Cu/Zn‑SOD has been shown to have a higher O2

·− level under various environmental stresses conditions [63]. Additionally, the repression of SOD1 and SOD2 causes yeast cells sensitive to ethanol and heat stress [58]. Moreover, SOD2 activates oxidative response regulator Yap1 [58], so when SOD1 and SOD2 are knocked out, the ability to survive under oxidative impulse is severely suppressed [103]. In addition, the inactivation of the Yap1 cancelled the activation of SOD and catalase [108].

Evident correlation among functional factors

Many cellular functional compounds are helpful to the envi‑ronmental stress tolerance of yeast. In addition to the end‑products of the signalling pathways, such as glycerol, cell wall components, and YAP1 pathway‑regulated antioxidants (Table 1). The integrity of cell membranes has also attracted much attention. For example, Yang et al. [109] found that S. cerevisiae strain with increasing content of unsaturated fatty acid (UFA) and cell membrane fluidity can tolerate high concentrations (up to 25 vol%) of ethanol. Researchers have found that the increasing content of some amino acids helps yeast to resist environmental stresses. For instance, Wang et al. [110] proved that overexpressing key gene PRO1 encoding proline improved the tolerance of S. cerevisiae to furfural, acetic acid, and phenol stress. Trehalose is a vital functional compound being proved to help yeasts to resist various environmental stresses, such as desiccation [111], freezing [112], heat stress [113], etc. The pentose phosphate pathway [114] is the main source of NADPH, an important cofactor for cellular ROS elimination.

These compounds can be biosynthesized in yeast cells. Generally, the increase in concentration of one functional compound might correspondingly reduce the synthesis of the other compound with a similar function, unless the sole improvement of one compound is not enough to meet the demand for stress resistance. Interestingly, we found that the synthesis of these compounds is interrelated (Fig. 4).

Fig. 4 The correlation of the biosynthesis process in factors involved in different functions. One of the functional factors in yeast cells may be involved in the synthesis of another functional factor. According to the protective mechanism of functional factors, researchers can specifically protect yeast cells from stress damage by coordinating the synthesis of these functional factors

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Understanding the synthesis of stress‑resistant compounds in yeast cells can help improve the stress tolerance of yeast through a combination of genetic engineering and metabolic engineering.

How to improve the environmental stress tolerance of yeasts?

Resistance to environmental stresses varies from strain to strain, also relates to the certain type of environmen‑tal stresses and the duration of exposure to environmental stresses conditions. To improve yeast fermentation ability, several methods have been previously employed to improve the environmental stress tolerance of yeasts (Table 3). Since environmental stress tolerance mechanism is still obscure, the majority of the methods depend on random mutagenesis, such as adaptive laboratory evolution [115], UV mutagenesis [116] (Table 3). A new genome shuffling technique with CRISPR system was used to realize environmental stress tolerance by genome evolution under environmental stress conditions [117]. Increasing the ploidy of yeast is also an option for researchers to improve yeast tolerance [118].

Regarding rational methods, the principle of improving cell tolerance mainly relies on the existing understanding of the mechanism of yeast cell tolerance. Therefore, rational mutation of key genes and increasing the content of func‑tional factors are usually used as effective means to improve

the environmental stress tolerance of yeasts (Table 3). Based on the understanding of the yeast stress response mecha‑nism, the researchers might use many rational methods to improve the stress tolerance of yeast. Regrettably, strains exhibiting multi‑tolerance simultaneously have not been developed yet. In addition to the way cells are modified based on the cellular environmental stress response mecha‑nism, the direct addition of functional compounds through exogenous sources can also improve the strain’s environmen‑tal stress tolerance to a certain extent.

During the fermentation process, yeast cells inevitably encounter many environmental stresses that seriously affect the fermentation efficiency. Improving the environmental stress tolerance of yeasts is beneficial to the smooth comple‑tion of the fermentation. In ethanol fermentation, the yeast cells are exposed to high concentrations of ethanol, which are toxic and hinder the further fermentation process of yeast cells [109]. Irrational strain breeding methods are very com‑mon that a superior S. cerevisiae strain F23 was acquired using strategies of ethanol domestication, UV mutagenesis, and protoplast fusion [109]. Thanks to the ethanol tolerant yeast obtained irrationally, researchers can study its toler‑ance mechanism through omics methods, etc. Understanding the environmental stress tolerance mechanism of yeast is one of the foundations for rationally constructing tolerant yeast. By validating the function of OLE1 in the variability of membrane fluidity in strain F23, OLE1‑overexpressed transformant successfully resists ethanol stress [109]. In

Table 3 Methods to improve the environmental stress tolerance of yeasts

Environmental stresses Irrational methods Rational methods

Yeast species Technique variation Refs. Yeast species Technique variation Refs.

Acid S. cerevisiae Serial microaerobic batch cultivation

[120] S. cerevisiae Deletion of key gene OPI1 and improve the biosynthesis of phosphatidylcholine

[121]

S. cerevisiae Adaptive laboratory evolution [122] S. cerevisiae Overexpression of RCK1 coding for a protein kinase involved in reducing ROS level

[123]

Oxidative S. cerevisiae Global transcription machin‑ery engineering

[124] Yarrowia lipolytica Upregulating oxidative stress defense pathways

[125]

Hyperosmotic Saccharomy-ces pastori-anus

Ethyl methanesulfonate mutagenesis

[126] Candida glycerinogenes Improving intracellular gamma‑aminobutyric acid accumulation

[127]

High temperature S. cerevisiae Spore‑to‑cell hybridization technique

[128] S. cerevisiae Rational synthetic combina‑tion with HSP and SOD genetic devices

[129]

P. pastoris UV mutagensis [130] S. cerevisiae Strengthen the antioxidant defense system

[131]

Ethanol S. cerevisiae Metabolic engineering and genome shuffling

[132] S. cerevisiae Multilevel defense system [133]

Lager yeast Interspecific hybridization [134] S. cerevisiae Coordination of the CWI and HOG pathways

[51]

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addition, fermentation inhibitors involved in lignocellulose hydrolysates are unavoidable problems to produce biofu‑els and biochemicals by yeast cells. Kim et al. constructed robust S. cerevisiae with improved furan derivatives and ace‑tic acid tolerance through modulation of spermidine contents by metabolic engineering method [119].

Conclusion and perspectives

Concerning sustainable development, environmental stress‑tolerant yeasts are extremely vital for industrial fermenta‑tion. Yeast cells may encounter multiple environmental stresses at the same time. Mechanisms required for one environmental stress tolerance may also be responsible for other environmental stress tolerance. At present, a lot of articles have well summarized the damages caused by environmental stresses. However, the understanding of the tactics that yeast cells respond to environmental stresses is still incomplete. We summarized from the perspectives of signalling pathways and their cross‑talk mechanism to figure out the cell response in yeasts under environmental stresses. Additionally, we summarized the functional compounds and their inter‑correlations in response to environmental stresses. This review provided a general awareness and understand‑ing of how yeast cells respond to environmental stresses. Besides, it has initially sorted out the associations between the various environmental stress tolerance mechanisms and the associations between functional factors in yeast. It pro‑vides a reference for improving the environmental stress tolerance of yeast from the perspective of overall regulation.

Based on the summary of the mechanisms of environ‑mental stress tolerance in yeast, we would like to give some advice to other researchers. When studying several environ‑mental stresses simultaneously, you can start with the cell wall integrity pathway and changes in cell membrane com‑positions. Also, HSPs and UPS can be the focus of research since almost all kinds of environmental stresses can lead to misfolding and aggregation of intracellular proteins. When studying osmotic tolerance, attention can be paid to the syn‑thesis of the HOG pathway and the accumulation of intra‑cellular glycerol. Under the conditions of oxidative stress, attention should be paid to the YAP1/SKN7 pathway and the antioxidant defence system. In addition to focusing on intracellular trehalose content, studies on the mechanism of thermotolerance in yeast can refer to the anti‑oxidative stress mechanism in most cases. The cellular level of amino acid can be changed to acquire an environmental stress‑tolerant yeast.

The environmental stress tolerance mechanism of yeast is mainly based on two aspects: cells resist damages caused by environmental stresses and cell self‑repairing after being damaged. This understanding provides ideas for rational

transformation to improve yeast stress tolerance. Through the continuous understanding of the cellular environmen‑tal stress tolerance mechanism, more and more key genes and key pathways have been unearthed. With the develop‑ment of gene editing technology, more and more research‑ers use CRISPR system for genome editing in yeast. Lian et al. [135] described a combinatorial metabolic engineering strategy that combines transcriptional activation, transcrip‑tional interference, and gene deletion in S. cerevisiae. In the future, through CRISPR system, it may be possible to simul‑taneously knock out unfavourable genes, activate favourable genes, and inhibit genes suitable for low‑dose expression in yeast to achieve the environmental stress tolerance of yeast.

Despite several signalling pathways that promote environ‑mental stress tolerance in yeasts have been demonstrated, the inter‑relationship of these pathways is not clear enough, and there may be some key links that have not been found. In the future, researches on the environmental stress tolerance of yeasts need to be carried out in terms of finding more func‑tional pathways and modifying existing metabolic pathways, then trying to resolve the completed environmental stress tolerance mechanism.

Acknowledgements This work is supported by the National Key Research and Development Program of China [grant number 2021YFC2100203]; the National Natural Science Foundation of China [grant number 32072162]; the Postgraduate Research & Practice Inno‑vation Program of Jiangsu Province [grant number KYCX18_1791], and the National First‑Class Discipline Program of Light Industry Technology and Engineering [grant number LITE2018‑09].

Declarations

Conflict of interest The authors declare that they have no conflict of interest.

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