Sulfur Starvation Research Articles

Sulfur starvation-induced autophagy in Saccharomyces cerevisiae involves SAM-dependent signaling and transcription activator Met4

Autophagy is a key lysosomal degradative mechanism allowing a prosurvival response to stresses, especially nutrient starvation. Here we investigate the mechanism of autophagy induction in response to sulfur starvation in Saccharomyces cerevisiae. We found that sulfur deprivation leads to rapid and widespread transcriptional induction of autophagy-related (ATG) genes in ways not seen under nitrogen starvation. This distinctive response depends mainly on the transcription activator of sulfur metabolism Met4. Consistently, Met4 is essential for autophagy under sulfur starvation. Depletion of either cysteine, methionine or SAM induces autophagy flux. However, only SAM depletion can trigger strong transcriptional induction of ATG genes and a fully functional autophagic response. Furthermore, combined inactivation of Met4 and Atg1 causes a dramatic decrease in cell survival under sulfur starvation, highlighting the interplay between sulfur metabolism and autophagy to maintain cell viability. Thus, we describe a pathway of sulfur starvation-induced autophagy depending on Met4 and involving SAM as signaling sulfur metabolite.

Transcriptional and metabolic profiling of sulfur starvation response in two monocots

BackgroundSulfur (S) is a mineral nutrient essential for plant growth and development, which is incorporated into diverse molecules fundamental for primary and secondary metabolism, plant defense, signaling, and maintaining cellular homeostasis. Although, S starvation response is well documented in the dicot model Arabidopsis thaliana, it is not clear if the same transcriptional networks control the response also in the monocots.ResultsWe performed series of physiological, expression, and metabolite analyses in two model monocot species, one representing the C3 plants, Oryza sativa cv. kitaake, and second representing the C4 plants, Setaria viridis. Our comprehensive transcriptomic analysis revealed twice as many differentially expressed genes (DEGs) in S. viridis than in O. sativa under S-deficiency, consistent with a greater loss of sulfur and S-containing metabolites under these conditions. Surprisingly, most of the DEGs and enriched gene ontology terms were species-specific, with an intersect of only 58 common DEGs. The transcriptional networks were different in roots and shoots of both species, in particular no genes were down-regulated by S-deficiency in the roots of both species.ConclusionsOur analysis shows that S-deficiency seems to have different physiological consequences in the two monocot species and their nutrient homeostasis might be under distinct control mechanisms.

N2-Acetylornithine deacetylase functions as a Cys-Gly dipeptidase in the cytosolic glutathione degradation pathway in Arabidopsis thaliana.

Glutathione (GSH) is required for various physiological processes in plants, including redox regulation and detoxification of harmful compounds. GSH also functions as a repository for assimilated sulfur and is actively catabolized in plants. In Arabidopsis, GSH is mainly degraded initially by cytosolic enzymes, γ-glutamyl cyclotransferase, and γ-glutamyl peptidase, which release cysteinylglycine (Cys-Gly). However, the subsequent enzyme responsible for catabolizing this dipeptide has not been identified to date. In the present study, we identified At4g17830 as a Cys-Gly dipeptidase, namely cysteinylglycine peptidase 1 (CGP1). CGP1 complemented the phenotype of the yeast mutant that cannot degrade Cys-Gly. The Arabidopsis cgp1 mutant had lower Cys-Gly degradation activity than the wild type and showed perturbed concentrations of thiol compounds. Recombinant CGP1 showed reasonable Cys-Gly degradation activity in vitro. Metabolomic analysis revealed that cgp1 exhibited signs of severe sulfur deficiency, such as elevated accumulation of O-acetylserine (OAS) and the decrease in sulfur-containing metabolites. Morphological changes observed in cgp1, including longer primary roots of germinating seeds, were also likely associated with sulfur starvation. Notably, At4g17830 has previously been reported to encode an N2-acetylornithine deacetylase (NAOD) that functions in the ornithine biosynthesis. The cgp1 mutant did not show a decrease in ornithine content, whereas the analysis of CGP1 structure did not rule out the possibility that CGP1 has Cys-Gly dipeptidase and NAOD activities. Therefore, we propose that CGP1 is a Cys-Gly dipeptidase that functions in the cytosolic GSH degradation pathway and may play dual roles in GSH and ornithine metabolism.

Collective production of hydrogen sulfide gas enables budding yeast lacking MET17 to overcome their metabolic defect.

Assimilation of sulfur is vital to all organisms. In S. cerevisiae, inorganic sulfate is first reduced to sulfide, which is then affixed to an organic carbon backbone by the Met17 enzyme. The resulting homocysteine can then be converted to all other essential organosulfurs such as methionine, cysteine, and glutathione. This pathway has been known for nearly half a century, and met17 mutants have long been classified as organosulfur auxotrophs, which are unable to grow on sulfate as their sole sulfur source. Surprisingly, we found that met17Δ could grow on sulfate, albeit only at sufficiently high cell densities. We show that the accumulation of hydrogen sulfide gas underpins this density-dependent growth of met17Δ on sulfate and that the locus YLL058W (HSU1) enables met17Δ cells to assimilate hydrogen sulfide. Hsu1 protein is induced during sulfur starvation and under exposure to high sulfide concentrations in wild-type cells, and the gene has a pleiotropic role in sulfur assimilation. In a mathematical model, the low efficiency of sulfide assimilation in met17Δ can explain the observed density-dependent growth of met17Δ on sulfate. Thus, having uncovered and explained the paradoxical growth of a commonly used "auxotroph," our findings may impact the design of future studies in yeast genetics, metabolism, and volatile-mediated microbial interactions.

Sulfate limitation increases specific plasmid DNA yield and productivity in E. coli fed-batch processes

Plasmid DNA (pDNA) is a key biotechnological product whose importance became apparent in the last years due to its role as a raw material in the messenger ribonucleic acid (mRNA) vaccine manufacturing process. In pharmaceutical production processes, cells need to grow in the defined medium in order to guarantee the highest standards of quality and repeatability. However, often these requirements result in low product titer, productivity, and yield. In this study, we used constraint-based metabolic modeling to optimize the average volumetric productivity of pDNA production in a fed-batch process. We identified a set of 13 nutrients in the growth medium that are essential for cell growth but not for pDNA replication. When these nutrients are depleted in the medium, cell growth is stalled and pDNA production is increased, raising the specific and volumetric yield and productivity. To exploit this effect we designed a three-stage process (1. batch, 2. fed-batch with cell growth, 3. fed-batch without cell growth). The transition between stage 2 and 3 is induced by sulfate starvation. Its onset can be easily controlled via the initial concentration of sulfate in the medium. We validated the decoupling behavior of sulfate and assessed pDNA quality attributes (supercoiled pDNA content) in E. coli with lab-scale bioreactor cultivations. The results showed an increase in supercoiled pDNA to biomass yield by 33% and an increase of supercoiled pDNA volumetric productivity by 13 % upon limitation of sulfate. In conclusion, even for routinely manufactured biotechnological products such as pDNA, simple changes in the growth medium can significantly improve the yield and quality.Graphical

Identification of ascorbate- and salicylate-responsive miRNAs and verification of the spectral control of miR395 in Arabidopsis.

We assumed that miRNAs might regulate the physiological and biochemical processes in plants through their effects on the redox system and phytohormones. To check this hypothesis, the transcriptome profile of wild-type Arabidopsis and lines with decreased ascorbate (Asc), glutathione (GSH), or salicylate (Sal) levels were compared. GSH deficiency did not influence the miRNA expression, whereas lower levels of Asc and Sal reduced the accumulation of 9 and 44 miRNAs, respectively, but only four miRNAs were upregulated. Bioinformatics analysis revealed that their over-represented target genes are associated with the synthesis of nitrogen-containing and aromatic compounds, nucleic acids, and sulphate assimilation. Among them, the sulphate reduction-related miR395 - ATP-sulfurylase couple was selected to check the assumed modulating role of the light spectrum. A greater induction of the Asc- and Sal-responsive miR395 was observed under sulphur starvation in far-red light compared to white and blue light in wild-type and GSH-deficient Arabidopsis lines. Sal deficiency inhibited the induction of miR395 by sulphur starvation in blue light, whereas Asc deficiency greatly reduced it independently of the spectrum. Interestingly, sulphur starvation decreased only the level of ATP sulfurylase 4 among the miR395 target genes in far-red light. The expression level of ATP sulfurylase 3 was higher in far-red light than in blue light in wild-type and Asc-deficient lines. The results indicate the coordinated control of miRNAs by the redox and hormonal system since 11 miRNAs were affected by both Asc and Sal deficiency. This process can be modulated by light spectrum, as shown for miR395.

Network analysis for identifying potential anti-virulence targets from whole transcriptome of Pseudomonas aeruginosa and Staphylococcus aureus exposed to certain anti-pathogenic polyherbal formulations

Introduction: Antimicrobial resistance (AMR) is a serious global threat. Identification of novel antibacterial targets is urgently warranted to help antimicrobial drug discovery programs. This study attempted identification of potential targets in two important pathogens Pseudomonas aeruginosa and Staphylococcus aureus. Methods: Transcriptomes of P. aeruginosa and S. aureus exposed to two different quorum-modulatory polyherbal formulations were subjected to network analysis to identify the most highly networked differentially expressed genes (hubs) as potential anti-virulence targets. Results: Genes associated with denitrification and sulfur metabolism emerged as the most important targets in P. aeruginosa. Increased buildup of nitrite (NO2) in P. aeruginosa culture exposed to the polyherbal formulation Panchvalkal was confirmed through in vitro assay too. Generation of nitrosative stress and inducing sulfur starvation seemed to be effective anti-pathogenic strategies against this notorious gram-negative pathogen. Important targets identified in S. aureus were the transcriptional regulator sarA, immunoglobulin-binding protein Sbi, serine protease SplA, the saeR/S response regulator system, and gamma-hemolysin components hlgB and hlgC. Conclusion: Further validation of the potential targets identified in this study is warranted through appropriate in vitro and in vivo assays in model hosts. Such validated targets can prove vital to many antibacterial drug discovery programs globally.

Network analysis for identifying potential anti-virulence targets from whole transcriptome of Pseudomonas aeruginosa and Staphylococcus aureus exposed to certain anti-pathogenic polyherbal formulations.

Antimicrobial resistance (AMR) is a serious global threat. Identification of novel antibacterial targets is urgently warranted to help antimicrobial drug discovery programs. This study attempted identification of potential targets in two important pathogens Pseudomonas aeruginosa and Staphylococcus aureus. Transcriptomes of P. aeruginosa and S. aureus exposed to two different quorum-modulatory polyherbal formulations were subjected to network analysis to identify the most highly networked differentially expressed genes (hubs) as potential anti-virulence targets. Genes associated with denitrification and sulfur metabolism emerged as the most important targets in P. aeruginosa. Increased buildup of nitrite (NO2) in P. aeruginosa culture exposed to the polyherbal formulation Panchvalkal was confirmed through in vitro assay too. Generation of nitrosative stress and inducing sulfur starvation seemed to be effective anti-pathogenic strategies against this notorious gram-negative pathogen. Important targets identified in S. aureus were the transcriptional regulator sarA, immunoglobulin-binding protein Sbi, serine protease SplA, the saeR/S response regulator system, and gamma-hemolysin components hlgB and hlgC. Further validation of the potential targets identified in this study is warranted through appropriate invitro and in vivo assays in model hosts. Such validated targets can prove vital to many antibacterial drug discovery programs globally.

The Arabidopsis ATP-Binding Cassette E protein ABCE2 is a conserved component of the translation machinery.

ATP-Binding Cassette E (ABCE) proteins dissociate cytoplasmic ribosomes after translation terminates, and contribute to ribosome recycling, thus linking translation termination to initiation. This function has been demonstrated to be essential in animals, fungi, and archaea, but remains unexplored in plants. In most species, ABCE is encoded by a single-copy gene; by contrast, Arabidopsis thaliana has two ABCE paralogs, of which ABCE2 seems to conserve the ancestral function. We isolated apiculata7-1 (api7-1), the first viable, hypomorphic allele of ABCE2, which has a pleiotropic morphological phenotype reminiscent of mutations affecting ribosome biogenesis factors and ribosomal proteins. We also studied api7-2, a null, recessive lethal allele of ABCE2. Co-immunoprecipitation experiments showed that ABCE2 physically interacts with components of the translation machinery. An RNA-seq study of the api7-1 mutant showed increased responses to iron and sulfur starvation. We also found increased transcript levels of genes related to auxin signaling and metabolism. Our results support for the first time a conserved role for ABCE proteins in translation in plants, as previously shown for the animal, fungal, and archaeal lineages. In Arabidopsis, the ABCE2 protein seems important for general growth and vascular development, likely due to an indirect effect through auxin metabolism.

Bioreduction mechanisms of high-concentration hexavalent chromium using sulfur salts by photosynthetic bacteria

Eliminating “sulfur starvation” caused by competition for sulfate transporters between chromate and sulfate is crucial to enhance the content of sulfur-containing compounds and improve the tolerance and reduction capability of Cr(VI) in bacteria. In this study, the effects of sulfur salts on the Cr(VI) bioremediation and the possible mechanism were investigated in Rhodobacter sphaeroides SC01 by cell imaging, spectroscopy, and biochemical measurements. The results showed that, when the concentration of metabisulfite was 2.0 g L−1, and the initial OD600 was 0.33, the reduction rate of R. sphaeroides SC01 reached up to 91.3% for 500 mg L−1 Cr(VI) exposure at 96 h. Moreover, thiosulfate and sulfite also markedly increased the concentration of reduced Cr(VI) in R. sphaeroides SC01. Furthermore, the characterization results revealed that –OH, –CONH, –COOH, –SO3, -PO3, and -S-S- played a major role in the adsorption of Cr, and Cr(III) reduced by bacteria was bioprecipitated in the production of Cr2P3S9 and CrPS4. In addition, R. sphaeroids SC01 combined with metabisulfite significantly increased the activity of glutathione peroxidase and the content of glutathione (GSH) and total sulfhydryl while decreasing reactive oxygen species (ROS) accumulation and cell death induced by Cr(VI) toxic. Overall, the results of this research revealed a highly efficient and reliable strategy for Cr(VI) removal by photosynthetic bacteria combined with sulfur salts in high-concentration Cr(VI)-contaminated wastewater.

Cytokinin modulates the metabolic network of sulfur and glutathione.

The phytohormone cytokinin is implicated in a range of growth, developmental, and defense processes. A growing body of evidence supports a crosstalk between cytokinin and nutrient signaling pathways, such as nitrate availability. Cytokinin signaling regulates sulfur-responsive gene expression, but the underlying molecular mechanisms and their impact on sulfur-containing metabolites have not been systematically explored. Using a combination of genetic and pharmacological tools, we investigated the interplay between cytokinin signaling and sulfur homeostasis. Exogenous cytokinin triggered sulfur starvation-like gene expression accompanied by a decrease in sulfate and glutathione content. This process was uncoupled from the activity of the major transcriptional regulator of sulfate starvation signaling SULFUR LIMITATION 1 and an important glutathione-degrading enzyme, γ-glutamyl cyclotransferase 2;1, expression of which was robustly up-regulated by cytokinin. Conversely, glutathione accumulation was observed in mutants lacking the cytokinin receptor ARABIDOPSIS HISTIDINE KINASE 3 and in cytokinin-deficient plants. Cytokinin-deficient plants displayed improved root growth upon exposure to glutathione-depleting chemicals which was attributed to a higher capacity to maintain glutathione levels. These results shed new light on the interplay between cytokinin signaling and sulfur homeostasis. They position cytokinin as an important modulator of sulfur uptake, assimilation, and remobilization in plant defense against xenobiotics and root growth.

Starch accumulation dynamics and transcriptome analysis of Chlorella sorokiniana during transition of sulfur nutritional status

Microalgae can effectively accumulate starch by using nutritional limitation methods in the context of bioalcohol fuel production. However, relatively few studies have focused on starch accumulation in microalgae and its molecular basis, especially under sulfur limitation conditions. In this study, the starch accumulation dynamics and physiological responses of Chlorella sorokiniana under sulfur starvation (SS) and sulfur replenishment (SR) conditions were investigated, and the genes involved in the transcriptional regulation were explored using RNA-seq. The starch content in C. sorokiniana cells significantly increased from 1.6% to 55.0% of dry weight within 24 h under SS conditions, and then, it decreased to 3.4% within 12 h after transition to SR conditions. However, cell growth was inhibited, and pigment content decreased under SS conditions. Using RNA-seq analysis, a total of 9720 differentially expressed genes (DEGs) induced by sulfur status were obtained. These genes were narrowed down to 454 starvation and replenishment cross-validated (SRV)-DEGs, among which 283 SRV-DEGs were significantly up-regulated and 171 SRV-DEGs were down-regulated under SS conditions, and returned to their previous state under SR conditions. The SRV-DEGs enriched in the sulfate metabolism pathway were all up-regulated under SS conditions after 6 h to speed up the sulfur metabolic cycle, and the transcriptional abundance of a sulfate transporter (SULTR4), cysteine synthase[O-acetylserine(thiol)-lyase] (OASTL), serine acetyltransferase (SAT), and methanethiol oxidase (SELENBP1) increased 8.6-fold, 12.6-fold, 8.7-fold, and 12.4-fold, respectively. Protein synthesis was correspondingly inhibited, which resulted in the reallocation of carbon and elevated the starch synthesis pathway, in which the expressions of glycogen branching enzyme (GBE) and starch synthase (SS) were up-regulated 12.0- and 3.0-fold, respectively. The fatty acid desaturase (FAD) and phosphatidic acid phosphatase (PAP) in the lipid synthesis pathway were strongly up-regulated 8.8- and 16.2-fold, respectively, indicating the competitive synthesis of lipids. The down-regulation of SRV-DEGs associated with carbon fixation, such as those in the Calvin cycle, possibly affected cell growth. The time-resolved transcriptional analysis identified the SRV-DEGs, revealing the underlying starch accumulation mechanism, as well as the relationship with cell growth and lipid synthesis.

Glutathione degradation activity of γ-glutamyl peptidase 1 manifests its dual roles in primary and secondary sulfur metabolism in Arabidopsis.

Glutathione (GSH) functions as a major sulfur repository and hence occupies an important position in primary sulfur metabolism. GSH degradation results in sulfur reallocation and is believed to be carried out mainly by γ-glutamyl cyclotransferases (GGCT2;1, GGCT2;2, and GGCT2;3), which, however, do not fully explain the rapid GSH turnover. Here, we discovered that γ-glutamyl peptidase 1 (GGP1) contributes to GSH degradation through a yeast complementation assay. Recombinant proteins of GGP1, as well as GGP3, showed high degradation activity of GSH, but not of oxidized glutathione (GSSG), in vitro. Notably, the GGP1 transcripts were highly abundant in rosette leaves, in agreement with the ggp1 mutants constantly accumulating more GSH regardless of nutritional conditions. Given the lower energy requirements of the GGP- than the GGCT-mediated pathway, the GGP-mediated pathway could be a more efficient route for GSH degradation than the GGCT-mediated pathway. Therefore, we propose a model wherein cytosolic GSH is degraded chiefly by GGP1 and likely also by GGP3. Another noteworthy fact is that GGPs are known to process GSH conjugates in glucosinolate and camalexin synthesis; indeed, we confirmed that the ggp1 mutant contained higher levels of O-acetyl-l-Ser, a signaling molecule for sulfur starvation, and lower levels of glucosinolates and their degradation products. The predicted structure of GGP1 further provided a rationale for this hypothesis. In conclusion, we suggest that GGP1 and possibly GGP3 play vital roles in both primary and secondary sulfur metabolism.

Effects of seawater sulfur starvation and enrichment on Gracilaria gracilis growth and biochemical composition

The genus Gracilaria, largest biomass producer in coastal regions, encompasses a wide range of species including Gracilaria gracilis. Nowadays, there is a spate of interest in its culture in lagoon where the water sulfate concentration is variable. A laboratory culture was carried out to determine the sulfate concentration effect on their growth as well as their biochemical composition, which were 2.5, 27 or 50 mM, referred to as SSS (sulfur starved seawater), SW (seawater) and SES (sulfur enriched seawater).We found that the sulfate content of the surrounding medium is a key parameter influencing both the alga growth and its composition. However, seawater proved to be the most suitable environment to sustain alga growth, proteins, R-phycoerythrin and agar yields, but sulfur enrichment and starvation affects them. The sulfate degree of agar and therefore its quality is related to the medium sulfate concentration. We conclude that sulfur starvation (2.5 mM) for three weeks, led to severe growth retardation, lower agar yield and quality and indicated the limit potential of G. gracilis for mariculture under these conditions. These results demonstrated that the success of G. gracilis culture in the lagoon is feasible if sulfate concentration is closer to that of seawater.

Improving carbohydrate accumulation in Chlamydomonas debaryana induced by sulfur starvation using response surface methodology.

Most methods that promote carbohydrate production negatively affect cell growth and microalgal biomass production. This study explores, in a two-stage cultivation strategy, in Chlamydomonas debaryana the optimization of certain culture conditions for high carbohydrate production without loss of biomass. In the first stage, the interaction between sodium bicarbonate supplementation, aeration, and different growth periods was optimized using the response surface methodology (RMS). The 3-factor Box-Behnken design (BBD) was applied, and a second-order polynomial regression analysis was used to analyze the experimental data. The results showed that 0.45g L-1 of sodium bicarbonate combined with a good aerated agitation (0.6 L min-1) and a cultivation period of 18days are optimal to produce 5.02g L-1 of biomass containing 43% of carbohydrates.Under these optimized growth conditions, accumulation of carbohydrates was studied using different modes of nutritional stress. The results indicated that carbohydrate content was improved and the maximum accumulation (about 60% of the dry weight) was recorded under sulfur starvation with only a 14% reduction in biomass as compared to control. This study showed promising results as to biomass production and carbohydrate yield by the microalgae C. debaryana in view of production of third-generation biofuels.

Coordinated bacterial and plant sulfur metabolism in Enterobacter sp. SA187–induced plant salt stress tolerance

Enterobacter sp. SA187 is a root endophytic bacterium that maintains growth and yield of plants under abiotic stress conditions. In this work, we compared the metabolic wirings of Arabidopsis and SA187 in the free-living and endophytic interaction states. The interaction of SA187 with Arabidopsis induced massive changes in bacterial gene expression for chemotaxis, flagellar biosynthesis, quorum sensing, and biofilm formation. Besides modification of the bacterial carbon and energy metabolism, various nutrient and metabolite transporters and the entire sulfur pathway were up-regulated. Under salt stress, Arabidopsis resembled plants under sulfate starvation but not when colonized by SA187, which reprogramed the sulfur regulon of Arabidopsis. In accordance, salt hypersensitivity of multiple Arabidopsis sulfur metabolism mutants was partially or completely rescued by SA187 as much as by the addition of sulfate, L-cysteine, or L-methionine. Many components of the sulfur metabolism that are localized in the chloroplast were partially rescued by SA187. Finally, salt-induced accumulation of reactive oxygen species as well as the hypersensitivity of LSU mutants were suppressed by SA187. LSUs encode a central regulator linking sulfur metabolism to chloroplast superoxide dismutase activity. The coordinated regulation of the sulfur metabolic pathways in both the beneficial microorganism and the host plant is required for salt stress tolerance in Arabidopsis and might be a common mechanism utilized by different beneficial microbes to mitigate the harmful effects of different abiotic stresses on plants.

Aspects of the Neurospora crassa Sulfur Starvation Response Are Revealed by Transcriptional Profiling and DNA Affinity Purification Sequencing.

ABSTRACTAccurate nutrient sensing is important for rapid fungal growth and exploitation of available resources. Sulfur is an important nutrient source found in a number of biological macromolecules, including proteins and lipids. The model filamentous fungus Neurospora crassa is capable of utilizing sulfur found in a variety of sources from amino acids to sulfate. During sulfur starvation, the transcription factor CYS-3 is responsible for upregulation of genes involved in sulfur uptake and assimilation. Using a combination of RNA sequencing and DNA affinity purification sequencing, we performed a global survey of the N. crassa sulfur starvation response and the role of CYS-3 in regulating sulfur-responsive genes. The CYS-3 transcription factor bound the promoters and regulated genes involved in sulfur metabolism. Additionally, CYS-3 directly activated the expression of a number of uncharacterized transporter genes, suggesting that regulation of sulfur import is an important aspect of regulation by CYS-3. CYS-3 also directly regulated the expression of genes involved in mitochondrial electron transfer. During sulfur starvation, genes involved in nitrogen metabolism, such as amino acid and nucleic acid metabolic pathways, along with genes encoding proteases and nucleases that are necessary for scavenging nitrogen, were activated. Sulfur starvation also caused changes in the expression of genes involved in carbohydrate metabolism, such as those encoding glycosyl hydrolases. Thus, our data suggest a connection between sulfur metabolism and other aspects of cellular metabolism.IMPORTANCE Identification of nutrients present in the environment is a challenge common to all organisms. Sulfur is an important nutrient source found in proteins, lipids, and electron carriers that are required for the survival of filamentous fungi such as Neurospora crassa. Here, we transcriptionally profiled the response of N. crassa to characterize the global response to sulfur starvation. We also used DNA affinity purification sequencing to identify the direct downstream targets of the transcription factor responsible for regulating genes involved in sulfur uptake and assimilation. Along with genes involved in sulfur metabolism, this transcription factor regulated a number of uncharacterized transporter genes and genes involved in mitochondrial electron transfer. Our data also suggest a connection between sulfur, nitrogen, and carbon metabolism, indicating that the regulation of a number of metabolic pathways is intertwined.

Resource allocation in Chlamydomonas reinhardtii exposed to sulphur starvation and anaerobiosis

ABSTRACT When environmental conditions change, microalgae can modulate the size of their macromolecular pools and their elemental composition in order to accommodate their nutritional and energetic demands. In this context, we evaluated how O2 levels and S starvation alter the resource partitioning in terms of organic and elemental composition in Chlamydomonas reinhardtii. The analysis of gene transcription, photosynthetic energy production and organic pool allocation allowed us to identify a two-phase cellular response related to external O2 concentration: (1) a short-term response where S starvation induced the expression of S starvation genes immediately after the start of the perturbation; (2) a long-term response that consisted of a drastic reduction of photosynthetic energy production and changes in C allocation following S deprivation, which was related to oxygen conditions. In anaerobic conditions, S starvation did not affect the pools of carbohydrates and lipids, whereas the protein pool decreased slightly over time. On the other hand, in aerobic conditions, S starvation modulated all organic pools: carbohydrates, lipids and proteins. We conclude that the energy levels produced by fermentation metabolism are not sufficient to sustain the acclimation process in C. reinhardtii; moreover, S availability affects C. reinhardtii more than do oxygen levels.

Biodesulfurization Induces Reprogramming of Sulfur Metabolism in Rhodococcus qingshengii IGTS8: Proteomics and Untargeted Metabolomics.

ABSTRACTSulfur metabolism in fuel-biodesulfurizing bacteria and the underlying physiological adaptations are not understood, which has impeded the development of a commercially viable bioprocess for fuel desulfurization. To fill these knowledge gaps, we performed comparative proteomics and untargeted metabolomics in cultures of the biodesulfurization reference strain Rhodococcus qingshengii IGTS8 grown on either inorganic sulfate or the diesel-borne organosulfur compound dibenzothiophene as a sole sulfur source. Dibenzothiophene significantly altered the biosynthesis of many sulfur metabolism proteins and metabolites in a growth phase-dependent manner, which enabled us to reconstruct the first experimental model for sulfur metabolism in a fuel-biodesulfurizing bacterium. All key pathways related to assimilatory sulfur metabolism were represented in the sulfur proteome, including uptake of the sulfur sources, sulfur acquisition, and assimilatory sulfate reduction, in addition to biosynthesis of key sulfur-containing metabolites such as S-adenosylmethionine, coenzyme A, biotin, thiamin, molybdenum cofactor, mycothiol, and ergothioneine (low-molecular weight thiols). Fifty-two proteins exhibited significantly different abundance during at least one growth phase. Sixteen proteins were uniquely detected and 47 proteins were significantly more abundant in the dibenzothiophene culture during at least one growth phase. The sulfate-free dibenzothiophene-containing culture reacted to sulfate starvation by restricting sulfur assimilation, enforcing sulfur-sparing, and maintaining redox homeostasis. Biodesulfurization triggered alternative pathways for sulfur assimilation different from those operating in the inorganic sulfate culture. Sulfur metabolism reprogramming and metabolic switches in the dibenzothiophene culture were manifested in limiting sulfite reduction and biosynthesis of cysteine, while boosting the production of methionine via the cobalamin-independent pathway, as well as the biosynthesis of the redox buffers mycothiol and ergothioneine. The omics data underscore the key role of sulfur metabolism in shaping the biodesulfurization phenotype and highlight potential targets for improving the biodesulfurization catalytic activity via metabolic engineering.IMPORTANCE For many decades, research on biodesulfurization of fossil fuels was conducted amid a large gap in knowledge of sulfur metabolism and its regulation in fuel-biodesulfurizing bacteria, which has impeded the development of a commercially viable bioprocess. In addition, lack of understanding of biodesulfurization-associated metabolic and physiological adaptations prohibited the development of efficient biodesulfurizers. Our integrated omics-based findings reveal the assimilatory sulfur metabolism in the biodesulfurization reference strain Rhodococcus qingshengii IGTS8 and show how sulfur metabolism and oxidative stress response were remodeled and orchestrated to shape the biodesulfurization phenotype. Our findings not only explain the frequently encountered low catalytic activity of native fuel-biodesulfurizing bacteria but also uncover unprecedented potential targets in sulfur metabolism that could be exploited via metabolic engineering to boost the biodesulfurization catalytic activity, a prerequisite for commercial application.

Response to sulfur in Schizosaccharomyces pombe

ABSTRACTSulfur is an essential component of various biologically important molecules, including methionine, cysteine and glutathione, and it is also involved in coping with oxidative and heavy metal stress. Studies using model organisms, including budding yeast (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe), have contributed not only to understanding various cellular processes but also to understanding the utilization and response mechanisms of each nutrient, including sulfur. Although fission yeast can use sulfate as a sulfur source, its sulfur metabolism pathway is slightly different from that of budding yeast because it does not have a trans-sulfuration pathway. In recent years, it has been found that sulfur starvation causes various cellular responses in S. pombe, including sporulation, cell cycle arrest at G2, chronological lifespan extension, autophagy induction and reduced translation. This MiniReview identifies two sulfate transporters in S. pombe, Sul1 (encoded by SPBC3H7.02) and Sul2 (encoded by SPAC869.05c), and summarizes the metabolic pathways of sulfur assimilation and cellular response to sulfur starvation. Understanding these responses, including metabolism and adaptation, will contribute to a better understanding of the various stress and nutrient starvation responses and chronological lifespan regulation caused by sulfur starvation.