Genome-Wide Investigation of the Cysteine Synthase Gene Family Shows That Overexpression of CSase Confers Alkali Tolerance to Alfalfa (Medicago sativa L.)

Alfalfa is widely grown worldwide as a perennial high-quality legume forage and as a good ecological landcover. The cysteine synthase (CSase) gene family is actively involved in plant growth and development and abiotic stress resistance but has not been systematically investigated in alfalfa. We identified 39 MsCSase genes on 4 chromosomes of the alfalfa genome. Phylogenetic analysis demonstrated that these genes were clustered into six subfamilies, and members of the same subfamily had similar physicochemical properties and sequence structures. Overexpression of the CSase gene in alfalfa increased alkali tolerance. Compared with control plants, the overexpression lines presented higher proline, soluble sugars, and cysteine and reduced glutathione contents and superoxide dismutase and peroxidase activities as well as lower hydrogen peroxide and superoxide anion contents after alkali stress. The relative expression of γ-glutamyl cysteine synthetase gene (a downstream gene of CSase) in the overexpression lines was much higher than that in the control line. The CSase gene enhanced alkalinity tolerance by regulating osmoregulatory substances and improving antioxidant capacity. These results provide a reference for studying the CSase gene family in alfalfa and expanding the alkali tolerance gene resources of forage plants.

Serine acetyltransferase from Neisseria gonorrhoeae; structural and biochemical basis of inhibition

Serine acetyltransferase (SAT) catalyzes the first step in the two-step pathway to synthesize L-cysteine in bacteria and plants. SAT synthesizes O-acetylserine from substrates L‑serine and acetyl coenzyme A and is a key enzyme for regulating cellular cysteine levels by feedback inhibition of L-cysteine, and its involvement in the cysteine synthase complex. We have performed extensive structural and kinetic characterization of the SAT enzyme from the antibiotic-resistant pathogen Neisseria gonorrhoeae. Using X-ray crystallography, we have solved the structures of NgSAT with the non-natural ligand, L-malate (present in the crystallization screen) to 2.01 Å and with the natural substrate L-serine (2.80 Å) bound. Both structures are hexamers, with each monomer displaying the characteristic left-handed parallel β-helix domain of the acyltransferase superfamily of enzymes. Each structure displays both extended and closed conformations of the C-terminal tail.  L‑malate bound in the active site results in an interesting mix of open and closed active site conformations, exhibiting a structural change mimicking the conformation of cysteine (inhibitor) bound structures from other organisms. Kinetic characterization shows competitive inhibition of L-cysteine with substrates L-serine and acetyl coenzyme A. The SAT reaction represents a key point for the regulation of cysteine biosynthesis and controlling cellular sulfur due to feedback inhibition by L-cysteine and formation of the cysteine synthase complex. Data presented here provide the structural and mechanistic basis for inhibitor design and given this enzyme is not present in humans could be explored to combat the rise of extensively antimicrobial-resistant N. gonorrhoeae.

Transcriptome and proteome profiling revealed molecular mechanism of selenium responses in bread wheat (Triticum aestivum L.)

Background Although selenium (Se) plays important roles in scavenging free radicals, alleviating oxidative stresses, and strengthening immune system, the knowledge about Se responses in bread wheat is still limited. In order to clarify the molecular mechanism of Se responses in bread wheat, 2-week-old wheat seedlings of cultivar ‘Jimai22’ treated with 10 μM disodium selenate (Na2SeO4) for 0, 3, and 24 h were collected and analyzed by transcriptional sequencing and tandem mass tag-based (TMT) quantitative proteomics. Results At least 11,656 proteins and 133,911 genes were identified, and proteins including ATP sulfurylase (APS), cysteine synthase (CS), SeCys lyase, sulfate transporters, glutathione S-transferase (GSTs), glutathione peroxidase (GSH-Px), glutaredoxins (GRXs), superoxide dismutases (SODs), catalases (CATs), heat shock proteins (HSPs), UDP-glycose flavonoid glycosyltransferases (UFGTs), sucrose-6-phosphate hydrolases (Suc-6-PHs), archaeal phosphoglucose isomerases (APGIs), malate synthases (MSs), and endo-1,4-beta-xylanase (Xyn) in Se accumulation, ROS scavenging, secondary metabolism, and carbohydrate metabolism were significantly differently expressed. Conclusions This is the first complementary analyses of the transcriptome and proteome related with selenium responses in bread wheat. Our work enhances the understanding about the molecular mechanism of selenium responses in bread wheat.

L-Cysteine Synthase Enhanced Sulfide Biotransformation in Subtropical Marine Mangrove Sediments as Revealed by Metagenomics Analysis

High sulfides concentrations can be poisonous to environment because of anthropogenic waste production or natural occurrences. How to elucidate the biological transformation mechanisms of sulfide pollutants in the subtropical marine mangrove ecosystem has gained increased interest. Thus, in the present study, the sulfide biotransformation in subtropical mangroves ecosystem was accurately evaluated using metagenomic sequencing and quantitative polymerase chain reaction analysis. Most abundant genes were related to the organic sulfur transformation. Furthermore, an ecological model of sulfide conversion was constructed. Total phosphorus was the dominant environmental factor that drove the sulfur cycle and microbial communities. We compared mangrove and non-mangrove soils and found that the former enhanced metabolism that was related to sulfate reduction when compared to the latter. Total organic carbon, total organic nitrogen, iron, and available sulfur were the key environmental factors that effectively influenced the dissimilatory sulfate reduction. The taxonomic assignment of dissimilatory sulfate-reducing genes revealed that Desulfobacterales and Chromatiales were mainly responsible for sulfate reduction. Chromatiales were most sensitive to environmental factors. The high abundance of cysE and cysK could contribute to the coping of the microbial community with the toxic sulfide produced by Desulfobacterales. Collectively, these findings provided a theoretical basis for the mechanism of the sulfur cycle in subtropical mangrove ecosystems.

Transcriptome and Proteome Profiling Revealed Molecular Mechanism of Selenium Responses in Bread Wheat (Triticum Aestivum L.)

Background: Although selenium (Se) plays important roles in scavenging free radicals, alleviating oxidative stresses, and strengthening immune system, the knowledge about Se response in bread wheat is still limited. In order to clarify the molecular mechanism of Se response in bread wheat, 2-week-old wheat seedlings of cultivar ‘Jimai22’ treated with 10 μM disodium selenate (Na2SeO4) for 0 h, 3 h, and 24 h were collected and analyzed by transcriptional sequencing and tandem mass tag-based (TMT) quantitative proteomics. Results: At least 11656 proteins and 133911 genes were identified, and proteins including ATP sulfurylase (APS), cysteine synthase (CS), SeCys lyase, sulfate transporters, glutathione S-transferase (GSTs), glutathione peroxidase (GSH-Px), glutaredoxins (GRXs), superoxide dismutases (SODs), catalases (CATs), heat shock proteins (HSPs), UDP-glycose flavonoid glycosyltransferases (UFGTs), sucrose-6-phosphate hydrolases (Suc-6-PHs), archaeal phosphoglucose isomerases (APGIs), malate synthases (MSs), and endo-1,4-beta-xylanase (Xyn) in Se accumulation, ROS scavenging, secondary metabolism, and carbohydrate metabolism were significantly differently expressed.Conclusions: This is the first complementary analyses of the transcriptome and proteome related with selenium responses in bread wheat. Our work enhances the understanding about the molecular mechanism of selenium responses in bread wheat.

A Competitive O-Acetylserine Sulfhydrylase Inhibitor Modulates the Formation of Cysteine Synthase Complex

Cysteine is the main precursor of sulfur-containing biological molecules in bacteria and contributes to the control of the cell redox state. Hence, this amino acid plays an essential role in microbial survival and pathogenicity and the reductive sulfate assimilation pathway is considered a promising target for the development of new antibacterials. Serine acetyltransferase (SAT) and O-acetylserine sulfhydrylase (OASS-A), the enzymes catalyzing the last two steps of cysteine biosynthesis, engage in the formation of the cysteine synthase (CS) complex. The interaction between SAT and OASS-A finely tunes cysteine homeostasis, and the development of inhibitors targeting either protein–protein interaction or the single enzymes represents an attractive strategy to undermine bacterial viability. Given the peculiar mode of interaction between SAT and OASS-A, which exploits the insertion of SAT C-terminal sequence into OASS-A active site, we tested whether a recently developed competitive inhibitor of OASS-A exhibited any effect on the CS stability. Through surface plasmon resonance spectroscopy, we (i) determined the equilibrium constant for the Salmonella Typhimurium CS complex formation and (ii) demonstrated that the inhibitor targeting OASS-A active site affects CS complex formation. For comparison, the Escherichia coli CS complex was also investigated, with the aim of testing the potential broad-spectrum activity of the candidate antimicrobial compound.