Assimilation Pathway Research Articles

Sulfate-reducing bacteria block cadmium and lead uptake in rice by regulating sulfur metabolism.

This study was dedicated to investigating the role of sulfur metabolic processes in sulfate-reducing bacteria in plant resistance to heavy metal contamination. We constructed sulfate-reducing bacterial communities based on the functional properties of sulfate-reducing strains, and then screened out the most effective sulfate-reducing bacterial community SYN1, that prevented Cd and Pb uptake in rice through hydroponic experiment. This community lowered Cd levels in the roots and upper roots by 36.60% and 39.88%, respectively, and Pb levels by 35.96% and 51.54%. We also compared two treatment groups, inoculated with SYN1 and exogenously added GSH, and found that both enhanced the antioxidant response of the plants, increased the lignin and GSH contents and the expression of genes related to the phenylpropane biosynthesis pathway (OsCAD, Os4CL, OsCOMT, OsPOD, OsC3H, and OsPAL), and decreased the expression of heavy metal transporter genes (OsHMA2, OsIRT1) expression. There were no significant differences between the two treatments. Sulfate-reducing bacteria produce GSH through the sulfur assimilation pathway, and GSH can directly chelate heavy metals or enhance plant antioxidant enzyme activities and regulate processes such as uptake and translocation of heavy metals, thus enhancing plant resistance to heavy metal toxicity.

Unraveling nitrogen metabolism, cold and stress adaptation in polar Bosea sp. PAMC26642 through comparative genome analysis

Nitrogen metabolism, related genes, and other stress-resistance genes are poorly understood in Bosea strain. To date, most of the research work in Bosea strains has been focused on thiosulfate oxidation and arsenic reduction. This work aimed to better understand and identify genomic features that enable thiosulfate-oxidizing lichen-associated Bosea sp. PAMC26642 from the Arctic region of Svalbard, Norway, to withstand harsh environments. Comparative genomic analysis was performed using various bioinformatics tools to compare Bosea sp. PAMC26642 with other strains of the same genus, emphasizing nitrogen metabolism and stress adaptability. During genomic analysis of Bosea sp. PAMC26642, assimilatory nitrogen metabolic pathway and its associated enzymes such as nitrate reductase, NAD(P)H-nitrite reductase, ferredoxin-nitrite reductase, glutamine synthetase, glutamine synthase, and glutamate dehydrogenase were identified. In addition, carbonic anhydrase, cyanate lyase, and nitronate monooxygenase were also identified. Furthermore, the strain demonstrated nitrate reduction at two different temperatures (15°C and 25°C). Enzymes associated with various stress adaptation pathways, including oxidative stress (superoxide dismutase, catalase, and thiol peroxidase), osmotic stress (OmpR), temperature stress (Csp and Hsp), and heavy metal resistance, were also identified. The average Nucleotide Identity (ANI) value is found to be below the threshold of 94-95%, indicating this bacterium might be a potential new species. This study is very helpful in determining the diversity of thiosulfate-oxidizing nitrate-reducing bacteria, as well as their ability to adapt to extreme environments. These bacteria can be used in the future for environmental, biotechnological, and agricultural purposes, particularly in processes involving sulfur and nitrogen transformation.

Nitrogen Assimilation Plays a Role in Balancing the Chloroplastic Glutathione Redox Potential Under High Light Conditions.

Nitrate reduction requires reducing equivalents produced by the photosynthetic electron transport chain. Therefore, it has been suggested that nitrate assimilation provides a sink for electrons under high light conditions. We tested this hypothesis by monitoring photosynthetic efficiency and the chloroplastic glutathione redox potential (chl-EGSH) of plant lines with mutated glutamine synthetase 2 (GS2) and ferredoxin-dependent glutamate synthase 1 (GOGAT1). Mutant lines incorporated significantly less isotopically-labelled nitrate into amino acids than wild-type plants, demonstrating impaired nitrogen assimilation. When nitrate assimilation was compromised, photosystem II (PSII) proved more vulnerable to photodamage. The effect of the nitrate assimilation pathway on the chl- EGSH was monitored using the chloroplast-targeted roGFP2 biosensor (chl-roGFP2). Remarkably, while oxidation followed by reduction of chl-roGFP2 was detected in WT plants in response to high light, oxidation values were stable in the mutant lines, suggesting that chl-EGSH relaxation after high light-induced oxidation is achieved by diverting excess electrons to the nitrogen assimilation pathway. Importantly, similar ΦPSII and chl-roGFP2 patterns were observed at elevated CO2, suggesting that mutant phenotypes are not associated with photorespiration activity. Together, these findings indicate that the nitrogen assimilation pathway serves as a sustainable energy dissipation route, ensuring efficient photosynthetic activity and fine-tuning redox metabolism under light-saturated conditions.

Evolutionary engineering of Saccharomyces cerevisiae: Crafting a synthetic methylotroph via self-reprogramming.

Methanol, as a non-edible feedstock, offers a promising sustainable alternative to sugar-based substrates in biochemical production. Despite progress in engineering methanol assimilation in nonmethylotrophs, the full transformation into methanol-dependent synthetic methylotrophs remains a formidable challenge. Here, moving beyond the conventional rational design principle, we engineered a synthetic methylotrophic Saccharomyces cerevisiae through genome rearrangement and adaptive laboratory evolution. This evolutionarily advanced strain unexpectedly shed the heterologous methanol assimilation pathway and demonstrated the robust growth on sole methanol. We discovered that the evolved strain likely realized methanol assimilation through a previously unidentified Adh2-Sfa1-rGly (ASrG) pathway, facilitating the concurrent assimilation of formate and CO2. Furthermore, the incorporation of electron transfer material C3N4 quantum dots obviously enhanced methanol-dependent growth, emphasizing the role of energy availability in the ASrG pathway. This breakthrough introduces a previously unidentified C1 utilization pathway and highlights the exceptional adaptability and self-evolving capacity of the S. cerevisiae metabolic network.

Advancements in Biological Conversion of C1 Feedstocks: Sustainable Bioproduction and Environmental Solutions.

The use of one-carbon (C1) feedstocks, including carbon dioxide (CO2), carbon monoxide (CO), formate (HCO2H), methanol (CH3OH), and methane (CH4), presents a significant opportunity for sustainable bioproduction and environmental conservation. This Perspective explores the development of biological methods for converting C1 feedstocks into valuable products, emphasizing major progress from engineering native C1 assimilation pathways to the creation of synthetic autotrophs and methylotrophs that utilize these carbon sources. Additionally, we discuss hybrid approaches that merge biological and electrochemical systems, particularly for the conversion of CO2. This Perspective underscores the importance of C1 bioconversion in promoting sustainable biotechnological strategies for a low-carbon future.

Nitrate assimilation pathway is impacted in young tobacco plants overexpressing a constitutively active nitrate reductase or displaying a defective CLCNt2

BackgroundWe have previously shown that the expression of a constitutively active nitrate reductase variant and the suppression of CLCNt2 gene function (belonging to the chloride channel (CLC) gene family) in field-grown tobacco reduces tobacco-specific nitrosamines (TSNA) accumulation in cured leaves and cigarette smoke. In both cases, TSNA reductions resulted from a strong diminution of free nitrate in the leaf, as nitrate is a precursor of the TSNA-producing nitrosating agents formed during tobacco curing and smoking. These nitrosating agents modify tobacco alkaloids to produce TSNAs, the most problematic of which are NNN (N-nitrosonornicotine) and NNK (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone). The expression of a deregulated nitrate reductase enzyme (DNR) that is no longer responsive to light regulation is believed to diminish free nitrate pools by immediately channeling incoming nitrate into the nitrate assimilation pathway. The reduction in nitrate observed when the two tobacco gene copies encoding the vacuolar nitrate transporter CLCNt2 were down-regulated by RNAi-mediated suppression or knocked out using the CRISPR-Cas technology was mechanistically distinct; likely attributable to the inability of the tobacco cell to efficiently sequester nitrate into the vacuole where this metabolite is protected from further assimilation. In this study, we used transcriptomic and metabolomic analyses to compare the nitrate assimilation response in tobacco plants either expressing DNR or lacking CLCNt2 function.ResultsWhen grown in a controlled environment, both DNR and CLCNt2-KO (CLCKO) plants exhibited (1) reduced nitrate content in the leaf; (2) increased N-assimilation into the amino acids Gln and Asn; and (3) a similar pattern of differential regulation of several genes controlling stress responses, including water stress, and cell wall metabolism in comparison to wild-type plants. Differences in gene regulation were also observed between DNR and CLCKO plants, including genes encoding nitrite reductase and asparagine synthetase.ConclusionsOur data suggest that even though both DNR and CLCKO plants display common characteristics with respect to nitrate assimilation, cellular responses, water stress, and cell wall remodeling, notable differences in gene regulatory patterns between the two low nitrate plants are also observed. These findings open new avenues in using plants fixing more nitrogen into amino acids for plant improvement or nutrition perspectives.

New-to-nature CO2-dependent acetyl-CoA assimilation enabled by an engineered B12-dependent acyl-CoA mutase

Acetyl-CoA is a key metabolic intermediate and the product of various natural and synthetic one-carbon (C1) assimilation pathways. While an efficient conversion of acetyl-CoA into other central metabolites, such as pyruvate, is imperative for high biomass yields, available aerobic pathways typically release previously fixed carbon in the form of CO2. To overcome this loss of carbon, we develop a new-to-nature pathway, the Lcm module, in this study. The Lcm module provides a direct link between acetyl-CoA and pyruvate, is shorter than any other oxygen-tolerant route and notably fixes CO2, instead of releasing it. The Lcm module relies on the new-to-nature activity of a coenzyme B12-dependent mutase for the conversion of 3-hydroxypropionyl-CoA into lactyl-CoA. We demonstrate Lcm activity of the scaffold enzyme 2-hydroxyisobutyryl-CoA mutase from Bacillus massiliosenegalensis, and further improve catalytic efficiency 10-fold by combining in vivo targeted hypermutation and adaptive evolution in an engineered Escherichia coli selection strain. Finally, in a proof-of-principle, we demonstrate the complete Lcm module in vitro. Overall, our work demonstrates a synthetic CO2-incorporating acetyl-CoA assimilation route that expands the metabolic solution space of central carbon metabolism, providing options for synthetic biology and metabolic engineering.

Metabolic regulation of 5-oxoproline for enhanced heat tolerance in perennial ryegrass

Pyroglutamic acid [(5-oxoproline (5-oxp)], a non-protein amino acid, can be converted to glutamate to regulate amino acid metabolism in plants. Its roles in plant adaptation to abiotic stresses, including heat stress, are not well understood. The objectives of this study were to determine whether exogenous application of 5-oxp could promote heat tolerance in cool-season perennial grass species and identify the major metabolic pathways that could be activated or responsive to 5-oxp for enhancing heat tolerance. Perennial ryegrass (Lolium perenne L.) plants were foliar-sprayed with 5-oxp or water (untreated control) prior to and during the exposure to heat stress (35/33 ℃, day/night temperature) or ambient temperature (25/22 ℃, day/night temperature, non-stress control) in controlled-environment growth chambers. Application of 5-oxp improved the heat tolerance of perennial ryegrass, as manifested by the chlorophyll content, photochemical efficiency, cell membrane stability, and antioxidant enzyme activities increasing by 31.2%, 25.7%, 37.2%, and 57.1-258.3%, as well as the reduction in hydrogen peroxide production by 36.8%. Metabolic profiling identified metabolites up-regulated by 5-oxp that are involved in the metabolic pathways of carbon assimilation in photosynthesis, glycolysis and the tricarboxylic acid cycle of respiration, proteinogenic amino acid metabolism, glutathione metabolism, and nucleotide metabolism for DNA or RNA synthesis and ATP generation. The up-regulation or activation of those metabolic processes could contribute to 5-oxp-mediated enhancement in the heat tolerance of perennial ryegrass.

The microbial phosphorus cycle in aquatic ecosystems.

Phosphorus is an essential element for life, and phosphorus cycling is crucial to planetary habitability. In aquatic environments, microorganisms are a major component of phosphorus cycling and rapidly transform the diverse chemical forms of phosphorus through various uptake, assimilation and release pathways. Recent discoveries have revealed a more dynamic and complex aquatic microbial phosphorus cycle than previously understood. Some microorganisms have been shown to use and produce new phosphorus compounds, including those in reduced forms. New findings have also raised numerous unanswered questions that warrant further investigation. There is an expanding influence of human activity on aquatic ecosystems. Advancements in understanding the phosphorus biogeochemistry of evolving aquatic environments offer a unique opportunity to comprehend, anticipate and mitigate the effect of human activities. In this Review, I discuss the wealth of new aquatic phosphorus cycle research, spanning disciplines from omics and physiology to global biogeochemical modelling, with a focus on the current comprehension of how aquatic microorganisms sense, transport, assimilate, store, produce and release phosphorus. Of note, I delve into cellular phosphorus allocation, an underexplored topic with wide-ranging implications for energy and element flux in aquatic ecosystems.

Role of membrane H+ transport and plasmalemma excitability in pattern formation, long-distance transport and photosynthesis of Characean algae

Illuminated giant cells of Characeae comprise alternating areas with H+ pump activity and zones with high conductivity for H+/OH–, which create counter-directed H+ flows between the medium and the cytoplasm. In areas where H+ enters the cell, the pH on the surface (pHo) increases to pH 10, while the cytoplasmic pH (pHc) decreases. The lack of the permeant substrate of photosynthesis (CO2) and the acidic pHc shift in the region of external alkaline zones redirect electron transport in chloroplasts from CO2-dependent (assimilatory) pathway to O2 reduction. This electron transport route is associated with an increase in thylakoid membrane ΔpH and an enhanced nonphotochemical quenching (NPQ) of chlorophyll excitations, which underlies strict coordination between nonuniform distributions of pHo and photosynthetic activity in resting cells. When the action potential (AP) is generated, the longitudinal pH profile is temporarily smoothed out, while the heterogeneity of the distribution of NPQ and PSII photochemical activity (YII) sharply increases. The damping of the pHo profile is due to the suppression of the H+ pump and passive H+ conductance under the influence of an almost 100-fold increase in the cytoplasmic of Ca2+ level ([Ca2+]c) during AP. The increase in [Ca2+]c stimulates photoreduction of O2 in chloroplasts under external alkaline zones and, at the same time, arrests the cytoplasmic streaming, which causes the accumulation of excess amounts of H2O2 in the cytoplasm in areas of intense production of this metabolite, with a weak effect on areas of CO2 assimilation. These changes enhance the nonuniform distribution of cell photosynthesis and account for the long-term oscillations of chlorophyll fluorescence Fm' and the quantum efficiency of linear electron flow in microscopic cell areas after the AP generation.

Genomic insights into key mechanisms for carbon, nitrogen, and phosphate assimilation by the acidophilic, halotolerant genus Acidihalobacter members.

In-depth comparative genomic analysis was conducted to predict carbon, nitrogen, and phosphate assimilation pathways in the halotolerant, acidophilic genus Acidihalobacter. The study primarily aimed to understand how the metabolic capabilities of each species can determine their roles and effects on the microbial ecology of their unique saline and acidic environments, as well as in their potential application to saline water bioleaching systems. All four genomes encoded the genes for the complete tricarboxylic acid cycle, including 2-oxoglutarate dehydrogenase, a key enzyme absent in obligate chemolithotrophic acidophiles. Genes for a unique carboxysome shell protein, csoS1D, typically found in halotolerant bacteria but not in acidophiles, were identified. All genomes contained lactate and malate utilization genes, but only A. ferrooxydans DSM 14175T contained genes for the metabolism of propionate. Genes for phosphate assimilation were present, though organized differently across species. Only A. prosperus DSM 5130T and A. aeolianus DSM 14174T genomes contained nitrogen fixation genes, while A. ferrooxydans DSM 14175T and A. yilgarnensis DSM 105917T possessed genes for urease transporters and respiratory nitrate reductases, respectively. The findings suggest that all species can fix carbon dioxide but can also potentially utilize exogenous carbon sources and that the non-nitrogen-fixing species rely on alternative nitrogen assimilation mechanisms.

Response and adaptation of Chlorella pyrenoidosa to 6PPD: Physiological and genetic mechanisms

The extensive contamination of the tire antidegradant N-(1,3-dimethylbutyl)-N′-phenyl-p-phenylenediamine (6PPD) in aquatic environments have raised concerns about its potential threats to aquatic organisms. Here, the responses of green algae Chlorella pyrenoidosa (C. pyrenoidosa) to 6PPD exposure were investigated for the first time. The growth of C. pyrenoidosa experienced three sequential phases, including inhibition, recovery and stimulation. Physiological and transcriptome analysis suggested that the growth inhibition was associated with the suppressed nitrogen assimilation and amino acid biosynthesis pathways, among which nitrate transporter (NRT) 2.1 was a key target of 6PPD. Molecular docking revealed the steadily binding of 6PPD to the substrate entry region of NRT 2.1 via hydrogen bonds and π − cation interaction, blocking the acquisition of extracellular inorganic nitrogen. Along with the removal of 6PPD through abiotic processes and biodegradation, an adaptive metabolic shift in cells not only facilitated growth recovery but also triggered a compensatory stimulation phase. With regard to microalgal adaptation, upregulated DNA replication and repair pathways served to maintain the integrity of the genetic information, enhanced photosynthesis cascades and central carbon metabolism improved carbon flux and energy conversion to microalgal biomass, recovered amino acid biosynthesis produced essential proteins for multiple metabolisms. The results provide new insights into microalgal molecular responses to 6PPD exposure, facilitating a better understanding of ecological consequences of 6PPD in the environment.

Taxonomic and functional partitioning of Chloroflexota populations under ferruginous conditions at and below the sediment-water interface.

The adaptation of the phylum Chloroflexota to various geochemical conditions is thought to have originated in primitive microbial ecosystems, involving hydrogenotrophic energy conservation under ferruginous anoxia. Oligotrophic deep waters displaying anoxic ferruginous conditions, such as those of Lake Towuti, and their sediments may thus constitute a preferential ecological niche for investigating metabolic versatility in modern Chloroflexota. Combining pore water geochemistry, cell counts, sulfate reduction rates, and 16S rRNA genes with in-depth analysis of metagenome-assembled genomes, we show that Chloroflexota benefit from cross-feeding on metabolites derived from canonical respiration chains and fermentation. Detailing their genetic contents, we provide molecular evidence that Anaerolineae have metabolic potential to use unconventional electron acceptors, different cytochromes, and multiple redox metalloproteins to cope with oxygen fluctuations, and thereby effectively colonizing the ferruginous sediment-water interface. In sediments, Dehalococcoidia evolved to be acetogens, scavenging fatty acids, haloacids, and aromatic acids, apparently bypassing specific steps in carbon assimilation pathways to perform energy-conserving secondary fermentations combined with CO2 fixation via the Wood-Ljungdahl pathway. Our study highlights the partitioning of Chloroflexota populations according to alternative electron acceptors and donors available at the sediment-water interface and below. Chloroflexota would have developed analogous primeval features due to oxygen fluctuations in ancient ferruginous ecosystems.

Trans-aconitic acid assimilation system as a widespread bacterial mechanism for environmental adaptation.

The ability of bacteria to use natural carbon sources greatly affects their growth and survival in the environment. Bacteria have evolved versatile abilities to use environmental carbon sources, but their diversity and assimilation pathways remain largely unexplored. Trans-aconitic acid, a geometric isomer of cis-aconitic acid involved in the tricarboxylic acid cycle, has long been considered a natural carbon source metabolizable by bacteria. However, its catabolism and ecological role in linking bacterial interactions with the environments remain unclear. Here, we identify a regulatory system in Bacillus velezensis FZB42 that is capable of sensing and catabolizing trans-aconitic acid. The system consists of a tar operon, an adjacent positive regulatory gene tarR, and a shared promoter. After receiving the trans-aconitic acid signal, the TarR protein interacts directly with the promoter, initiating the expression of the membrane transporter TarB and aconitate isomerase TarA encoded by the operon, which function in importing the trans-aconitic acid and isomerizing it into the central intermediate cis-aconitic acid. Subsequent soil colonization experiments reveal that trans-aconitic acid assimilating ability can give its coding bacteria a growth and competitive advantage. Bioinformatics analyses coupled with bacterial isolation experiments further show that the assimilation system of trans-aconitic acid is widely distributed in the bacterial domain, and its assimilating bacteria also extensively distributed in nature, indicating an important role of trans-aconitic acid metabolism in bacterial carbon acquisition. This work emphasizes the importance of metabolic adaptation to environmental carbon sources for bacterial survival and may provide inspiration for engineering microbes with enhanced environmental competitiveness.

An acidophilic fungus promotes prey digestion in a carnivorous plant.

Leaves of the carnivorous sundew plants (Drosera spp.) secrete mucilage that hosts microorganisms, but whether this microbiota contributes to prey digestion is unclear. We identified the acidophilic fungus Acrodontium crateriforme as the dominant species in the mucilage microbial communities, thriving in multiple sundew species across the global range. The fungus grows and sporulates on sundew glands as its preferred acidic environment, and its presence in traps increased the prey digestion process. A. crateriforme has a reduced genome similar to other symbiotic fungi. During A. crateriforme-Drosera spatulata coexistence and digestion of prey insects, transcriptomes revealed significant gene co-option in both partners. Holobiont expression patterns during prey digestion further revealed synergistic effects in several gene families including fungal aspartic and sedolisin peptidases, facilitating prey digestion in leaves, as well as nutrient assimilation and jasmonate signalling pathway expression. This study establishes that botanical carnivory is defined by adaptations involving microbial partners and interspecies interactions.

Transcriptional Downregulation of Methanol Metabolism Key Genes During Yeast Death in Engineered Pichia pastoris.

Pichia pastoris possesses the unique ability to utilize methanol as its sole carbon source, which makes it a proper host for producing various high-value-added products via metabolic engineering. Nevertheless, cell death has been observed during the fermentation of modified P. pastoris, with limited literature elucidating the underlying causes and mechanisms. Understanding the death mechanisms during methanol-based fermentation is crucial for optimizing fermentation strategies, enhancing the accumulation of target products, and reducing production costs. Here, we first sought to eliminate the potential causes of cell death during fermentation, such as inadequate inorganic salts and toxic by-product accumulation. The elimination of these potential causes was achieved efficiently utilizing the high-throughput fermentation equipment. Subsequently, we established a correlation between yeast cell death and the duration of the methanol metabolism period by monitoring the growth of the yeast at different fermentation stages. A critical revelation from this work came from analyzing the yeast's transcriptomic data at various stages of methanol metabolism. It was observed that a significant characteristic of yeast cell death during fermentation was the marked down-regulation of transcript levels of key enzymes involved in the methanol assimilation pathway and genes related to their biosynthesis process. The findings of this work are crucial for better understanding the causes and mechanisms of cell death for engineered P. pastoris during methanol-utilized fermentation.

Inhibition of sulfur assimilation by S-benzyl-L-cysteine: Impacts on growth, photosynthesis, and leaf proteome of maize plants

Sulfur is an essential nutrient for various physiological processes, including protein synthesis and enzyme activation. We aimed to evaluate how S-benzyl-L-cysteine (SBC), an inhibitor of the sulfur assimilation pathway, affects maize plants' growth, photosynthesis, and leaf proteomic profile. Thus, maize plants were grown for 14 days in vermiculite supplemented with SBC. Photosynthesis was assessed using light and CO2 response curves and chlorophyll a fluorescence. Leaf proteome analysis was conducted to evaluate photosynthetic protein biosynthesis, and ROS content was quantified to assess oxidative stress. Applying SBC resulted in a significant decrease in the growth of maize plants. The gas exchange analysis revealed that maize plants exhibited a diminished rate of CO2 assimilation attributable to both stomatal and non-stomatal limitations. Furthermore, SBC suppressed the activity of important elements involved in the photosynthetic electron transport chain (including photosystems I and II, cytochrome b6f, and ATP synthase) and enzymes responsible for the Calvin cycle, some of which have sulfur-containing prosthetic groups. Consequently, the diminished electron flow rate resulted in a substantial increase in the levels of ROS within the leaves. Our research highlights the crucial role of SBC in disrupting maize photosynthesis by limiting L-cysteine and assimilated sulfur availability, which are essential for the synthesis of protein and prosthetic groups and photosynthetic processes, emphasizing the potential of OAS-TL as a new herbicide site of action.

Enhanced S-adenosyl-L-methionine synthesis in Saccharomyces cerevisiae using metabolic engineering strategies

S-adenosyl-L-methionine (SAM) plays pivotal roles in various physiological processes. With its increasing application in the treatment of diseases such as liver disease, depression, osteoarthritis and Alzheimer's, interest in SAM production aroused. Currently, Saccharomyces cerevisiae is the main industrial producer of SAM. With the surge in demand for SAM, improving the SAM biosynthesis is of importance. In this study, a multimodule engineering strategy was employed to improving SAM production: 1) Enhancing the gene expression of the sulfur assimilation pathway; 2) Strengthening the metabolic flux of the SAM synthesis pathway; 3) Weakening the SAM degradation pathway; 4) Increasing ATP supply. The resulting engineered mutant SC06 (S. cerevisiae CEN.PK2–1CΔgal80∷Tcyc1-sam2-Pgal1-Pgal10-met14-Tadh1, Δlsc2∷Tcyc1-hom6-Pgal1-Pgal10-met6-Tadh1, Δsah1Δmls1) displayed the highest SAM titer of 240.86 mg/L, which was 10.22-fold increase compared with the original strain. With optimized conditions, the SAM titer of mutant SC06 in shake flask fermentation reached 473.02 mg/L with a specific yield of 127.18 mg/g dry cell weight (DCW). In a 5 L fermenter with fed-batch fermentation, the maximal SAM yield of mutant SC06 reached 1.25 g/L with a specific yield of 166.67 mg/g DCW after 58 h cultivation. Therefore, the established metabolic engineering strategies displayed promising efficiency in improving the SAM productivity of S. cerevisiae CEN.PK2–1C, which may provide a useful tool for the improvement of SAM-producing strains.

Boreal moss-microbe interactions are revealed through metagenome assembly of novel bacterial species

Moss-microbe interactions contribute to ecosystem processes in boreal forests. Yet, how host-specific characteristics and the environment drive the composition and metabolic potential of moss microbiomes is still poorly understood. In this study, we use shotgun metagenomics to identify the taxonomy and metabolic potential of the bacteria of four moss species of the boreal forests of Northern Québec, Canada. To characterize moss bacterial community composition and diversity, we assembled the genomes of 110 potentially novel bacterial species. Our results highlight that moss genus, species, gametophyte section, and to a lesser extent soil pH and soil temperature, drive moss-associated bacterial community composition and diversity. In the brown gametophyte section, two Stigonema spp. showed partial pathway completeness for photosynthesis and nitrogen fixation, while all brown-associated Hyphomicrobiales had complete assimilatory nitrate reduction pathways and many nearly complete carbon fixation pathways. Several brown-associated species showed partial to complete pathways for coenzyme M and F420 biosynthesis, important for methane metabolism. In addition, green-associated Hyphomicrobiales (Methylobacteria spp.) displayed potential for the anoxygenic photosystem II pathway. Overall, our findings demonstrate how host-specific characteristics and environmental factors shape the composition and metabolic potential of moss bacteria, highlighting their roles in carbon fixation, nitrogen cycling, and methane metabolism in boreal forests.

Enhanced nutrients removal from low C/N ratio rural sewage by embedding heterotrophic nitrifying bacteria and activated alumina in a tidal flow constructed wetland

Rural sewage treatment facilitates nitrogen and phosphorus removal yet can be costly. To address this challenge, a cost-effective embedding material mainly consisting of heterotrophic nitrifying bacteria, activated alumina (AA), and a solid carbon source (HPMC) was applied to a tidal flow constructed wetlands (TFCWs); aimed at stable nitrogen and phosphorus removal under low carbon-to-nitrogen (C/N) ratios. The TFCWs could be shortened to 16 d of startup duration time compared with the control group; and improved the ammonia nitrogen (NH4+-N), total nitrogen (TN), and total phosphorus (TP) removal efficiencies to 98 %, 93 %, and 68 %, respectively. Also, effluent NH4+-N, TN, and TP in the enhanced TFCWs could be stable at 0.52 ± 0.18, 1.23 ± 0.45, and 0.75 ± 0.25 mg/L, respectively. Microbial community analysis revealed that AA and HPMC were enriched Pseudomonas sp., which potentially accelerated the NH4+-N assimilation pathway and phosphate biological removal. Embedding materials-TFCWs can provide new solutions for integrated rural sewage technology.