3-Nitropropionic acid (3-NPA) is a toxic nitroaliphatic compound, but the genomic basis of bacterial adaptation to 3-NPA remains incompletely resolved, especially in organisms lacking a canonical nitronate monooxygenase. Here, we used phenotype screening, whole-genome resequencing, targeted UHPLC-MS/MRM, and composite RNA-seq profiling to investigate 3-NPA utilization in Leclercia barmai EMC7. Following attempted Tn5-Mob mutagenesis and kanamycin-based recovery, two independently derived mutants, LTM01 and LTM14, showed impaired growth under 3-NPA-dependent minimal conditions. No stable transposon insertion was detected in either mutant. Comparative genome analysis instead revealed convergent loss-of-function (LOF) mutations affecting amino acid metabolism, nitrogen allocation, central carbon metabolism, cofactor-linked functions, transport, and metal-redox homeostasis, while the predicted flavin-dependent nitro-redox candidate and canonical nitrate/nitrite reduction loci were not disrupted. Targeted UHPLC-MS/MRM analysis showed that wild-type EMC7 depleted approximately 50% of 3-NPA within 36 h, whereas the mutants displayed poor 3- NPA-utilization phenotypes under the same growth framework. Wild-type RNA-seq under glucose-3NPA relative to glucose-KNO3 identified a broad expression profile involving flavin-redox functions, nitrogen assimilation, oxidative-stress response, envelope stress, efflux, cofactor metabolism, and iron homeostasis. Integration of mutant LOF profiles with wild-type gene expression data showed that several disrupted genes or functional counterparts belonged to modules showing expression changes during the wild-type glucose-3NPA response. These findings identify candidate adaptive modules associated with 3-NPA utilization in EMC7, including nitrogen redistribution, carbon entry, sulfur-redox support, cofactor supply, transport, envelope remodeling, and metal-redox control. These results extend the landscape of metabolic adaptation beyond canonical pathways in environmental bacteria.
Sucrose non-fermenting-1-related protein kinase (SnRK) is a plant serine/threonine kinase that mediates stress signaling, yet its function in the mangrove Avicennia marina remains unexplored. In this study, we identified 46 SnRK genes in A. marina, classifying them into three subfamilies with high phylogenetic conservation. Family expansion occurred mainly through 20 segmental duplication events under purifying selection (Ka/Ks < 1). Promoter regions were enriched in stress- and hormone-responsive elements, and expression profiling showed distinct tissue-specific and stress-responsive patterns. Our study specifically focused on the functional characterization of AmSnRK2.7. AmSnRK2.7-overexpressed Arabidopsis thaliana increases salt tolerance by altering the expression of ion transport genes and mediating Na+ efflux from the roots. Further investigation revealed that AmSnRK2.7 interacts with and phosphorylates AmENO2 (enolase, a key enzyme in the glycolysis pathway). Meanwhile, measurements of ATP content in wild type, AmSnRK2.7-overexpressed Arabidopsis lines and the atsnrk2.6 (homologous gene of AmSnRK2.7 in A. thaliana) mutant confirm that the AmSnRK2.7-AmENO2 module can affect the energy-driven Na+ efflux. Our findings provide key insights into the role of SnRK gene family in mangrove adaptation to saline intertidal habitat, and suggest AmSnRK2.7-AmENO2 module plays a role in salt tolerance.
Jin-Yu Liu, Yudan Xiang, Lingyu Song et al.· Plant, Cell and Environment· 0 citations
GLORI sequencing is applied to generate single-base resolution transcriptome-wide m6A maps in seven bacterial species to provide a quantitative atlas of bacterial m6A and establish a foundation for understanding its regulatory and evolutionary roles.
Youyue Li, Letong Xu, Na Liu et al.· Cell Reports· 0 citations
ABSTRACT The regulatory landscape responsible for lignocellulose degradation in white-rot basidiomycete fungi remains largely unexplored. In this study, we characterize a novel transcriptional activator, Bhr1, in the white-rot fungus Dichomitus squalens. Bhr1 exhibits an unusual domain architecture that combines a septin-like P-loop NTPase fold with Zn(II)2Cys6 DNA-binding domains and plays a critical role in activating (hemi-)cellulase enzyme production when D. squalens is exposed to mannose-rich substrates. Using CRISPR/Cas9-mediated gene editing, we generated a bhr1 disruption mutant that displayed distinct phenotypes and enzyme activity profiles on mannose and guar gum compared to the wild type. RNA sequencing data indicate that Bhr1 induces specific (hemi-)cellulase-encoding genes without altering the expression of genes encoding sugar transporters or sugar metabolic enzymes. Phylogenetic analyses show that Bhr1 is basidiomycete specific and largely restricted to saprotrophic and plant-associated Agaricomycetes fungi. Based on the domain architecture of Bhr1 and the effects of its disruption in D. squalens, our findings reveal a lineage-specific regulatory innovation in basidiomycetes that is distinct from those described in ascomycetes. Elucidating the function and evolutionary conservation of Bhr1 advances our understanding of lignocellulose degradation at the molecular level in basidiomycete fungi and may inform studies of their ecological adaptation and the development of biotechnological applications. IMPORTANCE Understanding the transcriptional regulatory mechanisms in white-rot fungi, such as Dichomitus squalens, is crucial for advancing our knowledge of lignocellulose degradation. This study identifies D. squalens Bhr1 as a key regulator of (hemi-)cellulase production on mannose-rich substrates and further distinguishes basidiomycete transcription factors involved in plant biomass degradation from their ascomycete counterparts. Our findings highlight the significance of lineage-specific regulators in facilitating adaptive enzyme production for efficient biomass utilization, which is critical to carbon cycling in terrestrial ecosystems. This work establishes a foundation for exploring novel regulatory strategies among wood-degrading fungi, potentially enabling targeted strain engineering in biotechnological applications. Understanding the transcriptional regulatory mechanisms in white-rot fungi, such as Dichomitus squalens, is crucial for advancing our knowledge of lignocellulose degradation. This study identifies D. squalens Bhr1 as a key regulator of (hemi-)cellulase production on mannose-rich substrates and further distinguishes basidiomycete transcription factors involved in plant biomass degradation from their ascomycete counterparts. Our findings highlight the significance of lineage-specific regulators in facilitating adaptive enzyme production for efficient biomass utilization, which is critical to carbon cycling in terrestrial ecosystems. This work establishes a foundation for exploring novel regulatory strategies among wood-degrading fungi, potentially enabling targeted strain engineering in biotechnological applications.
Victor M. Gonzalez Ramos, A. Lipzen, Hope Hundley et al.· Microbiology spectrum· 0 citations
A novel component of the maize nitrogen-signaling network is uncovered and provide a candidate gene for further functional studies on low-nitrogen adaptation in maize.
ABSTRACT Nitrous oxide (N2O) is a potent greenhouse gas and ozone-depleting substance, with a global warming potential 273 times greater than CO2 over a 100-year horizon. Microbial reduction of N2O to dinitrogen represents a key pathway for mitigating emissions under diverse environmental conditions. Here, we report the first comprehensive genomic and physiological characterization of Stutzerimonas frequens strain E49, a newly isolated N2O-reducing bacterium obtained from activated sludge treating landfill leachate. Whole-genome sequencing revealed a 4.51-Mbp circular chromosome and a 35.3-kbp plasmid with high completeness. Functional annotation identified a complete denitrification gene set, including nosZ, as well as the ectABCD-ask gene cluster associated with ectoine biosynthesis, suggesting adaptation to osmotic stress. Strain E49, a uniform rod-shaped bacterium (1.5–2.5 μm), efficiently reduced N2O under anaerobic conditions in the absence of an externally supplied organic carbon source, achieving a biomass-specific rate of 0.70 ± 0.02 µmol-N2O/mg-biomass/h and a cell-specific rate of 7.93 ± 0.23 × 10−10 µmol-N2O/cell/h. Among the cultivation regimens tested, nitrate-free DSMZ 1180 medium supplemented with NH4Cl yielded the highest activity, indicating medium-dependent regulation of N2O respiration. Comparative analysis showed that strain E49 outperformed several reported N2O-reducing isolates under carbon-limited conditions. These findings demonstrate the metabolic versatility of strain E49 and highlight its potential role as a biological sink for N2O in low-nutrient environments. IMPORTANCE Nitrous oxide is a powerful greenhouse gas that contributes to climate change and ozone depletion. Microorganisms that convert nitrous oxide into nitrogen gas play an essential role in reducing these emissions. In this study, we investigated Stutzerimonas frequens strain E49, a bacterium isolated from wastewater treatment sludge. We found that this organism can efficiently reduce nitrous oxide even without an external supply of organic carbon, which is typically required by most bacteria. This suggests that the bacterium can rely on internal energy reserves to carry out this process. We also identified the genetic basis for its nitrous oxide reduction and its ability to adapt to environmental stress. These findings improve our understanding of how nitrous oxide-reducing bacteria function in nutrient-limited environments and may support the development of strategies to mitigate emissions in wastewater treatment and other engineered systems. Nitrous oxide is a powerful greenhouse gas that contributes to climate change and ozone depletion. Microorganisms that convert nitrous oxide into nitrogen gas play an essential role in reducing these emissions. In this study, we investigated Stutzerimonas frequens strain E49, a bacterium isolated from wastewater treatment sludge. We found that this organism can efficiently reduce nitrous oxide even without an external supply of organic carbon, which is typically required by most bacteria. This suggests that the bacterium can rely on internal energy reserves to carry out this process. We also identified the genetic basis for its nitrous oxide reduction and its ability to adapt to environmental stress. These findings improve our understanding of how nitrous oxide-reducing bacteria function in nutrient-limited environments and may support the development of strategies to mitigate emissions in wastewater treatment and other engineered systems.
