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The indica and japonica rice ( Oryza sativa ) subspecies differ in nitrate (NO 3 − ) assimilation capacity and nitrogen (N) use efficiency (NUE). Here, we show that a major component of this difference is conferred by allelic variation at OsNR2 , a gene encoding a NADH/NADPH-dependent NO 3 − reductase (NR). Selection-driven allelic divergence has resulted in variant indica and japonica OsNR2 alleles encoding structurally distinct OsNR2 proteins, with indica OsNR2 exhibiting greater NR activity. Indica OsNR2 also promotes NO 3 − uptake via feed-forward interaction with OsNRT1.1B , a gene encoding a NO 3 − uptake transporter. These properties enable indica OsNR2 to confer increased effective tiller number, grain yield and NUE on japonica rice, effects enhanced by interaction with an additionally introgressed indica OsNRT1.1B allele. In consequence, indica OsNR2 provides an important breeding resource for the sustainable increases in japonica rice yields necessary for future global food security. Indica rice has higher nitrate assimilation and nitrogen use efficiency (NUE) than japonica rice, but the mechanism is unclear. Here, the authors reveal that the difference is partly due to allelic variation of a nitrate reductase encoding gene and this indica allele can increase yield potential and NUE.

Algae have reemerged as potential next-generation feedstocks for biofuels, but strain improvement and progress in algal biology research have been limited by the lack of advanced molecular tools for most eukaryotic microalgae. Here we describe the development of an efficient transformation method for Nannochloropsis sp., a fast-growing, unicellular alga capable of accumulating large amounts of oil. Moreover, we provide additional evidence that Nannochloropsis is haploid, and we demonstrate that insertion of transformation constructs into the nuclear genome can occur by high-efficiency homologous recombination. As examples, we generated knockouts of the genes encoding nitrate reductase and nitrite reductase, resulting in strains that were unable to grow on nitrate and nitrate/nitrite, respectively. The application of homologous recombination in this industrially relevant alga has the potential to rapidly advance algal functional genomics and biotechnology.

Plants have evolved adaptive strategies that involve transcriptional networks to cope with and survive environmental challenges. Key transcriptional regulators that mediate responses to environmental fluctuations in nitrate have been identified; however, little is known about how these regulators interact to orchestrate nitrogen (N) responses and cell-cycle regulation. Here we report that teosinte branched1/cycloidea/proliferating cell factor1-20 (TCP20) and NIN-like protein (NLP) transcription factors NLP6 and NLP7, which act as activators of nitrate assimilatory genes, bind to adjacent sites in the upstream promoter region of the nitrate reductase gene, NIA1, and physically interact under continuous nitrate and N-starvation conditions. Regions of these proteins necessary for these interactions were found to include the type I/II Phox and Bem1p (PB1) domains of NLP6&7, a protein-interaction module conserved in animals for nutrient signaling, and the histidine- and glutamine-rich domain of TCP20, which is conserved across plant species. Under N starvation, TCP20-NLP6&7 heterodimers accumulate in the nucleus, and this coincides with TCP20 and NLP6&7-dependent up-regulation of nitrate assimilation and signaling genes and down-regulation of the G₂/M cell-cycle marker gene, CYCB1;1. TCP20 and NLP6&7 also support root meristem growth under N starvation. These findings provide insights into how plants coordinate responses to nitrate availability, linking nitrate assimilation and signaling with cell-cycle progression.

Dissimilatory nitrate reduction to ammonia (DNRA) is a common biochemical process in the nitrogen cycle in natural and man-made habitats, but its significance in wastewater treatment plants is not well understood. Several ammonifying Trichlorobacter strains (former Geobacter ) were previously enriched from activated sludge in nitrate-limited chemostats with acetate as electron ( e ) donor, demonstrating their presence in these systems. Here, we isolated and characterized the new species Trichlorobacter ammonificans strain G1 using a combination of low redox potential and copper-depleted conditions. This allowed purification of this DNRA organism from competing denitrifiers. T. ammonificans is an extremely specialized ammonifier, actively growing only with acetate as e -donor and carbon source and nitrate as e -acceptor, but H 2 can be used as an additional e -donor. The genome of G1 does not encode the classical ammonifying modules NrfAH/NrfABCD. Instead, we identified a locus encoding a periplasmic nitrate reductase immediately followed by an octaheme cytochrome c that is conserved in many Geobacteraceae species. We purified this octaheme cytochrome c protein (TaNiR), which is a highly active dissimilatory ammonifying nitrite reductase loosely associated with the cytoplasmic membrane. It presumably interacts with two ferredoxin subunits (NapGH) that donate electrons from the menaquinol pool to the periplasmic nitrate reductase (NapAB) and TaNiR. Thus, the Nap-TaNiR complex represents a novel type of highly functional DNRA module. Our results indicate that DNRA catalyzed by octaheme nitrite reductases is a metabolic feature of many Geobacteraceae , representing important community members in various anaerobic systems, such as rice paddy soil and wastewater treatment facilities.

To gain a better insight into the selenium nanoparticle (nSe) benefits/toxicity, this experiment was carried out to address the behavior of bitter melon seedlings to nSe (0, 1, 4, 10, 30, and 50 mgL-1) or bulk form (selenate). Low doses of nSe increased biomass accumulation, while concentrations of 10 mgL-1 and above were associated with stem bending, impaired root meristem, and severe toxicity. Responses to nSe were distinct from that of bulk in that the nano-type exhibited a higher efficiency to stimulate growth and organogenesis than the bulk. The bulk form displayed higher phytotoxicity than the nano-type counterpart. According to the MSAP-based analysis, nSe mediated substantial variation in DNA cytosine methylation, reflecting the epigenetic modification. By increasing the concentration of nSe, the expression of the WRKY1 transcription factor linearly up-regulated (mean = 7.9-fold). Transcriptions of phenylalanine ammonia-lyase (PAL) and 4-Coumarate: CoA-ligase (4CL) genes were also induced. The nSe treatments at low concentrations enhanced the activity of leaf nitrate reductase (mean = 52%) in contrast with the treatment at toxic concentrations. The toxic concentration of nSe increased leaf proline concentration by 80%. The nSe supplement also stimulated the activities of peroxidase (mean = 35%) and catalase (mean = 10%) enzymes. The nSe-treated seedlings exhibited higher PAL activity (mean = 39%) and soluble phenols (mean = 50%). The nSe toxicity was associated with a disrupted differentiation of xylem conducting tissue. The callus formation and performance of the explants originated from the nSe-treated seedlings had a different trend than that of the control. This experiment provides new insights into the nSe-associated advantage/ cytotoxicity and further highlights the necessity of designing convincing studies to introduce novel methods for plant cell/tissue cultures and agriculture.

Bacteria of the SAR11 clade constitute up to one half of all microbial cells in the oxygen-rich surface ocean. SAR11 bacteria are also abundant in oxygen minimum zones (OMZs), where oxygen falls below detection and anaerobic microbes have vital roles in converting bioavailable nitrogen to N 2 gas. Anaerobic metabolism has not yet been observed in SAR11, and it remains unknown how these bacteria contribute to OMZ biogeochemical cycling. Here, genomic analysis of single cells from the world’s largest OMZ revealed previously uncharacterized SAR11 lineages with adaptations for life without oxygen, including genes for respiratory nitrate reductases (Nar). SAR11 nar genes were experimentally verified to encode proteins catalysing the nitrite-producing first step of denitrification and constituted ~40% of OMZ nar transcripts, with transcription peaking in the anoxic zone of maximum nitrate reduction activity. These results link SAR11 to pathways of ocean nitrogen loss, redefining the ecological niche of Earth’s most abundant organismal group. Bacteria of the SAR11 clade constitute up to one half of all marine microbes and are thought to require oxygen for growth; here, a subgroup of SAR11 bacteria are shown to thrive in ocean oxygen minimum zones and to encode abundant respiratory nitrate reductases. An anoxic niche for SAR11 bacteria SAR11 bacteria, the most abundant type of microbe in the world's oceans, are thought to require oxygen for growth, yet they are also abundant in waters where oxygen levels are low. Frank Stewart and colleagues show here that a subgroup of SAR11 bacteria that thrives in ocean oxygen minimum zones have adapted to the microaerobic/anaerobic conditions there, and they encode abundant respiratory nitrate reductases that perform the first step in denitrification. These results redefine the ecological niche of Earth's most abundant organismal group and suggest that they are substantial contributors to nitrogen loss in oxygen minimum zones.

Small interfering RNAs (siRNAs) are essential for proper development and immunity in eukaryotes 1 . Plants produce siRNAs with lengths of 21, 22 or 24 nucleotides. The 21- and 24-nucleotide species mediate cleavage of messenger RNAs and DNA methylation 2 , 3 , respectively, but the biological functions of the 22-nucleotide siRNAs remain unknown. Here we report the identification and characterization of a group of endogenous 22-nucleotide siRNAs that are generated by the DICER-LIKE 2 (DCL2) protein in plants. When cytoplasmic RNA decay and DCL4 are deficient, the resulting massive accumulation of 22-nucleotide siRNAs causes pleiotropic growth disorders, including severe dwarfism, meristem defects and pigmentation. Notably, two genes that encode nitrate reductases— NIA1 and NIA2 —produce nearly half of the 22-nucleotide siRNAs. Production of 22-nucleotide siRNAs triggers the amplification of gene silencing and induces translational repression both gene specifically and globally. Moreover, these 22-nucleotide siRNAs preferentially accumulate upon environmental stress, especially those siRNAs derived from NIA1/2 , which act to restrain translation, inhibit plant growth and enhance stress responses. Thus, our research uncovers the unique properties of 22-nucleotide siRNAs, and reveals their importance in plant adaptation to environmental stresses. Characterization of 22-nucleotide short interfering RNAs in plants finds that they accumulate in response to environmental stress, causing translational repression, inhibition of plant growth and enhanced stress responses.

Diatoms are unicellular algae that accumulate significant amounts of triacylglycerols as storage lipids when their growth is limited by nutrients. Using biochemical, physiological, bioinformatics, and reverse genetic approaches, we analyzed how the flux of carbon into lipids is influenced by nitrogen stress in a model diatom, Phaeodactylum tricornutum . Our results reveal that the accumulation of lipids is a consequence of remodeling of intermediate metabolism, especially reactions in the tricarboxylic acid and the urea cycles. Specifically, approximately one-half of the cellular proteins are cannibalized; whereas the nitrogen is scavenged by the urea and glutamine synthetase/glutamine 2-oxoglutarate aminotransferase pathways and redirected to the de novo synthesis of nitrogen assimilation machinery, simultaneously, the photobiological flux of carbon and reductants is used to synthesize lipids. To further examine how nitrogen stress triggers the remodeling process, we knocked down the gene encoding for nitrate reductase, a key enzyme required for the assimilation of nitrate. The strain exhibits 40–50% of the mRNA copy numbers, protein content, and enzymatic activity of the wild type, concomitant with a 43% increase in cellular lipid content. We suggest a negative feedback sensor that couples photosynthetic carbon fixation to lipid biosynthesis and is regulated by the nitrogen assimilation pathway. This metabolic feedback enables diatoms to rapidly respond to fluctuations in environmental nitrogen availability. Significance When starved for nutrients, diatoms redirect carbon toward biosynthesis of storage lipids, triacylglycerols (TAGs). We examined how this modification is achieved in the diatom Phaeodactylum tricornutum. Under nitrogen stress, the cells cannibalized their photosynthetic apparatus while recycling intracellular nitrogen and redirecting it to synthesize nitrogen assimilation enzymes. Simultaneously, they allocated newly fixed carbon toward lipids. In contrast, a nitrate reductase knocked-down strain shunted ∼40% more carbon toward TAGs than the wild type without losing photosynthetic capacity. Our results show that diatoms can remodel their intermediate metabolism on environmental cues and reveal that a key signal in this remodeling is associated with nitrogen assimilation. This insight informs a strategy of developing a much more efficient pathway to produce algal-based biofuels.

Lelliottia amnigena PTJIIT1005 is a bacterium that utilizes nitrate as the sole nitrogen source and can remediate nitrate from media . The annotation was done related to nitrogen metabolic genes using the PATRIC, RAST tools, and PGAP from the genome sequence of this bacterium. Multiple sequence alignments and phylogenetic analysis of respiratory nitrate reductase, assimilatory nitrate reductase, nitrite reductase, glutamine synthetase, hydroxylamine reductase, nitric oxide reductase genes from PTJIIT1005 were done to find out sequence identities with the most similar species. The identification of operon arrangement in bacteria was also identified. The PATRIC KEGG feature mapped the N-metabolic pathway to identify the chemical process, and the 3D structure of representative enzymes was also elucidated. The putative protein 3D structure was analyzed using I-TASSER software. It gave good quality protein models of all nitrogen metabolism genes and showed good sequence identity with reference templates, approximately 81–99%, except for two genes; assimilatory nitrate reductase and nitrite reductase. This study suggested that PTJIIT1005 can remove N-nitrate from water because of having N-assimilation and denitrification genes.

In sweet potato, rational nitrogen (N) assimilation and distribution are conducive to inhibiting vine overgrowth. Nitrate (NO 3 - ) is the main N form absorbed by roots, and cultivar is an important factor affecting N utilization. Herein, a hydroponic experiment was conducted that included four NO 3 - concentrations of 0 (N0), 4 (N1), 8 (N2) and 16 (N3) mmol L -1 with two cultivars of Jishu26 (J26, N-sensitive) and Xushu32 (X32, N-tolerant). For J26, with increasing NO 3 - concentrations, the root length and root surface area significantly decreased. However, no significant differences were observed in these parameters for X32. Higher NO 3 - concentrations upregulated the expression levels of the genes that encode nitrate reductase ( NR2 ), nitrite reductase ( NiR2 ) and nitrate transporter ( NRT1.1 ) in roots for both cultivars. The trends in the activities of NR and NiR were subject to regulation of NR2 and NiR2 transcription, respectively. For both cultivars, N2 increased the N accumulated in leaves, growth points and roots. For J26, N3 further increased the N accumulation in these organs. Under higher NO 3 - nutrition, compared with X32, J26 exhibited higher expression levels of the NiR2 , NR2 and NRT1.1 genes, a higher influx NO 3 - rate in roots, and higher activities of NR and NiR in leaves and roots. Conclusively, the regulated effects of NO 3 - supplies on root growth and NO 3 - utilization were more significant for J26. Under high NO 3 - conditions, J26 exhibited higher capacities of NO 3 - absorption and distributed more N in leaves and in growth points, which may contribute to higher growth potential in shoots and more easily cause vine overgrowth.