Engineering Papers⌕ Search

SEARCH · Engineering Papers

Results for “Functional genomics”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,135 records · Page 63
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Respiratory Vinyl Chloride Reductive Dechlorination to Ethene in TceA-Expressing Dehalococcoides mccartyi

Bioremediation of chlorinated ethenes in anoxic aquifers hinges on organohalide-respiring Dehalococcoidia expressing vinyl chloride (VC) reductive dehalogenase (RDase). The tceA gene encoding the trichloroethene-dechlorinating RDase TceA is frequently detected in contaminated groundwater but not recognized as a biomarker for VC detoxification. We demonstrate that tceA-carrying Dehalococcoides mccartyi (Dhc) strains FL2 and 195 grow with VC as an electron acceptor when sufficient vitamin B 12 (B 12 ) is provided. Strain FL2 cultures that received 50 μg L –1 B 12 completely dechlorinated VC to ethene at rates of 14.80 ± 1.30 μM day –1 and attained 1.64 ± 0.11 × 10 8 cells per μmol of VC consumed. Strain 195 attained similar growth yields of 1.80 ± 1.00 × 10 8 cells per μmol of VC consumed, and both strains could be consecutively transferred with VC as the electron acceptor. Proteomic analysis demonstrated TceA expression in VC-grown strain FL2 cultures. Resequencing of the strain FL2 and strain 195 tceA genes identified non-synonymous substitutions, although their consequences for TceA function are currently unknown. The finding that Dhc strains expressing TceA respire VC can explain ethene formation at chlorinated solvent sites, where quantitative polymerase chain reaction analysis indicates that tceA dominates the RDase gene pool.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2021

A Century of Synergy in Termite Symbiosis Research: Linking the Past with New Genomic Insights

Termites have long been studied for their symbiotic associations with gut microbes. In the late nineteenth century, this relationship was poorly understood and captured the interest of parasitologists such as Joseph Leidy; this research led to that of twentieth-century biologists and entomologists including Cleveland, Hungate, Trager, and Lüscher. Early insights came via microscopy, organismal, and defaunation studies, which led to descriptions of microbes present, descriptions of the roles of symbionts in lignocellulose digestion, and early insights into energy gas utilization by the host termite. Focus then progressed to culture-dependent microbiology and biochemical studies of host–symbiont complementarity, which revealed specific microhabitat requirements for symbionts and noncellulosic mechanisms of symbiosis (e.g., N 2 fixation). Today, knowledge on termite symbiosis has accrued exponentially thanks to omic technologies that reveal symbiont identities, functions, and interdependence, as well as intricacies of host–symbiont complementarity. Moving forward, the merging of classical twentieth-century approaches with evolving omic tools should provide even deeper insights into host–symbiont interplay.

Entomology↗
DOE OSTI2026

Bioenergy Cropping Reduces the Spatiotemporal Scaling of Soil Bacterial Biodiversity

Widespread bioenergy cropping can transform landscapes, strongly affecting biodiversity. However, the impact of bioenergy cropping on the spatiotemporal scaling of soil biodiversity remains virtually unknown, despite its profound implications for the functioning of the ecological community. Here, we investigated how bioenergy cropping influenced the spatiotemporal scaling of soil bacterial biodiversity in marginal soils (sandy loam and clay loam soils) in Oklahoma, USA. We detected strong, significant species-time-area relationships (STARs) and phylogenetic-time-area relationships (PTARs) in bacterial communities and their lineages, suggesting that STARs and PTARs exist in microbial ecology within the studied system. Also, spatiotemporal scaling rates (the slopes of STAR and PTAR models) varied substantially among bacterial lineages and were positively correlated with their 16S rRNA gene copy numbers, a genomic trait indicative of microbial growth potentials. Strikingly, bioenergy cropping significantly reduced spatiotemporal scaling rates by 6.8%-14.1%, with a more pronounced reduction observed in sandy loam soils, where those rates were significantly lower than in clay loam soils. The heterogeneity of soil phosphorus and carbon resulted in variations in bacterial spatiotemporal scaling rates. Collectively, our findings suggest that bioenergy cropping may alleviate rapid shifts in soil biodiversity across space and time, thereby stabilizing soil biodiversity and supporting its role as part of sustainable land management and climate mitigation strategies.

bacterial diversity↗
DOE OSTI2023

Rapid assembly of SARS-CoV-2 genomes reveals attenuation of the Omicron BA.1 variant through NSP6

Although the SARS-CoV-2 Omicron variant (BA.1) spread rapidly across the world and effectively evaded immune responses, its viral fitness in cell and animal models was reduced. The precise nature of this attenuation remains unknown as generating replication-competent viral genomes is challenging because of the length of the viral genome (~30 kb). Here, we present a plasmid-based viral genome assembly and rescue strategy (pGLUE) that constructs complete infectious viruses or noninfectious subgenomic replicons in a single ligation reaction with >80% efficiency. Fully sequenced replicons and infectious viral stocks can be generated in 1 and 3 weeks, respectively. By testing a series of naturally occurring viruses as well as Delta-Omicron chimeric replicons, we show that Omicron nonstructural protein 6 harbors critical attenuating mutations, which dampen viral RNA replication and reduce lipid droplet consumption. Thus, pGLUE overcomes remaining barriers to broadly study SARS-CoV-2 replication and reveals deficits in nonstructural protein function underlying Omicron attenuation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Multiple Wheat Genomes Reveal Novel Gli-2 Sublocus Location and Variation of Celiac Disease Epitopes in Duplicated α-Gliadin Genes

The seed protein α-gliadin is a major component of wheat flour and causes gluten-related diseases. However, due to the complexity of this multigene family with a genome structure composed of dozens of copies derived from tandem and genome duplications, little was known about the variation between accessions, and thus little effort has been made to explicitly target α-gliadin for bread wheat breeding. Here, we analyzed genomic variation in α-gliadins across 11 recently published chromosome-scale assemblies of hexaploid wheat, with validation using long-read data. We unexpectedly found that the Gli-B2 locus is not a single contiguous locus but is composed of two subloci, suggesting the possibility of recombination between the two during breeding. We confirmed that the number of immunogenic epitopes among 11 accessions varied. The D subgenome of a European spelt line also contained epitopes, in agreement with its hybridization history. Evolutionary analysis identified amino acid sites under diversifying selection, suggesting their functional importance. The analysis opens the way for improved grain quality and safety through wheat breeding.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

The anaerobic fungus Caecomyces churrovis produces H2 via a non-3 bifurcating NADH-dependent enzyme complex

Anaerobic fungi (AF) decompose lignocellulose-based biomass into fermentable sugars through the production of powerful biomass-degrading enzymes. AF are unusual among fungi in that they generate energy via hydrogenosomes, which are also associated with the release of H2 though yet unknown metabolic mechanisms. In particular, it remains unclear how NAD(P)+ is regenerated within hydrogenosomes and how H2 is formed. Here, we reveal the molecular mechanism for hydrogenosomal H2 production in the AF strain C. churrovis by combining genomic search, proteomic analysis, and enzymology. Our enzyme assays on the large organelle fraction of C. churrovis revealed the activity of H2:NAD+ oxidoreductase but not pyruvate:ferredoxin oxidoreductase activity. We identified genes encoding [FeFe] hydrogenase (Hyd) and NADH dehydrogenase subunits E and F (NuoE, NuoF) in C. churrovis, and confirmed their expression in the isolated hydrogenosomal fractions by proteomic analysis. Combining the individually purified proteins, we found that the assay system consisting of Hyd-Strep and NuoEF-Strep reduced NAD+ with H2. Furthermore, this system formed H2 directly from NADH independent of ferredoxin, functioning as a non-bifurcating NADH-dependent enzyme rather than an electron-bifurcating enzyme. We identified homologs of hydrogenosomal NuoE, NuoF, and Hyd in many other AF, indicating this pathway is widely conserved among the early-branching AF. This work demonstrates the existence of a non-bifurcating NADH-dependent enzyme complex in eukaryotes. Moreover, this complex could be a target for controlling AF H2 production and altering fungal metabolism.

fungi↗
DOE OSTI2021

Enhancing transcription in Escherichia coli and Pseudomonas putida using bacteriophage lambda anti-terminator protein Q

Functional characterization of metagenomic DNA often involves expressing heterologous DNA in genetically tractable microorganisms such as Escherichia coli . Functional expression of heterologous genes can suffer from limitations due to the lack of recognition of foreign promoters or presence of intrinsic terminators on foreign DNA between a vector-based promoter and the transcription start site. Anti-terminator proteins are a possible solution to overcome this limitation. When bacteriophage lambda infects E. coli , it relies on the host transcription machinery to transcribe and express phage DNA. Lambda anti-terminator protein Q (λQ) regulates the expression of late-genes of phage lambda. E. coli RNA polymerase recognizes the P R ' promoter on the lambda genome and forms a complex with λQ, to overcome the terminator t R '. In this study we show the use of λQ to efficiently transcribe a capsular polysaccharide cluster, cps3 , from Lactobacillus plantarum containing intrinsic terminators in Escherichia coli . In addition, we expand the use of anti-terminator λQ in Pseudomonas putida . The results show ~ fivefold higher expression of a fluorescent reporter locate ~ 12.5kbp downstream from the promoter, when the transcription is driven by P R ' promoter in presence of λQ compared to a lac promoter. These results suggest that λQ could be used in metabolic engineering to enhance expression of heterologous DNA.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

A spatio-temporally constrained gene regulatory network directed by PBX1/2 acquires limb patterning specificity via HAND2

A lingering question in developmental biology has centered on how transcription factors with widespread distribution in vertebrate embryos can perform tissue-specific functions. Here, using the murine hindlimb as a model, we investigate the elusive mechanisms whereby PBX TALE homeoproteins, viewed primarily as HOX cofactors, attain context-specific developmental roles despite ubiquitous presence in the embryo. We first demonstrate that mesenchymal-specific loss of PBX1/2 or the transcriptional regulator HAND2 generates similar limb phenotypes. By combining tissue-specific and temporally controlled mutagenesis with multi-omics approaches, we reconstruct a gene regulatory network (GRN) at organismal-level resolution that is collaboratively directed by PBX1/2 and HAND2 interactions in subsets of posterior hindlimb mesenchymal cells. Genome-wide profiling of PBX1 binding across multiple embryonic tissues further reveals that HAND2 interacts with subsets of PBX-bound regions to regulate limb-specific GRNs. Our research elucidates fundamental principles by which promiscuous transcription factors cooperate with cofactors that display domain-restricted localization to instruct tissue-specific developmental programs.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Evidence for a cytokine-sensitive network of iron-associated genes that protects pancreatic islets against ferroptosis

Background/Objectives: The micronutrient iron is closely connected to inflammation and is among the complex factors contributing to beta-cell failure in diabetes. High levels of dietary iron increase the risk of developing type 2 diabetes, and excessive iron uptake by beta-cells can cause oxidative stress and inhibit function. Elevated levels of proinflammatory cytokines in obese individuals, such as interleukin (IL)-1beta and IL-6, increase the risk of developing type 2 diabetes, and there is evidence that these low levels of circulating cytokines can lead to islet dysfunction. Methods: In this study, gene microarray and other data were analyzed for expression differences in islets treated for 48 h with 10 pg/mL IL-1beta + 20 pg/mL IL-6 as a model of low-grade inflammation versus untreated. Results: Three iron-associated genes were among the most cytokine-sensitive in the mouse genome: Hamp, Steap4, and Lcn2. These proteins are all involved with increasing/retaining cellular iron. We hypothesized that increased cellular iron would lead to increased susceptibility to ferroptosis. Surprisingly, 24 h pre-exposure to low-grade inflammation, which upregulates this iron-gene network, prevented subsequent erastin-induced ferroptosis. We also found that Steap4 overexpression reduced islet dysfunction caused by high-dose proinflammatory cytokines (10× low-dose), suggesting an overall protective effect. Steap4 overexpression also upregulated Hamp and Lcn2, suggesting Steap4 regulates these cytokine-sensitive iron genes.; in contrast, ferritin and ferroportin gene expression, which are not sensitive to cytokines, were unchanged. Conclusions: These data suggest an inflammation-induced network of genes involved in cellular iron uptake and retention plays a protective role in islets against oxidative stress and ferroptosis.

IL-1β↗
DOE OSTI2024

Functional plasticity of HCO 3 – uptake and CO 2 fixation in Cupriavidus necator H16

Despite its prominence, the ability to engineer Cupriavidus necator H16 for inorganic carbon uptake and fixation is underexplored. We tested the roles of endogenous and heterologous genes on C. necator inorganic carbon metabolism. Deletion of β-carbonic anhydrase can had the most deleterious effect on C. necator autotrophic growth. Replacement of this native uptake system with several classes of dissolved inorganic carbon (DIC) transporters from Cyanobacteria and chemolithoautotrophic bacteria recovered autotrophic growth and supported higher cell densities compared to wild-type (WT) C. necator in batch culture. Strains expressing Halothiobacillus neopolitanus DAB2 (hnDAB2) and diverse rubisco homologs grew in CO 2 similarly to the wild-type strain. Our experiments suggest that the primary role of carbonic anhydrase during autotrophic growth is to support anaplerotic metabolism, and an array of DIC transporters can complement this function. This work demonstrates flexibility in HCO 3 - uptake and CO 2 fixation in C. necator, providing new pathways for CO 2 -based biomanufacturing.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

The impact of G-quadruplex dynamics on inter-tetrad electronic couplings: a hybrid computational study

The G-quadruplex is a fascinating nucleic acid motif with implications in biology, medicine, and nanotechnologies. G-quadruplexes can form in the telomeres at the edges of chromosomes and in other guanine-rich regions of the genome. They can also be engineered for exploitation as biological materials for nanodevices. Their higher stiffness and higher charge transfer rates make them better candidates in nanodevices than duplex DNA. For the development of molecular nanowires, it is important to optimize electron transport along the wire axis. One powerful basis to do so is by manipulating the structure, based on known effects that structural changes have on electron transport. Here, for this work, we investigate such effects, by a combination of classical simulations of the structure and dynamics and quantum calculations of electronic couplings. We find that this structure–function relationship is complex. A single helix shape parameter alone does not embody such complexity, but rather a combination of distances and angles between stacked bases influences charge transfer efficiency. By analyzing linear combinations of shape descriptors for different topologies, we identify the structural features that most affect charge transfer efficiency. We discuss the transferability of the proposed model and the limiting effects of inherent flexibility.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2024

Unified understanding of the impact of semiflexibility, concentration, and molecular weight on macromolecular-scale ring diffusion

Conformationally fluctuating, globally compact macromolecules such as polymeric rings, single-chain nanoparticles, microgels, and many-arm stars display complex dynamic behaviors due to their rich topological structure and intermolecular organization. Synthetic rings are hybrid objects with conformations that display both ideal random walk and compact globular features, which can serve as models of genomic DNA. To date, emphasis has been placed on the effect of ring molecular weight on their unusual behaviors. Here, we combine simulations and a microscopic force-level theory to build a unified understanding for how key aspects of ring dynamics depend on different tunable molecular properties including backbone rigidity, monomer concentration, degree of traditional entanglement, and molecular weight. Our large-scale molecular dynamics simulations of ring melts with very different backbone stiffnesses reveal unanticipated behaviors which agree well with our generalized theory. This includes a universal master curve for center-of-mass diffusion constants as a function of molecular weight scaled by a chemistry and thermodynamic state-dependent critical molecular weight that generalizes the concept of an entanglement cross-over for linear chains. The key physics is how backbone rigidity and monomer concentration induced changes of the entanglement length, interring packing, degree of interpenetration, and liquid compressibility slow down space-time dynamic-force correlations on macromolecular scales. A power law decay of the center-of-mass diffusion constant with inverse molecular weight squared is the first consequence, followed by an ultraslow activated hopping transport regime. Our results set the stage to address slow dynamics and kinetic arrest in different families of compact synthetic and biological polymeric systems.

Science & Technology - Other Topics↗
DOE OSTI2022

Expression and structure of the Chlamydia trachomatis DksA ortholog

Chlamydia trachomatis is a bacterial obligate intracellular parasite and a significant cause of human disease, including sexually transmitted infections and trachoma. The bacterial RNA polymerase-binding protein DksA is a transcription factor integral to the multicomponent bacterial stress response pathway known as the stringent response. The genome of C. trachomatis encodes a DksA ortholog (DksA Ct ) that is maximally expressed at 15–20 h post infection, a time frame correlating with the onset of transition between the replicative reticulate body (RB) and infectious elementary body (EB) forms of the pathogen. Ectopic overexpression of DksA Ct in C. trachomatis prior to RB–EB transitions during infection of HeLa cells resulted in a 39.3% reduction in overall replication (yield) and a 49.6% reduction in recovered EBs. While the overall domain organization of DksA Ct is similar to the DksA ortholog of Escherichia coli (DksA Ec ), DksA Ct did not functionally complement DksA Ec . Transcription of dksA Ct is regulated by tandem promoters, one of which also controls expression of nrdR, encoding a negative regulator of deoxyribonucleotide biosynthesis. The phenotype resulting from ectopic expression of DksA Ct and the correlation between dksA Ct and nrdR expression is consistent with a role for DksA Ct in the C. trachomatis developmental cycle.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

CRISPR–Cas9-mediated nuclear transport and genomic integration of nanostructured genes in human primary cells

DNA nanostructures are a promising tool to deliver molecular payloads to cells. DNA origami structures, where long single-stranded DNA is folded into a compact nanostructure, present an attractive approach to package genes; however, effective delivery of genetic material into cell nuclei has remained a critical challenge. Here, we describe the use of DNA nanostructures encoding an intact human gene and a fluorescent protein encoding gene as compact templates for gene integration by CRISPR-mediated homology-directed repair (HDR). Our design includes CRISPR–Cas9 ribonucleoprotein binding sites on DNA nanostructures to increase shuttling into the nucleus. We demonstrate efficient shuttling and genomic integration of DNA nanostructures using transfection and electroporation. These nanostructured templates display lower toxicity and higher insertion efficiency compared to unstructured double-stranded DNA templates in human primary cells. Furthermore, our study validates virus-like particles as an efficient method of DNA nanostructure delivery, opening the possibility of delivering nanostructures in vivo to specific cell types. Together, these results provide new approaches to gene delivery with DNA nanostructures and establish their use as HDR templates, exploiting both their design features and their ability to encode genetic information. This work also opens a door to translate other DNA nanodevice functions, such as biosensing, into cell nuclei.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Binding profiles for 961 Drosophila and C. elegans transcription factors reveal tissue-specific regulatory relationships

A catalog of transcription factor (TF) binding sites in the genome is critical for deciphering regulatory relationships. Here, we present the culmination of the efforts of the modENCODE (model organism Encyclopedia of DNA Elements) and modERN (model organism Encyclopedia of Regulatory Networks) consortia to systematically assay TF binding events in vivo in two major model organisms,Drosophila melanogaster(fly) andCaenorhabditis elegans(worm). These data sets comprise 605 TFs identifying 3.6 M sites in the fly and 356 TFs identifying 0.9 M sites in the worm, and represent the majority of the regulatory space in each genome. We demonstrate that TFs associate with chromatin in clusters termed “metapeaks,” that larger metapeaks have characteristics of high-occupancy target (HOT) regions, and that the importance of consensus sequence motifs bound by TFs depends on metapeak size and complexity. Combining ChIP-seq data with single-cell RNA-seq data in a machine-learning model identifies TFs with a prominent role in promoting target gene expression in specific cell types, even differentiating between parent–daughter cells during embryogenesis. These data are a rich resource for the community that should fuel and guide future investigations into TF function. To facilitate data accessibility and utility, all strains expressing green fluorescent protein (GFP)-tagged TFs are available at the stock centers for each organism. The chromatin immunoprecipitation sequencing data are available through the ENCODE Data Coordinating Center, GEO, and through a direct interface that provides rapid access to processed data sets and summary analyses, as well as widgets to probe the cell-type-specific TF–target relationships.

Biochemistry & Molecular Biology↗
DOE OSTI2020

Ubiquity and functional uniformity in CO2 concentrating mechanisms in multiple phyla of Bacteria is suggested by a diversity and prevalence of genes encoding candidate dissolved inorganic carbon transporters

ABSTRACT Autotrophic microorganisms catalyze the entry of dissolved inorganic carbon (DIC; = CO2 + HCO3− + CO32−) into the biological component of the global carbon cycle, despite dramatic differences in DIC abundance and composition in their sometimes extreme environments. “Cyanobacteria” are known to have CO2 concentrating mechanisms (CCMs) to facilitate growth under low CO2 conditions. These CCMs consist of carboxysomes, containing enzymes ribulose 1,5-bisphosphate oxygenase and carbonic anhydrase, partnered to DIC transporters. CCMs and their DIC transporters have been studied in a handful of other prokaryotes, but it was not known how common CCMs were beyond “Cyanobacteria”. Since it had previously been noted that genes encoding potential transporters were found neighboring carboxysome loci, α-carboxysome loci were gathered from bacterial genomes, and potential transporter genes neighboring these loci are described here. Members of transporter families whose members all transport DIC (CHC, MDT and Sbt) were common in these neighborhoods, as were members of the SulP transporter family, many of which transport DIC. 109 of 115 taxa with carboxysome loci have some form of DIC transporter encoded in their genomes, suggesting that CCMs consisting of carboxysomes and DIC transporters are widespread not only among “Cyanobacteria”, but also among members of “Proteobacteria” and “Actinobacteria”.

Scott, Kathleen M.↗
DOE OSTI2026

The anaerobic fungus Caecomyces churrovis produces H 2 via a non-bifurcating NADH-dependent enzyme complex

ABSTRACT Hydrogenosomes are mitochondria-derived organelles that produce ATP and H 2 to support energy metabolism in anaerobic eukaryotes. H 2 production allows reoxidation of reduced cofactors generated during fermentative metabolism; however, the metabolic mechanisms for H 2 production in anaerobic eukaryotes remains incompletely understood. In particular, it remains unclear whether anaerobic fungi (AF) hydrogenosomes use a ferredoxin-dependent pathway or a distinct mechanism to regenerate NAD(P) + and link electron transfer to H 2 formation. Here, by combining genomic search, proteomic analysis, and enzymology, we reveal the molecular mechanism for H 2 production in the AF strain Caecomyces churrovis . Our enzyme assays on the organelle fraction of C. churrovis revealed the activity of H 2 :NAD + oxidoreductase but not pyruvate:ferredoxin oxidoreductase, which is usually linked to H 2 formation. We identified genes encoding [FeFe] hydrogenase (Hyd) and NADH dehydrogenase subunits E and F (NuoE, NuoF) in C. churrovis , and confirmed their expression in the isolated hydrogenosomal fractions by proteomic analysis. Combining the individually purified enzymes, we found Hyd and NuoEF proteins formed H 2 directly from NADH independently of ferredoxin, functioning as a non-bifurcating NADH-dependent enzyme rather than an electron-bifurcating enzyme. We identified homologs of hydrogenosomal NuoE, NuoF, and Hyd in many other AF, indicating this pathway is commonly shared among the AF. This work demonstrates the existence of a non-bifurcating NADH-dependent enzyme complex in eukaryotes. Moreover, this complex could potentially be exploited as a target for controlling AF H 2 production and altering fungal metabolism. IMPORTANCE H 2 production is a prominent feature of anaerobic energy metabolism, yet our understanding of eukaryotic mechanisms remains limited. Anaerobic fungi (AF) are key decomposers of lignocellulose and contribute to hydrogen flux in anaerobic environments. Although it has been more than 40 years since the H 2 production from AF was first reported, the molecular mechanism for hydrogenosomal H 2 production and redox balance remains unclear. We demonstrate that AF produce H 2 from NADH utilizing a non-bifurcating NADH-dependent enzyme complex rather than an electron-bifurcating, ferredoxin-dependent variant. We show that this enzyme complex is conserved across multiple AF lineages and thus demonstrate the occurrence of a non-bifurcating NADH-dependent enzyme in eukaryotes. This discovery expands our understanding of eukaryotic hydrogenosomal metabolism, reveals a previously unknown strategy for redox balancing, and highlights potential targets for manipulating H 2 production. These insights have broad implications for microbial energy metabolism, anaerobic ecosystems, and bioengineering of H 2 -producing systems.

Zhang, Bo [Department of Chemical Engineering, Uni↗
DOE OSTI2022

Validation and functional characterization of transcription factors in wheat using cell-free protein expression and high-throughput sequencing technologies

Transcription factors (TFs) are critical biomolecules that control and regulate gene expression in every organism. In wheat, there are 3,606 genes annotated as putative TFs which are primarily uncharacterized. In this work, using a DAP-seq approach, we have validated and characterized a select number of TFs (5bl and 5dl) belonging to GeBP family of proteins, responsible for plant cellular growth, development, and differentiation. We found that top DNA binding motifs for 5bl and 5dl are (G/T)N(T/G)GTGGT and (C/G)AA(C/G)AA respectively. These motifs/peaks are enriched in the promoter regions of the genes, which are associated with rRNA biogenesis and protein maturation pathways. This study is also a first demonstration that DAP-seq can be applied for such a complex and large genome such as wheat.

59 BASIC BIOLOGICAL SCIENCES↗