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At least 91 records · Page 5
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

Sequencing the Genomes of the First Terrestrial Fungal Lineages: What Have We Learned?

The first genome sequenced of a eukaryotic organism was for Saccharomyces cerevisiae, as reported in 1996, but it was more than 10 years before any of the zygomycete fungi, which are the early-diverging terrestrial fungi currently placed in the phyla Mucoromycota and Zoopagomycota, were sequenced. The genome for Rhizopus delemar was completed in 2008; currently, more than 1000 zygomycete genomes have been sequenced. Genomic data from these early-diverging terrestrial fungi revealed deep phylogenetic separation of the two major clades—primarily plant—associated saprotrophic and mycorrhizal Mucoromycota versus the primarily mycoparasitic or animal-associated parasites and commensals in the Zoopagomycota. Genomic studies provide many valuable insights into how these fungi evolved in response to the challenges of living on land, including adaptations to sensing light and gravity, development of hyphal growth, and co-existence with the first terrestrial plants. Genome sequence data have facilitated studies of genome architecture, including a history of genome duplications and horizontal gene transfer events, distribution and organization of mating type loci, rDNA genes and transposable elements, methylation processes, and genes useful for various industrial applications. Pathogenicity genes and specialized secondary metabolites have also been detected in soil saprobes and pathogenic fungi. Novel endosymbiotic bacteria and viruses have been discovered during several zygomycete genome projects. Overall, genomic information has helped to resolve a plethora of research questions, from the placement of zygomycetes on the evolutionary tree of life and in natural ecosystems, to the applied biotechnological and medical questions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Widespread adenine N6-methylation of active genes in fungi

N6-methyldeoxyadenine (6mA) is a noncanonical DNA base modification present at low levels in plant and animal genomes, but its prevalence and association with genome function in other eukaryotic lineages remains poorly understood. We report that abundant 6mA is associated with transcriptionally active genes in early-diverging fungal lineages. Using single-molecule long-read sequencing of 16 diverse fungal genomes, we observed that up to 2.8% of all adenines were methylated in early-diverging fungi, far exceeding levels observed in other eukaryotes and more derived fungi. 6mA occurred symmetrically at ApT dinucleotides and was concentrated in dense methylated adenine clusters surrounding the transcriptional start sites of expressed genes; its distribution was inversely correlated with that of 5-methylcytosine. Our results show a striking contrast in the genomic distributions of 6mA and 5-methylcytosine and reinforce a distinct role for 6mA as a gene-expression-associated epigenomic mark in eukaryotes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2018

Blocking amino acid transporter Os AAP 3 improves grain yield by promoting outgrowth buds and increasing tiller number in rice

Summary Amino acid transporters ( AAT s) play indispensable roles in nutrient allocation during plant development. In this study, we demonstrated that inhibiting expression of the rice amino acid transporter Os AAP 3 increased grain yield due to a formation of larger numbers of tillers as a result of increased bud outgrowth. Elevated expression of Os AAP 3 in transgenic plants resulted in significantly higher amino acid concentrations of Lys, Arg, His, Asp, Ala, Gln, Gly, Thr and Tyr, and inhibited bud outgrowth and rice tillering. However, RNA i of Os AAP 3 decreased significantly Arg, Lys, Asp and Thr concentrations to a small extent, and thus promoted bud outgrowth, increased significantly tiller numbers and effective panicle numbers per plant, and further enhanced significantly grain yield and nitrogen use efficiency ( NUE ). The promoter sequences of Os AAP 3 showed some divergence between Japonica and Indica rice, and expression of the gene was higher in Japonica , which produced fewer tillers than Indica . We generated knockout lines of Os AAP 3 on Japonica ZH 11 and KY 131 using CRISPR technology and found that grain yield could be increased significantly. These results suggest that manipulation of Os AAP 3 expression could be used to increase grain yield in rice.

Lu, Kai↗
NASA NTRS1994

Characterization of a new thermophilic sulfate-reducing bacterium Thermodesulfovibrio yellowstonii, gen. nov. and sp. nov.: its phylogenetic relationship to Thermodesulfobacterium commune and their origins deep within the bacterial domain

A thermophilic sulfate-reducing vibrio isolated from thermal vent water in Yellowstone Lake, Wyoming, USA is described. The gram-negative, curved rod-shaped cells averaged 0.3 micrometer wide and 1.5 micrometers long. They were motile by means of a single polar flagellum. Growth was observed between 40 degrees and 70 degrees C with optimal growth at 65 degrees C. Cultures remained viable for one year at 27 degrees C although spore-formation was not observed. Sulfate, thiosulfate and sulfite were used as electron acceptors. Sulfur, fumarate and nitrate were not reduced. In the presence of sulfate, growth was observed only with lactate, pyruvate, hydrogen plus acetate, or formate plus acetate. Pyruvate was the only compound observed to support fermentative growth. Pyruvate and lactate were oxidized to acetate. Desulfofuscidin and c-type cytochromes were present. The G + C content was 29.5 mol%. The divergence in the 16 S ribosomal RNA sequences between the new isolate and Thermodesulfobacterium commune suggests that these two thermophilic sulfate-reducing bacteria represent different genera. These two bacteria depict a lineage that branches deeply within the Bacteria domain and which is clearly distinct from previously defined phylogenetic lines of sulfate-reducing bacteria. Strain YP87 is described as the type strain of the new genus and species Thermodesulfovibrio yellowstonii.

Non-NASA Center↗
DOE OSTI2023

Whole-genome sequencing distinguishes the two most common giant kelp ecomorphs

Abstract Giant kelp, Macrocystis pyrifera, exists as distinct morphological variants—or “ecomorphs”—in different populations, yet the mechanism for this variation is uncertain, and environmental drivers for either adaptive or plastic phenotypes have not been identified. The ecomorphs Macrocystis “pyrifera” and M. “integrifolia” are distributed throughout temperate waters of North and South America with almost no geographic overlap and exhibit an incongruous, non-mirrored, distribution across the equator. This study evaluates the degree of genetic divergence between M. “pyrifera” and M. “integrifolia” across 18 populations in Chile and California using whole-genome sequencing and single-nucleotide polymorphism markers. Our results based on a principal component analysis, admixture clustering by genetic similarity, and phylogenetic inference demonstrate that M. “pyrifera” and M. “integrifolia” are genetically distinguishable. Analyses reveal separation by Northern and Southern Hemispheres and between morphs within hemispheres, suggesting that the convergent “integrifolia” morphology arose separately in each hemisphere. This is the first study to use whole-genome sequencing to understand genetic divergence in giant kelp ecomorphs, identifying 83 potential genes under selection and providing novel insights about Macrocystis evolution that were not evident with previous genetic techniques. Future studies are needed to uncover the environmental forces driving local adaptation and presumed convergent evolution of these morphs.

Environmental Sciences & Ecology↗
DOE OSTI2019

Genome of Alaskapox Virus, A Novel Orthopoxvirus Isolated from Alaska

Since the eradication of smallpox, there have been increases in poxvirus infections and the emergence of several novel poxviruses that can infect humans and domestic animals. In 2015, a novel poxvirus was isolated from a resident of Alaska. Diagnostic testing and limited sequence analysis suggested this isolate was a member of the Orthopoxvirus (OPXV) genus but was highly diverged from currently known species, including Akhmeta virus. Here, we present the complete 210,797 bp genome sequence of the Alaska poxvirus isolate, containing 206 predicted open reading frames. Phylogenetic analysis of the conserved central region of the genome suggested the Alaska isolate shares a common ancestor with Old World OPXVs and is diverged from New World OPXVs. We propose this isolate as a member of a new OPXV species, Alaskapox virus (AKPV). The AKPV genome contained host range and virulence genes typical of OPXVs but lacked homologs of C4L and B7R, and the hemagglutinin gene contained a unique 120 amino acid insertion. Seven predicted AKPV proteins were most similar to proteins in non-OPXV Murmansk or NY_014 poxviruses. Genomic analysis revealed evidence suggestive of recombination with Ectromelia virus in two putative regions that contain seven predicted coding sequences, including the A-type inclusion protein.

60 APPLIED LIFE SCIENCES↗
NASA NTRS1991

Archaebacterial rhodopsin sequences: Implications for evolution

It was proposed over 10 years ago that the archaebacteria represent a separate kingdom which diverged very early from the eubacteria and eukaryotes. It follows that investigations of archaebacterial characteristics might reveal features of early evolution. So far, two genes, one for bacteriorhodopsin and another for halorhodopsin, both from Halobacterium halobium, have been sequenced. We cloned and sequenced the gene coding for the polypeptide of another one of these rhodopsins, a halorhodopsin in Natronobacterium pharaonis. Peptide sequencing of cyanogen bromide fragments, and immuno-reactions of the protein and synthetic peptides derived from the C-terminal gene sequence, confirmed that the open reading frame was the structural gene for the pharaonis halorhodopsin polypeptide. The flanking DNA sequences of this gene, as well as those of other bacterial rhodopsins, were compared to previously proposed archaebacterial consensus sequences. In pairwise comparisons of the open reading frame with DNA sequences for bacterio-opsin and halo-opsin from Halobacterium halobium, silent divergences were calculated. These indicate very considerable evolutionary distance between each pair of genes, even in the dame organism. In spite of this, three protein sequences show extensive similarities, indicating strong selective pressures.

Lanyi, J. K.↗
DOE OSTI2023

Co-crystal structures of the fluorogenic aptamer Beetroot show that close homology may not predict similar RNA architecture

Beetroot is a homodimeric in vitro selected RNA that binds and activates DFAME, a conditional fluorophore derived from GFP. It is 70% sequence-identical to the previously characterized homodimeric aptamer Corn, which binds one molecule of its cognate fluorophore DFHO at its interprotomer interface. We have now determined the Beetroot-DFAME co-crystal structure at 1.95 Å resolution, discovering that this RNA homodimer binds two molecules of the fluorophore, at sites separated by ~30 Å. In addition to this overall architectural difference, the local structures of the non-canonical, complex quadruplex cores of Beetroot and Corn are distinctly different, underscoring how subtle RNA sequence differences can give rise to unexpected structural divergence. Through structure-guided engineering, we generated a variant that has a 12-fold fluorescence activation selectivity switch toward DFHO. Beetroot and this variant form heterodimers and constitute the starting point for engineered tags whose through-space inter-fluorophore interaction could be used to monitor RNA dimerization.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Different tertiary interactions create the same important 3D features in a distinct flavivirus xrRNA

During infection by a flavivirus (FV), cells accumulate noncoding subgenomic flavivirus RNAs (sfRNAs) that interfere with several antiviral pathways. These sfRNAs are formed by structured RNA elements in the 3' untranslated region (UTR) of the viral genomic RNA, which block the progression of host cell exoribonucleases that have targeted the viral RNA. Previous work on these exoribonuclease-resistant RNAs (xrRNAs) from mosquito-borne FVs revealed a specific three-dimensional fold with a unique topology in which a ring-like structure protectively encircles the 5' end of the xrRNA. Conserved nucleotides make specific tertiary interactions that support this fold. Examination of more divergent FVs reveals differences in their 3' UTR sequences, raising the question of whether they contain xrRNAs and if so, how they fold. To answer this, we demonstrated the presence of an authentic xrRNA in the 3' UTR of the Tamana bat virus (TABV) and solved its structure by X-ray crystallography. The structure reveals conserved features from previously characterized xrRNAs, but in the TABV version these features are created through a novel set of tertiary interactions not previously seen in xrRNAs. This includes two important A–C interactions, four distinct backbone kinks, several ordered Mg 2+ ions, and a C + –G–C base triple. The discovery that the same overall architecture can be achieved by very different sequences and interactions in distantly related flaviviruses provides insight into the diversity of this type of RNA and will inform searches for undiscovered xrRNAs in viruses and beyond.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Reactivity, Regioselectivity, and Synthetic Application of 2-Pyrenyl Units in Scholl Reactions

We herein report the reactivity and regioselectivity of 2-pyrenyl as a coupling unit in Scholl reactions. On the basis of the Scholl reactions of hexaarylbenzene substrates, we have found that pyrenyl units are preferably oxidized over naphthyl and phenyl units under appropriate Scholl reaction conditions, allowing divergent synthesis through a highly controllable intramolecular coupling sequence. We find that the C1 and C3 positions of the 2-pyrenyl unit are the favorable sites for intramolecular coupling while C4 is not reactive to allow further coupling. The reactivity and regioselectivity pattern can be explained by the spin density distribution, which shows that carbon–carbon bonds preferably form at sites with higher positive spin density. Guided by these findings, we successfully synthesized a double helicene and a sextuple helicene through the controlled Scholl reactions of 2-pyrenyl units.

Scholl reactions↗
DOE OSTI2016

Single sample resolution of rare microbial dark matter in a marine invertebrate metagenome

Direct, untargeted sequencing of environmental samples (metagenomics) and de novo genome assembly enable the study of uncultured and phylogenetically divergent organisms. However, separating individual genomes from a mixed community has often relied on the differential-coverage analysis of multiple, deeply sequenced samples. In the metagenomic investigation of the marine bryozoan Bugula neritina, we uncovered seven bacterial genomes associated with a single B. neritina individual that appeared to be transient associates, two of which were unique to one individual and undetectable using certain “universal” 16S rRNA primers and probes. We recovered high quality genome assemblies for several rare instances of “microbial dark matter,” or phylogenetically divergent bacteria lacking genomes in reference databases, from a single tissue sample that was not subjected to any physical or chemical pre-treatment. One of these rare, divergent organisms has a small (593 kbp), poorly annotated genome with low GC content (20.9%) and a 16S rRNA gene with just 65% sequence similarity to the closest reference sequence. Lastly, our findings illustrate the importance of sampling strategy and de novo assembly of metagenomic reads to understand the extent and function of bacterial biodiversity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Convergent evolution of aerobic fermentation through divergent mechanisms acting on the same key glycolytic genes

Expression profiling by high throughput sequencing The model yeast Saccharomyces cerevisiae has evolved aerobic fermentation in part through a whole genome duplication and subsequent retention of glycolytic genes. To identify independent yeast lineages with a high glycolytic rate phenotype we developed an assay measuring extracellular acidification rates (ECAR) to evaluate the glycolytic rates across diverse yeast species. This assay identified a novel group of yeasts in the genus Saturnispora with rapid glycolytic rates. Through comparative approaches, we found that several glycolytic genes encoding hexokinases and enzymes in lower glycolysis had higher expression and modifications in promoter sequences in rapid ECAR species. Intriguingly, many of the upregulated genes are the same genes that are duplicated in S. cerevisiae. When the transcription factor required for their activation was deleted, the mutants had a slow glycolytic rate and more respiratory phenotype. This work shows how the aerobic fermentation phenotype has convergently evolved in Saturnispora through transcriptional rewiring. This divergent mechanism nevertheless impacts the same glycolytic genes, which suggests that there are evolutionary constraints on how aerobic fermentation can arise.

Horianopoulos, Linda C. [Department of Food Scienc↗
DOE OSTI2019

Ancient polymorphisms contribute to genome-wide variation by long-term balancing selection and divergent sorting in Boechera stricta

Genomic variation is widespread, and both neutral and selective processes can generate similar patterns in the genome. These processes are not mutually exclusive, so it is difficult to infer the evolutionary mechanisms that govern population and species divergence. Boechera stricta is a perennial relative of Arabidopsis thaliana native to largely undisturbed habitats with two geographic and ecologically divergent subspecies. Here, we delineate the evolutionary processes driving the genetic diversity and population differentiation in this species. Using whole-genome re-sequencing data from 517 B. stricta accessions, we identify four genetic groups that diverged around 30-180 thousand years ago, with long-term small effective population sizes and recent population expansion after the Last Glacial Maximum. We find three genomic regions with elevated nucleotide diversity, totaling about 10% of the genome. These three regions of elevated nucleotide diversity show excess of intermediate-frequency alleles, higher absolute divergence ( d XY ), and lower relative divergence ( F ST ) than genomic background, and significant enrichment in immune-related genes, reflecting long-term balancing selection. Scattered across the genome, we also find regions with both high F ST and d XY among the groups, termed F ST -islands. Population genetic signatures indicate that F ST -islands with elevated divergence, which have experienced directional selection, are derived from divergent sorting of ancient polymorphisms. Our results suggest that long-term balancing selection on disease resistance genes may have maintained ancestral haplotypes across different geographical lineages, and unequal sorting of balanced polymorphisms may have generated genomic regions with elevated divergence. This study highlights the importance of ancestral balanced polymorphisms as crucial components of genome-wide variation.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1972

Sequence data - Magnitude and implications of some ambiguities.

A stochastic model is applied to the divergence of the horse-pig lineage from a common ansestor in terms of the alpha and beta chains of hemoglobin and fibrinopeptides. The results are compared with those based on the minimum mutation distance model of Fitch (1972). Buckwheat and cauliflower cytochrome c sequences are analyzed to demonstrate their ambiguities. A comparative analysis of evolutionary rates for various proteins of horses and pigs shows that errors of considerable magnitude are introduced by Glx and Asx ambiguities into evolutionary conclusions drawn from sequences of incompletely analyzed proteins.

Holmquist, R.↗
DOE OSTI2020

Potent CRISPR-Cas9 inhibitors from Staphylococcus genomes

Anti-CRISPRs (Acrs) are small proteins that inhibit the RNA-guided DNA targeting activity of CRISPR-Cas enzymes. Encoded by bacteriophage and phage-derived bacterial genes, Acrs prevent CRISPR-mediated inhibition of phage infection and can also block CRISPR-Cas-mediated genome editing in eukaryotic cells. To identify Acrs capable of inhibiting Staphylococcus aureus Cas9 (SauCas9), an alternative to the most commonly used genome editing protein Streptococcus pyogenes Cas9 (SpyCas9), we used both self-targeting CRISPR screening and guilt-by-association genomic search strategies. Here we describe three potent inhibitors of SauCas9 that we name AcrIIA13, AcrIIA14, and AcrIIA15. These inhibitors share a conserved N-terminal sequence that is dispensable for DNA cleavage inhibition and have divergent C termini that are required in each case for inhibition of SauCas9-catalyzed DNA cleavage. In human cells, we observe robust inhibition of SauCas9-induced genome editing by AcrIIA13 and moderate inhibition by AcrIIA14 and AcrIIA15. We also find that the conserved N-terminal domain of AcrIIA13–AcrIIA15 binds to an inverted repeat sequence in the promoter of these Acr genes, consistent with its predicted helix-turn-helix DNA binding structure. These data demonstrate an effective strategy for Acr discovery and establish AcrIIA13–AcrIIA15 as unique bifunctional inhibitors of SauCas9.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Gene flow and introgression are pervasive forces shaping the evolution of bacterial species

Abstract Background Although originally thought to evolve clonally, studies have revealed that most bacteria exchange DNA. However, it remains unclear to what extent gene flow shapes the evolution of bacterial genomes and maintains the cohesion of species. Results Here, we analyze the patterns of gene flow within and between >2600 bacterial species. Our results show that fewer than 10% of bacterial species are truly clonal, indicating that purely asexual species are rare in nature. We further demonstrate that the taxonomic criterion of ~95% genome sequence identity routinely used to define bacterial species does not accurately represent a level of divergence that imposes an effective barrier to gene flow across bacterial species. Interruption of gene flow can occur at various sequence identities across lineages, generally from 90 to 98% genome identity. This likely explains why a ~95% genome sequence identity threshold has empirically been judged as a good approximation to define bacterial species. Our results support a universal mechanism where the availability of identical genomic DNA segments required to initiate homologous recombination is the primary determinant of gene flow and species boundaries in bacteria. We show that these barriers of gene flow remain porous since many distinct species maintain some level of gene flow, similar to introgression in sexual organisms. Conclusions Overall, bacterial evolution and speciation are likely shaped by similar forces driving the evolution of sexual organisms. Our findings support a model where the interruption of gene flow—although not necessarily the initial cause of speciation—leads to the establishment of permanent and irreversible species borders.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Different evolutionary pathways of HIV-1 between fetus and mother perinatal transmission pairs indicate unique immune selection in fetuses

Study of evolution and selection pressure on HIV-1 in fetuses will lead to a better understanding of the role of immune responses in shaping virus evolution and vertical transmission. Detailed genetic analyses of HIV-1 env gene from 12 in utero transmission pairs show that most infections (67%) occur within 2 months of childbirth. In addition, the env sequences from long-term-infected fetuses are highly divergent and form separate phylogenetic lineages from their cognate maternal viruses. Host-selection sites unique to neonate viruses are identified in regions frequently targeted by neutralizing antibodies and T cell immune responses. Identification of unique selection sites in the env gene of fetal viruses indicates that the immune system in fetuses is capable of exerting selection pressure on viral evolution. Studying selection and evolution of HIV-1 or other viruses in fetuses can be an alternative approach to investigate adaptive immunity in fetuses.

60 APPLIED LIFE SCIENCES↗