The reference genome and abiotic stress responses of the model perennial grass Brachypodium sylvaticum.

Perennial grasses are important forage crops and emerging biomass crops and have the potential to be more sustainable grain crops. However, most perennial grass crops are difficult experimental subjects due to their large size, difficult genetics, and/or their recalcitrance to transformation. Thus, a tractable model perennial grass could be used to rapidly make discoveries that can be translated to perennial grass crops. Brachypodium sylvaticum has the potential to serve as such a model because of its small size, rapid generation time, simple genetics, and transformability. Here, we provide a high-quality genome assembly and annotation for B. sylvaticum, an essential resource for a modern model system. In addition, we conducted transcriptomic studies under 4 abiotic stresses (water, heat, salt, and freezing). Our results indicate that crowns are more responsive to freezing than leaves which may help them overwinter. We observed extensive transcriptional responses with varying temporal dynamics to all abiotic stresses, including classic heat-responsive genes. These results can be used to form testable hypotheses about how perennial grasses respond to these stresses. Taken together, these results will allow B. sylvaticum to serve as a truly tractable perennial model system.

JGI Plant Gene Atlas: an updateable transcriptome resource to improve functional gene descriptions across the plant kingdom.

Gene functional descriptions offer a crucial line of evidence for candidate genes underlying trait variation. Conversely, plant responses to environmental cues represent important resources to decipher gene function and subsequently provide molecular targets for plant improvement through gene editing. However, biological roles of large proportions of genes across the plant phylogeny are poorly annotated. Here we describe the Joint Genome Institute (JGI) Plant Gene Atlas, an updateable data resource consisting of transcript abundance assays spanning 18 diverse species. To integrate across these diverse genotypes, we analyzed expression profiles, built gene clusters that exhibited tissue/condition specific expression, and tested for transcriptional response to environmental queues. We discovered extensive phylogenetically constrained and condition-specific expression profiles for genes without any previously documented functional annotation. Such conserved expression patterns and tightly co-expressed gene clusters let us assign expression derived additional biological information to 64 495 genes with otherwise unknown functions. The ever-expanding Gene Atlas resource is available at JGI Plant Gene Atlas (https://plantgeneatlas.jgi.doe.gov) and Phytozome (https://phytozome.jgi.doe.gov/), providing bulk access to data and user-specified queries of gene sets. Combined, these web interfaces let users access differentially expressed genes, track orthologs across the Gene Atlas plants, graphically represent co-expressed genes, and visualize gene ontology and pathway enrichments.

A view of the pan-genome of domesticated Cowpea (Vigna unguiculata [L.] Walp.).

Cowpea, Vigna unguiculata L. Walp., is a diploid warm-season legume of critical importance as both food and fodder in sub-Saharan Africa. This species is also grown in Northern Africa, Europe, Latin America, North America, and East to Southeast Asia. To capture the genomic diversity of domesticates of this important legume, de novo genome assemblies were produced for representatives of six subpopulations of cultivated cowpea identified previously from genotyping of several hundred diverse accessions. In the most complete assembly (IT97K-499-35), 26,026 core and 4963 noncore genes were identified, with 35,436 pan genes when considering all seven accessions. GO terms associated with response to stress and defense response were highly enriched among the noncore genes, while core genes were enriched in terms related to transcription factor activity, and transport and metabolic processes. Over 5 million single nucleotide polymorphisms (SNPs) relative to each assembly and over 40 structural variants >1 Mb in size were identified by comparing genomes. Vu10 was the chromosome with the highest frequency of SNPs, and Vu04 had the most structural variants. Noncore genes harbor a larger proportion of potentially disruptive variants than core genes, including missense, stop gain, and frameshift mutations; this suggests that noncore genes substantially contribute to diversity within domesticated cowpea.

Multiple origins, one evolutionary trajectory: gradual evolution characterizes distinct lineages of allotetraploid Brachypodium.

The "genomic shock" hypothesis posits that unusual challenges to genome integrity such as whole genome duplication may induce chaotic genome restructuring. Decades of research on polyploid genomes have revealed that this is often, but not always the case. While some polyploids show major chromosomal rearrangements and derepression of transposable elements in the immediate aftermath of whole genome duplication, others do not. Nonetheless, all polyploids show gradual diploidization over evolutionary time. To evaluate these hypotheses, we produced a chromosome-scale reference genome for the natural allotetraploid grass Brachypodium hybridum, accession "Bhyb26." We compared 2 independently derived accessions of B. hybridum and their deeply diverged diploid progenitor species Brachypodium stacei and Brachypodium distachyon. The 2 B. hybridum lineages provide a natural timecourse in genome evolution because one formed 1.4 million years ago, and the other formed 140 thousand years ago. The genome of the older lineage reveals signs of gradual post-whole genome duplication genome evolution including minor gene loss and genome rearrangement that are missing from the younger lineage. In neither B. hybridum lineage do we find signs of homeologous recombination or pronounced transposable element activation, though we find evidence supporting steady post-whole genome duplication transposable element activity in the older lineage. Gene loss in the older lineage was slightly biased toward 1 subgenome, but genome dominance was not observed at the transcriptomic level. We propose that relaxed selection, rather than an abrupt genomic shock, drives evolutionary novelty in B. hybridum, and that the progenitor species' similarity in transposable element load may account for the subtlety of the observed genome dominance.

Reference genome of the nutrition-rich orphan crop chia (Salvia hispanica) and its implications for future breeding.

Chia (Salvia hispanica L.) is one of the most popular nutrition-rich foods and pseudocereal crops of the family Lamiaceae. Chia seeds are a rich source of proteins, polyunsaturated fatty acids (PUFAs), dietary fibers, and antioxidants. In this study, we present the assembly of the chia reference genome, which spans 303.6 Mb and encodes 48,090 annotated protein-coding genes. Our analysis revealed that ~42% of the chia genome harbors repetitive content, and identified ~3 million single nucleotide polymorphisms (SNPs) and 15,380 simple sequence repeat (SSR) marker sites. By investigating the chia transcriptome, we discovered that ~44% of the genes undergo alternative splicing with a higher frequency of intron retention events. Additionally, we identified chia genes associated with important nutrient content and quality traits, such as the biosynthesis of PUFAs and seed mucilage fiber (dietary fiber) polysaccharides. Notably, this is the first report of in-silico annotation of a plant genome for protein-derived small bioactive peptides (biopeptides) associated with improving human health. To facilitate further research and translational applications of this valuable orphan crop, we have developed the Salvia genomics database (SalviaGDB), accessible at https://salviagdb.org.

Chia (Salvia hispanica) Gene Expression Atlas Elucidates Dynamic Spatio-Temporal Changes Associated With Plant Growth and Development.

Chia (Salvia hispanica L.), now a popular superfood and a pseudocereal, is one of the richest sources of dietary nutrients such as protein, fiber, and polyunsaturated fatty acids (PUFAs). At present, the genomic and genetic information available in the public domain for this crop are scanty, which hinders an understanding of its growth and development and genetic improvement. We report an RNA-sequencing (RNA-Seq)-based comprehensive transcriptome atlas of Chia sampled from 13 tissue types covering vegetative and reproductive growth stages. We used ~355 million high-quality reads of total ~394 million raw reads from transcriptome sequencing to generate de novo reference transcriptome assembly and the tissue-specific transcript assemblies. After the quality assessment of the merged assemblies and implementing redundancy reduction methods, 82,663 reference transcripts were identified. About 65,587 of 82,663 transcripts were translated into 99,307 peptides, and we were successful in assigning InterPro annotations to 45,209 peptides and gene ontology (GO) terms to 32,638 peptides. The assembled transcriptome is estimated to have the complete sequence information for ~86% of the genes found in the Chia genome. Furthermore, the analysis of 53,200 differentially expressed transcripts (DETs) revealed their distinct expression patterns in Chia's vegetative and reproductive tissues; tissue-specific networks and developmental stage-specific networks of transcription factors (TFs); and the regulation of the expression of enzyme-coding genes associated with important metabolic pathways. In addition, we identified 2,411 simple sequence repeats (SSRs) as potential genetic markers from the transcripts. Overall, this study provides a comprehensive transcriptome atlas, and SSRs, contributing to building essential genomic resources to support basic research, genome annotation, functional genomics, and molecular breeding of Chia.

Meteorological Controls on Local and Regional Volcanic Ash Dispersal.

Volcanic ash has the capacity to impact human health, livestock, crops and infrastructure, including international air traffic. For recent major eruptions, information on the volcanic ash plume has been combined with relatively coarse-resolution meteorological model output to provide simulations of regional ash dispersal, with reasonable success on the scale of hundreds of kilometres. However, to predict and mitigate these impacts locally, significant improvements in modelling capability are required. Here, we present results from a dynamic meteorological-ash-dispersion model configured with sufficient resolution to represent local topographic and convectively-forced flows. We focus on an archetypal volcanic setting, Soufrière, St Vincent, and use the exceptional historical records of the 1902 and 1979 eruptions to challenge our simulations. We find that the evolution and characteristics of ash deposition on St Vincent and nearby islands can be accurately simulated when the wind shear associated with the trade wind inversion and topographically-forced flows are represented. The wind shear plays a primary role and topographic flows a secondary role on ash distribution on local to regional scales. We propose a new explanation for the downwind ash deposition maxima, commonly observed in volcanic eruptions, as resulting from the detailed forcing of mesoscale meteorology on the ash plume.

The Physcomitrella patens chromosome-scale assembly reveals moss genome structure and evolution.

The draft genome of the moss model, Physcomitrella patens, comprised approximately 2000 unordered scaffolds. In order to enable analyses of genome structure and evolution we generated a chromosome-scale genome assembly using genetic linkage as well as (end) sequencing of long DNA fragments. We find that 57% of the genome comprises transposable elements (TEs), some of which may be actively transposing during the life cycle. Unlike in flowering plant genomes, gene- and TE-rich regions show an overall even distribution along the chromosomes. However, the chromosomes are mono-centric with peaks of a class of Copia elements potentially coinciding with centromeres. Gene body methylation is evident in 5.7% of the protein-coding genes, typically coinciding with low GC and low expression. Some giant virus insertions are transcriptionally active and might protect gametes from viral infection via siRNA mediated silencing. Structure-based detection methods show that the genome evolved via two rounds of whole genome duplications (WGDs), apparently common in mosses but not in liverworts and hornworts. Several hundred genes are present in colinear regions conserved since the last common ancestor of plants. These syntenic regions are enriched for functions related to plant-specific cell growth and tissue organization. The P. patens genome lacks the TE-rich pericentromeric and gene-rich distal regions typical for most flowering plant genomes. More non-seed plant genomes are needed to unravel how plant genomes evolve, and to understand whether the P. patens genome structure is typical for mosses or bryophytes.

Extensive gene content variation in the Brachypodium distachyon pan-genome correlates with population structure.

While prokaryotic pan-genomes have been shown to contain many more genes than any individual organism, the prevalence and functional significance of differentially present genes in eukaryotes remains poorly understood. Whole-genome de novo assembly and annotation of 54 lines of the grass Brachypodium distachyon yield a pan-genome containing nearly twice the number of genes found in any individual genome. Genes present in all lines are enriched for essential biological functions, while genes present in only some lines are enriched for conditionally beneficial functions (e.g., defense and development), display faster evolutionary rates, lie closer to transposable elements and are less likely to be syntenic with orthologous genes in other grasses. Our data suggest that differentially present genes contribute substantially to phenotypic variation within a eukaryote species, these genes have a major influence in population genetics, and transposable elements play a key role in pan-genome evolution.

Cambrian cinctan echinoderms shed light on feeding in the ancestral deuterostome.

Reconstructing the feeding mode of the latest common ancestor of deuterostomes is key to elucidating the early evolution of feeding in chordates and allied phyla; however, it is debated whether the ancestral deuterostome was a tentaculate feeder or a pharyngeal filter feeder. To address this, we evaluated the hydrodynamics of feeding in a group of fossil stem-group echinoderms (cinctans) using computational fluid dynamics. We simulated water flow past three-dimensional digital models of a Cambrian fossil cinctan in a range of possible life positions, adopting both passive tentacular feeding and active pharyngeal filter feeding. The results demonstrate that an orientation with the mouth facing downstream of the current was optimal for drag and lift reduction. Moreover, they show that there was almost no flow to the mouth and associated marginal groove under simulations of passive feeding, whereas considerable flow towards the animal was observed for active feeding, which would have enhanced the transport of suspended particles to the mouth. This strongly suggests that cinctans were active pharyngeal filter feeders, like modern enteropneust hemichordates and urochordates, indicating that the ancestral deuterostome employed a similar feeding strategy.

Uncertainty analysis of a model of wind-blown volcanic plumes.

Mathematical models of natural processes can be used as inversion tools to predict unobserved properties from measured quantities. Uncertainty in observations and model formulation impact on the efficacy of inverse modelling. We present a general methodology, history matching, that can be used to investigate the effect of observational and model uncertainty on inverse modelling studies. We demonstrate history matching on an integral model of volcanic plumes that is used to estimate source conditions from observations of the rise height of plumes during the eruptions of Eyjafjallajökull, Iceland, in 2010 and Grímsvötn, Iceland, in 2011. Sources of uncertainty are identified and quantified, and propagated through the integral plume model. A preliminary sensitivity analysis is performed to identify the uncertain model parameters that strongly influence model predictions. Model predictions are assessed against observations through an implausibility measure that rules out model inputs that are considered implausible given the quantified uncertainty. We demonstrate that the source mass flux at the volcano can be estimated from plume height observations, but the magmatic temperature, exit velocity and exsolved gas mass fraction cannot be accurately determined. Uncertainty in plume height observations and entrainment coefficients results in a large range of plausible values of the source mass flux. Our analysis shows that better constraints on entrainment coefficients for volcanic plumes and more precise observations of plume height are required to obtain tightly constrained estimates of the source mass flux.

Enhanced threat detection in experienced riot police officers: cognitive evidence from the face-in-the-crowd effect.

We explored how varying levels of professional expertise in hostile crowd management could enhance threat detection capabilities as assessed by the face in the crowd paradigm. Trainee police officers and more experienced police officers specialized in, and having extensive experience with, riot control, were compared with participants with no experience in hostile crowd management on their search times and accuracy levels in detecting angry and happy face targets against a display of emotional and neutral distractor faces. The experienced officers relative to their trainee counterparts and nonpolice controls showed enhanced detection for threatening faces in both types of display along with a greater degree of inhibitory control over angry face distractors. These findings help to reinforce the ecological validity of the face in the crowd paradigm and provide a new theoretical link for the role of individual differences on the attentional processing of socially relevant stimuli.

The influence of localised size reorganisation on short-duration bidispersed granular flows.

We investigate experimentally the runout resulting from the collapse of a granular column containing two particle species that differ in size only. The experimental configuration is strictly two-dimensional (only one particle per width of the experimental tank) and we explore both the role of the initial arrangement and proportion of the two particle sizes in the column, using high-speed videography, and by determining the centres of mass of the big and small particles in the initial column and the final deposit. The duration of the experiment is sufficiently short that large-scale segregation does not occur, however, we find a clear dependence of runout on both initial mixture arrangement and proportion for all conditions. We investigated this observation through detailed analysis of the flow front motion, and identify a characteristic "stopping" phase when dissipation dominates, and we apply a shallow layer model at the flow front to show how the initial mixture arrangement and proportion influence the effective coefficient of friction during emplacement. We find that a bidispersed mixture can induce a larger friction on emplacement than a monodispersed mixture, and the highest coefficient of friction was found for a well-mixed initial arrangement of particles at the proportion that shows maximum horizontal spreading of the flow. These observations suggest that downwards percolation of fine particles takes place at the front of the collapsing column, and so localised size segregation processes at the flow front can control flow mobility. This effect is likely to be important in controlling the mobility of large geophysical flows that occur on finite time scales, and whose deposits typically show granular segregation at the front and edges but not throughout the entire deposit.

Structure, dynamics, evolution, and function of a major scaffold component in the nuclear pore complex.

The nuclear pore complex, composed of proteins termed nucleoporins (Nups), is responsible for nucleocytoplasmic transport in eukaryotes. Nuclear pore complexes (NPCs) form an annular structure composed of the nuclear ring, cytoplasmic ring, a membrane ring, and two inner rings. Nup192 is a major component of the NPC's inner ring. We report the crystal structure of Saccharomyces cerevisiae Nup192 residues 2-960 [ScNup192(2-960)], which adopts an α-helical fold with three domains (i.e., D1, D2, and D3). Small angle X-ray scattering and electron microscopy (EM) studies reveal that ScNup192(2-960) could undergo long-range transition between "open" and "closed" conformations. We obtained a structural model of full-length ScNup192 based on EM, the structure of ScNup192(2-960), and homology modeling. Evolutionary analyses using the ScNup192(2-960) structure suggest that NPCs and vesicle-coating complexes are descended from a common membrane-coating ancestral complex. We show that suppression of Nup192 expression leads to compromised nuclear transport and hypothesize a role for Nup192 in modulating the permeability of the NPC central channel.

Assembly of macromolecular complexes by satisfaction of spatial restraints from electron microscopy images.

To obtain a structural model of a macromolecular assembly by single-particle EM, a large number of particle images need to be collected, aligned, clustered, averaged, and finally assembled via reconstruction into a 3D density map. This process is limited by the number and quality of the particle images, the accuracy of the initial model, and the compositional and conformational heterogeneity. Here, we describe a structure determination method that avoids the reconstruction procedure. The atomic structures of the individual complex components are assembled by optimizing a match against 2D EM class-average images, an excluded volume criterion, geometric complementarity, and optional restraints from proteomics and chemical cross-linking experiments. The optimization relies on a simulated annealing Monte Carlo search and a divide-and-conquer message-passing algorithm. Using simulated and experimentally determined EM class averages for 12 and 4 protein assemblies, respectively, we show that a few class averages can indeed result in accurate models for complexes of as many as five subunits. Thus, integrative structural biology can now benefit from the relative ease with which the EM class averages are determined.

Atomic structure of the nuclear pore complex targeting domain of a Nup116 homologue from the yeast, Candida glabrata.

The nuclear pore complex (NPC), embedded in the nuclear envelope, is a large, dynamic molecular assembly that facilitates exchange of macromolecules between the nucleus and the cytoplasm. The yeast NPC is an eightfold symmetric annular structure composed of ~456 polypeptide chains contributed by ~30 distinct proteins termed nucleoporins. Nup116, identified only in fungi, plays a central role in both protein import and mRNA export through the NPC. Nup116 is a modular protein with N-terminal "FG" repeats containing a Gle2p-binding sequence motif and a NPC targeting domain at its C-terminus. We report the crystal structure of the NPC targeting domain of Candida glabrata Nup116, consisting of residues 882-1034 [CgNup116(882-1034)], at 1.94 Å resolution. The X-ray structure of CgNup116(882-1034) is consistent with the molecular envelope determined in solution by small-angle X-ray scattering. Structural similarities of CgNup116(882-1034) with homologous domains from Saccharomyces cerevisiae Nup116, S. cerevisiae Nup145N, and human Nup98 are discussed.

Structure-function mapping of a heptameric module in the nuclear pore complex.

The nuclear pore complex (NPC) is a multiprotein assembly that serves as the sole mediator of nucleocytoplasmic exchange in eukaryotic cells. In this paper, we use an integrative approach to determine the structure of an essential component of the yeast NPC, the ~600-kD heptameric Nup84 complex, to a precision of ~1.5 nm. The configuration of the subunit structures was determined by satisfaction of spatial restraints derived from a diverse set of negative-stain electron microscopy and protein domain-mapping data. Phenotypic data were mapped onto the complex, allowing us to identify regions that stabilize the NPC's interaction with the nuclear envelope membrane and connect the complex to the rest of the NPC. Our data allow us to suggest how the Nup84 complex is assembled into the NPC and propose a scenario for the evolution of the Nup84 complex through a series of gene duplication and loss events. This work demonstrates that integrative approaches based on low-resolution data of sufficient quality can generate functionally informative structures at intermediate resolution.

Modeling of proteins and their assemblies with the integrative modeling platform.

To understand the workings of the living cell, we need to characterize protein assemblies that constitute the cell (for example, the ribosome, 26S proteasome, and the nuclear pore complex). A reliable high-resolution structural characterization of these assemblies is frequently beyond the reach of current experimental methods, such as X-ray crystallography, NMR spectroscopy, electron microscopy, footprinting, chemical cross-linking, FRET spectroscopy, small-angle X-ray scattering, and proteomics. However, the information garnered from different methods can be combined and used to build computational models of the assembly structures that are consistent with all of the available datasets. Here, we describe a protocol for this integration, whereby the information is converted to a set of spatial restraints and a variety of optimization procedures can be used to generate models that satisfy the restraints as much as possible. These generated models can then potentially inform about the precision and accuracy of structure determination, the accuracy of the input datasets, and further data generation. We also demonstrate the Integrative Modeling Platform (IMP) software, which provides the necessary computational framework to implement this protocol, and several applications for specific-use cases.