Proinflammatory microenvironments within the intestine regulate the differentiation of tissue-resident CD8⁺ T cells responding to infection.

We report that oral infection with Yersinia pseudotuberculosis results in the development of two distinct populations of pathogen-specific CD8(+) tissue-resident memory T cells (TRM cells) in the lamina propria. CD103(-) T cells did not require transforming growth factor-ß (TGF-ß) signaling but were true resident memory cells. Unlike CD103(+)CD8(+) T cells, which were TGF-ß dependent and were scattered in the tissue, CD103(-)CD8(+) T cells clustered with CD4(+) T cells and CX3CR1(+) macrophages and/or dendritic cells around areas of bacterial infection. CXCR3-dependent recruitment of cells to inflamed areas was critical for development of the CD103(-) population and pathogen clearance. Our studies have identified the 'preferential' development of CD103(-) TRM cells in inflammatory microenvironments within the lamina propria and suggest that this subset has a critical role in controlling infection.

Multiple wheat genomes reveal global variation in modern breeding.

Advances in genomics have expedited the improvement of several agriculturally important crops but similar efforts in wheat (Triticum spp.) have been more challenging. This is largely owing to the size and complexity of the wheat genome1, and the lack of genome-assembly data for multiple wheat lines2,3. Here we generated ten chromosome pseudomolecule and five scaffold assemblies of hexaploid wheat to explore the genomic diversity among wheat lines from global breeding programs. Comparative analysis revealed extensive structural rearrangements, introgressions from wild relatives and differences in gene content resulting from complex breeding histories aimed at improving adaptation to diverse environments, grain yield and quality, and resistance to stresses4,5. We provide examples outlining the utility of these genomes, including a detailed multi-genome-derived nucleotide-binding leucine-rich repeat protein repertoire involved in disease resistance and the characterization of Sm16, a gene associated with insect resistance. These genome assemblies will provide a basis for functional gene discovery and breeding to deliver the next generation of modern wheat cultivars.

Modulation of receptor-like transmembrane kinase 1 nuclear localization by DA1 peptidases in Arabidopsis.

The cleavage of intracellular domains of receptor-like kinases (RLKs) has an important functional role in the transduction of signals from the cell surface to the nucleus in many organisms. However, the peptidases that catalyze protein cleavage during signal transduction remain poorly understood despite their crucial roles in diverse signaling processes. Here, we report in the flowering plant Arabidopsis thaliana that members of the DA1 family of ubiquitin-regulated Zn metallopeptidases cleave the cytoplasmic kinase domain of transmembrane kinase 1 (TMK1), releasing it for nuclear localization where it represses auxin-responsive cell growth during apical hook formation by phosphorylation and stabilization of the transcriptional repressors IAA32 and IAA34. Mutations in DA1 family members exhibited reduced apical hook formation, and DA1 family-mediated cleavage of TMK1 was promoted by auxin treatment. Expression of the DA1 family-generated intracellular kinase domain of TMK1 by an auxin-responsive promoter fully restored apical hook formation in a tmk1 mutant, establishing the function of DA1 family peptidase activities in TMK1-mediated differential cell growth and apical hook formation. DA1 family peptidase activity therefore modulates TMK1 kinase activity between a membrane location where it stimulates acid cell growth and initiates an auxin-dependent kinase cascade controlling cell proliferation in lateral roots and a nuclear localization where it represses auxin-mediated gene expression and growth.

Ubiquitylation activates a peptidase that promotes cleavage and destabilization of its activating E3 ligases and diverse growth regulatory proteins to limit cell proliferation in Arabidopsis.

The characteristic shapes and sizes of organs are established by cell proliferation patterns and final cell sizes, but the underlying molecular mechanisms coordinating these are poorly understood. Here we characterize a ubiquitin-activated peptidase called DA1 that limits the duration of cell proliferation during organ growth in Arabidopsis thaliana The peptidase is activated by two RING E3 ligases, Big Brother (BB) and DA2, which are subsequently cleaved by the activated peptidase and destabilized. In the case of BB, cleavage leads to destabilization by the RING E3 ligase PROTEOLYSIS 1 (PRT1) of the N-end rule pathway. DA1 peptidase activity also cleaves the deubiquitylase UBP15, which promotes cell proliferation, and the transcription factors TEOSINTE BRANCED 1/CYCLOIDEA/PCF 15 (TCP15) and TCP22, which promote cell proliferation and repress endoreduplication. We propose that DA1 peptidase activity regulates the duration of cell proliferation and the transition to endoreduplication and differentiation during organ formation in plants by coordinating the destabilization of regulatory proteins.

Fast-forward breeding for a food-secure world.

Crop production systems need to expand their outputs sustainably to feed a burgeoning human population. Advances in genome sequencing technologies combined with efficient trait mapping procedures accelerate the availability of beneficial alleles for breeding and research. Enhanced interoperability between different omics and phenotyping platforms, leveraged by evolving machine learning tools, will help provide mechanistic explanations for complex plant traits. Targeted and rapid assembly of beneficial alleles using optimized breeding strategies and precise genome editing techniques could deliver ideal crops for the future. Realizing desired productivity gains in the field is imperative for securing an adequate future food supply for 10 billion people.

TGF-ß signaling to T cells inhibits autoimmunity during lymphopenia-driven proliferation.

T cell-specific deletion of the receptor for transforming growth factor-ß (TGF-ß) mediated by Cre recombinase expressed early in T cell development leads to early-onset lethal autoimmune disease that cannot be controlled by regulatory T cells. However, when we deleted that receptor through the use of Cre driven by a promoter that is active much later in T cell development, adult mice in which most peripheral CD4(+) or CD8(+) T cells lacked the receptor for TGF-ß showed no signs of autoimmunity. Because of their enhanced responses to weak stimulation of the T cell antigen receptor, when transferred into lymphopenic recipients, naive TGF-ß-unresponsive T cells underwent much more proliferation and differentiation into effector cells and induced lymphoproliferative disease. We propose that TGF-ß signaling controls the self-reactivity of peripheral T cells but that in the absence of TGF-ß signals, an added trigger such as lymphopenia is needed to drive overt autoimmune disease.

Understand memory, design better vaccines.

Naive lymphocytes have a finite lifespan and are continually replaced by input from generative organs. In contrast, memory cells or their progeny can last a lifetime. The expanded populations of memory cells and their more widespread distribution provide protection against recurrent infection.

Colloidal Deposition by Polymer-Surfactant Complexes with Dilution and Shear.

Deposition of silica microparticles on glass substrates was measured as a function of cationic polymer-anionic surfactant composition and shear rate. Particles were initially deposited in quiescent conditions in different polymer-surfactant compositions, which were chosen based on prior measurements of composition-dependent polymer-surfactant interactions and deposition behavior (up to 0.5 wt % polymer and 12 wt % surfactant). Programmed shear and dilution profiles in a flow cell together with optical microscopy observation were used to continuously track particle deposition, detachment, and redeposition. Knowledge of the shear-dependent torque on each particle provides information on adhesive torque mediated by polymer-surfactant complexes. Detachment of colloids initially deposited by depletion interactions occurs at low shear rates (∼100 s-1) due to lack of tangential forces or an adhesive torque. Further dilution produced redeposition of particles that resisted detachment (up to ∼2000 s-1) as the result of strong cationic polymer bridge formation, presumably due to preferential surfactant removal. Dilution from different initial compositions indicates a pathway dependence of polymer-surfactant de-complexation into shear-resistant cationic bridges. These findings demonstrate the ability to program deposition behavior via the informed design of initial polymer-surfactant compositions and shear profiles. The particle trajectory analysis developed in this work provides an assay to screen composition-dependent colloidal deposition in diverse materials and applications.

Energy landscapes on polymerized liquid crystal interfaces.

We measure and model monolayers of concentrated diffusing colloidal probes interacting with polymerized liquid crystal (PLC) planar surfaces. At topological defects in local nematic director profiles at PLC surfaces, we observe time-averaged two-dimensional particle density profiles of diffusing colloidal probes that closely correlate with spatial variations in PLC optical properties. An inverse Monte Carlo analysis of particle concentration profiles yields two-dimensional PLC interfacial energy landscapes on the kT-scale, which is the inherent scale of many interfacial phenomena (e.g., self-assembly, adsorption, diffusion). Energy landscapes are modelled as the superposition of macromolecular repulsion and van der Waals attraction based on an anisotropic dielectric function obtained from the liquid crystal birefringence. Modelled van der Waals landscapes capture most net energy landscape variations and correlate well with experimental PLC director profiles around defects. Some energy landscape variations near PLC defects indicate either additional local repulsive interactions or possibly the need for more rigorous van der Waals models with complete spectral data. These findings demonstrate direct, sensitive measurements of kT-scale van der Waals energy landscapes at PLC interfacial defects and suggest the ability to design interfacial anisotropic materials and van der Waals energy landscapes for colloidal assembly.

Genomic innovation for crop improvement.

Crop production needs to increase to secure future food supplies, while reducing its impact on ecosystems. Detailed characterization of plant genomes and genetic diversity is crucial for meeting these challenges. Advances in genome sequencing and assembly are being used to access the large and complex genomes of crops and their wild relatives. These have helped to identify a wide spectrum of genetic variation and permitted the association of genetic diversity with diverse agronomic phenotypes. In combination with improved and automated phenotyping assays and functional genomic studies, genomics is providing new foundations for crop-breeding systems.

Genome sequence of the progenitor of the wheat D genome Aegilops tauschii.

Aegilops tauschii is the diploid progenitor of the D genome of hexaploid wheat (Triticum aestivum, genomes AABBDD) and an important genetic resource for wheat. The large size and highly repetitive nature of the Ae. tauschii genome has until now precluded the development of a reference-quality genome sequence. Here we use an array of advanced technologies, including ordered-clone genome sequencing, whole-genome shotgun sequencing, and BioNano optical genome mapping, to generate a reference-quality genome sequence for Ae. tauschii ssp. strangulata accession AL8/78, which is closely related to the wheat D genome. We show that compared to other sequenced plant genomes, including a much larger conifer genome, the Ae. tauschii genome contains unprecedented amounts of very similar repeated sequences. Our genome comparisons reveal that the Ae. tauschii genome has a greater number of dispersed duplicated genes than other sequenced genomes and its chromosomes have been structurally evolving an order of magnitude faster than those of other grass genomes. The decay of colinearity with other grass genomes correlates with recombination rates along chromosomes. We propose that the vast amounts of very similar repeated sequences cause frequent errors in recombination and lead to gene duplications and structural chromosome changes that drive fast genome evolution.

Anisotropic particle multiphase equilibria in nonuniform fields.

We report a method to predict equilibrium concentration profiles of hard ellipses in nonuniform fields, including multiphase equilibria of fluid, nematic, and crystal phases. Our model is based on a balance of osmotic pressure and field mediated forces by employing the local density approximation. Implementation of this model requires development of accurate equations of state for each phase as a function of hard ellipse aspect ratio in the range k = 1-9. The predicted density profiles display overall good agreement with Monte Carlo simulations for hard ellipse aspect ratios k = 2, 4, and 6 in gravitational and electric fields with fluid-nematic, fluid-crystal, and fluid-nematic-crystal multiphase equilibria. The profiles of local order parameters for positional and orientational order display good agreement with values expected for bulk homogeneous hard ellipses in the same density ranges. Small discrepancies between predictions and simulations are observed at crystal-nematic and crystal-fluid interfaces due to limitations of the local density approximation, finite system sizes, and uniform periodic boundary conditions. The ability of the model to capture multiphase equilibria of hard ellipses in nonuniform fields as a function of particle aspect ratio provides a basis to control anisotropic particle microstructure on interfacial energy landscapes in diverse materials and applications.

Variation in Expression of the HECT E3 Ligase UPL3 Modulates LEC2 Levels, Seed Size, and Crop Yields in Brassica napus.

Identifying genetic variation that increases crop yields is a primary objective in plant breeding. We used association analyses of oilseed rape/canola (Brassica napus) accessions to identify genetic variation that influences seed size, lipid content, and final crop yield. Variation in the promoter region of the HECT E3 ligase gene BnaUPL3 C03 made a major contribution to variation in seed weight per pod, with accessions exhibiting high seed weight per pod having lower levels of BnaUPL3 C03 expression. We defined a mechanism in which UPL3 mediated the proteasomal degradation of LEC2, a master transcriptional regulator of seed maturation. Accessions with reduced UPL3 expression had increased LEC2 protein levels, larger seeds, and prolonged expression of lipid biosynthetic genes during seed maturation. Natural variation in BnaUPL3 C03 expression appears not to have been exploited in current B napus breeding lines and could therefore be used as a new approach to maximize future yields in this important oil crop.

Transforming growth factor-ß signaling controls the formation and maintenance of gut-resident memory T cells by regulating migration and retention.

Tissue-resident memory T (Trm) cells represent a population of memory CD8⁺ T cells that can act as first responders to local infection. The mechanisms regulating the formation and maintenance of intestinal Trm cells remain elusive. Here we showed that transforming growth factor-ß (TGF-ß) controlled both stages of gut Trm cell differentiation through different mechanisms. During the formation phase of Trm cells, TGF-ß signaling inhibited the migration of effector CD8⁺ T cells from the spleen to the gut by dampening the expression of integrin α4ß7. During the maintenance phase, TGF-ß was required for the retention of intestinal Trm cells at least in part through the induction of integrins αEß7 and α1, as well as CD69. Thus, the cytokine acts to control cytotoxic T cell differentiation in lymphoid and peripheral organs.

Design rules for 2D field mediated assembly of different shaped colloids into diverse microstructures.

Assembling different shaped particles into ordered microstructures is an open challenge in creating multifunctional particle-based materials and devices. Here, we report the two-dimensional (2D) AC electric field mediated assembly of different shaped colloidal particles into amorphous, liquid crystalline, and crystalline microstructures. Particle shapes investigated include disks, ellipses, squares, and rectangles, which show how systematic variations in anisotropy and corner curvature determine the number and type of resulting microstructures. AC electric fields induce dipolar interactions to control particle positional and orientational order. Microstructural states are determined via particle tracking to compute order parameters, which agree with computer simulations and show how particle packing and dipolar interactions together produce each structure. Results demonstrate how choice of particle shape and field conditions enable kinetically viable routes to assemble nematic, tetratic, and smectic liquid crystal structures as well as crystals with stretched 4- and 6-fold symmetry. Results show it is possible to assemble all corresponding hard particle phases, but also show how dipolar interactions influence and produce additional microstructures. Our findings provide design rules for the assembly of diverse microstructures of different shaped particles in AC electric fields, which could enable scalable and reconfigurable particle-based materials, displays, and printing technologies.

Hard superellipse phases: particle shape anisotropy & curvature.

We report computer simulations of two-dimensional convex hard superellipse particle phases vs. particle shape parameters including aspect ratio, corner curvature, and sidewall curvature. Shapes investigated include disks, ellipses, squares, rectangles, and rhombuses, as well as shapes with non-uniform curvature including rounded squares, rounded rectangles, and rounded rhombuses. Using measures of orientational order, order parameters, and a novel stretched bond orientational order parameter, we systematically identify particle shape properties that determine liquid crystal and crystalline phases including their coarse boundaries and symmetry. We observe phases including isotropic, nematic, tetratic, plastic crystals, square crystals, and hexagonal crystals (including stretched variants). Our results catalog known benchmark shapes, but include new shapes that also interpolate between known shapes. Our results indicate design rules for particle shapes that determine two-dimensional liquid, liquid crystalline, and crystalline microstructures that can be realized via particle assembly.

Hidden variation in polyploid wheat drives local adaptation.

Wheat has been domesticated into a large number of agricultural environments and has the ability to adapt to diverse environments. To understand this process, we survey genotype, repeat content, and DNA methylation across a bread wheat landrace collection representing global genetic diversity. We identify independent variation in methylation, genotype, and transposon copy number. We show that these, so far unexploited, sources of variation have had a significant impact on the wheat genome and that ancestral methylation states become preferentially "hard coded" as single nucleotide polymorphisms (SNPs) via 5-methylcytosine deamination. These mechanisms also drive local adaption, impacting important traits such as heading date and salt tolerance. Methylation and transposon diversity could therefore be used alongside SNP-based markers for breeding.

The RIG-I-like receptor LGP2 controls CD8(+) T cell survival and fitness.

The RIG-I-like receptors (RLRs) signal innate immune defenses upon RNA virus infection, but their roles in adaptive immunity have not been clearly defined. Here, we showed that the RLR LGP2 was not essential for induction of innate immune defenses, but rather was required for controlling antigen-specific CD8(+) T cell survival and fitness during peripheral T cell-number expansion in response to virus infection. Adoptive transfer and biochemical studies demonstrated that T cell-receptor signaling induced LGP2 expression wherein LGP2 operated to regulate death-receptor signaling and imparted sensitivity to CD95-mediated cell death. Thus, LGP2 promotes an essential prosurvival signal in response to antigen stimulation to confer CD8(+) T cell-number expansion and effector functions against divergent RNA viruses, including West Nile virus and lymphocytic choriomeningitis virus.

Droplet Formation and Growth Mechanisms in Reaction-Induced Spontaneous Emulsification of 3-(Trimethoxysilyl) Propyl Methacrylate.

Spontaneous emulsification of 3-(trimethoxysilyl) propyl methacrylate (TPM) can produce complex and active colloids, nanoparticles, or monodisperse Pickering emulsions. Despite the applicability of TPM in particle synthesis, the nucleation and growth mechanisms of TPM emulsions are still poorly understood. We investigate droplet formation and growth of TPM in aqueous solutions under quiescent conditions. Our results show that in the absence of stirring the mechanisms of diffusion and stranding likely drive the spontaneous emulsification of TPM through the formation of co-soluble species during hydrolysis. In addition, turbidity and dynamic light scattering experiments show that the pH modulates the growth mechanism. At pH 10.1, the droplets grow via Ostwald ripening, while at pH 11.5, the droplets grow via monomer addition. Adding surfactants [Tween, sodium dodecyl sulfate (SDS), or cetyltrimethylammonium bromide] leads to <100 nm droplets that are kinetically stable. The growth of Tween droplets occurs through addition of TPM species while the number density of droplets is kept constant. In addition, in the presence of the ionic surfactant SDS, electrostatic repulsion between the solubilized TPM species and SDS leads to a significant increase in the number density of droplets as well as additional nucleation events. Finally, imaging of the solubilization of TPM in capillaries shows that in the absence of a surfactant, TPM hydrolysis is likely the rate-limiting step for emulsification, whereas the presence of silica particles in the aqueous phase likely acts as a catalyst of TPM hydrolysis. Our experiments highlight the importance of diffusion and solubilization of TPM species in the aqueous phase in the nucleation and growth of droplets.

Anisotropic colloidal interactions & assembly in AC electric fields.

We match experimental and simulated configurations of anisotropic epoxy colloidal particles in high frequency AC electric fields by identifying analytical potentials for dipole-field and dipole-dipole interactions. We report an inverse Monte Carlo simulation algorithm to determine optimal fits of analytical potentials by matching simulated and experimental distribution functions for non-uniform liquid, liquid crystal, and crystal microstructures in varying amplitude electric fields. Two potentials that include accurate particle volume and dimensions along with a concentration dependent prefactor quantitatively capture experimental observations. At low concentrations, an effective ellipsoidal point dipole potential works well, whereas a novel stretched point dipole potential is found to be suitable at all concentrations, field amplitudes, and degrees of ordering. The simplicity, accuracy, and adjustability of the stretched point dipole potential suggest it can be applied to model field mediated microstructures and assembly of systematically varying anisotropic particle shapes.