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Overlapping coding regions balance selective forces between multiple genes. One possible division of nucleotide sequence is that the predominant selective force on a particular nucleotide can be attributed to just one gene. While this arrangement has been observed in regions in which one gene is structured and the other is disordered, we sought to explore how overlapping genes balance constraints when both protein products are structured over the same sequence. We use a combination of sequence analysis, functional assays, and selection experiments to examine an overlapped region in HIV-1 that encodes helical regions in both Env and Rev. We find that functional segregation occurs even in this overlap, with each protein spacing its functional residues in a manner that allows a mutable non-binding face of one helix to encode important functional residues on a charged face in the other helix. Additionally, our experiments reveal novel and critical functional residues in Env and have implications for the therapeutic targeting of HIV-1.

The HIV-1 Tat protein hijacks P-TEFb kinase to activate paused RNA polymerase II (RNAP II) at the viral promoter. Tat binds additional host factors, but it is unclear how they regulate RNAP II elongation. Here, we identify the cytoplasmic ubiquitin ligase UBE2O as critical for Tat transcriptional activity. Tat hijacks UBE2O to ubiquitinate the P-TEFb kinase inhibitor HEXIM1 of the 7SK snRNP, a fraction of which also resides in the cytoplasm bound to P-TEFb. HEXIM1 ubiquitination sequesters it in the cytoplasm and releases P-TEFb from the inhibitory 7SK complex. Free P-TEFb then becomes enriched in chromatin, a process that is also stimulated by treating cells with a CDK9 inhibitor. Finally, we demonstrate that UBE2O is critical for P-TEFb recruitment to the HIV-1 promoter. Together, the data support a unique model of elongation control where non-degradative ubiquitination of nuclear and cytoplasmic 7SK snRNP pools increases P-TEFb levels for transcriptional activation.

Anthropogenic activities have resulted in the intensified use of water resources. For example, open pit bitumen extraction by Canada's oil sands operations uses an estimated volume of three barrels of water for every barrel of oil produced. The waste tailings-oil sands process water (OSPW)-are stored in holding ponds, and present an environmental concern as they are comprised of residual hydrocarbons and metals. Following the hypothesis that endogenous OSPW microbial communities have an enhanced tolerance to heavy metals, we tested the capacity of planktonic and biofilm populations from OSPW to withstand metal ion challenges, using Cupriavidus metallidurans, a known metal-resistant organism, for comparison. The toxicity of the metals toward biofilm and planktonic bacterial populations was determined by measuring the minimum biofilm inhibitory concentrations (MBICs) and planktonic minimum inhibitory concentrations (MICs) using the MBEC ™ assay. We observed that the OSPW community and C. metallidurans had similar tolerances to 22 different metals. While thiophillic elements (Te, Ag, Cd, Ni) were found to be most toxic, the OSPW consortia demonstrated higher tolerance to metals reported in tailings ponds (Al, Fe, Mo, Pb). Metal toxicity correlated with a number of physicochemical characteristics of the metals. Parameters reflecting metal-ligand affinities showed fewer and weaker correlations for the community compared to C. metallidurans, suggesting that the OSPW consortia may have developed tolerance mechanisms toward metals present in their environment.

The HIV-1 protein Rev controls a critical step in viral replication by mediating the nuclear export of unspliced and singly-spliced viral mRNAs. Multiple Rev subunits assemble on the Rev Response Element (RRE), a structured region present in these RNAs, and direct their export through the Crm1 pathway. Rev-RRE assembly occurs via several Rev oligomerization and RNA-binding steps, but how these steps are coordinated to form an export–competent complex is unclear. Here, we report the first crystal structure of a Rev dimer-RRE complex, revealing a dramatic rearrangement of the Rev-dimer upon RRE binding through re-packing of its hydrophobic protein–protein interface. Rev-RNA recognition relies on sequence-specific contacts at the well-characterized IIB site and local RNA architecture at the second site. The structure supports a model in which the RRE utilizes the inherent plasticity of Rev subunit interfaces to guide the formation of a functional complex.

The HIV Rev protein routes viral RNAs containing the Rev Response Element (RRE) through the Crm1 nuclear export pathway to the cytoplasm where viral proteins are expressed and genomic RNA is delivered to assembling virions. The RRE assembles a Rev oligomer that displays nuclear export sequences (NESs) for recognition by the Crm1-RanGTP nuclear receptor complex. Here we provide the first view of an assembled HIV-host nuclear export complex using single-particle electron microscopy. Unexpectedly, Crm1 forms a dimer with an extensive interface that enhances association with Rev-RRE and poises NES binding sites to interact with a Rev oligomer. The interface between Crm1 monomers explains differences between Crm1 orthologs that alter nuclear export and determine cellular tropism for viral replication. The arrangement of the export complex identifies a novel binding surface to possibly target an HIV inhibitor and may point to a broader role for Crm1 dimerization in regulating host gene expression.

Bacterial chemotaxis systems are as diverse as the environments that bacteria inhabit, but how much environmental variation can cells tolerate with a single system? Diversification of a single chemotaxis system could serve as an alternative, or even evolutionary stepping-stone, to switching between multiple systems. We hypothesized that mutations in gene regulation could lead to heritable control of chemotactic diversity. By simulating foraging and colonization of E. coli using a single-cell chemotaxis model, we found that different environments selected for different behaviors. The resulting trade-offs show that populations facing diverse environments would ideally diversify behaviors when time for navigation is limited. We show that advantageous diversity can arise from changes in the distribution of protein levels among individuals, which could occur through mutations in gene regulation. We propose experiments to test our prediction that chemotactic diversity in a clonal population could be a selectable trait that enables adaptation to environmental variability.

More than 20 recurrent missense gain-of-function (GOF) mutations have been identified in the sodium-activated potassium (KNa) channel gene KCNT1 in patients with severe developmental and epileptic encephalopathies (DEEs), most of which are resistant to current therapies. Defining the neuron types most vulnerable to KCNT1 GOF will advance our understanding of disease mechanisms and provide refined targets for precision therapy efforts. Here, we assessed the effects of heterozygous expression of a Kcnt1 GOF variant (Kcnt1Y777H) on KNa currents and neuronal physiology among cortical glutamatergic and GABAergic neurons in mice, including those expressing vasoactive intestinal polypeptide (VIP), somatostatin (SST), and parvalbumin (PV), to identify and model the pathogenic mechanisms of autosomal dominant KCNT1 GOF variants in DEEs. Although the Kcnt1Y777H variant had no effects on glutamatergic or VIP neuron function, it increased subthreshold KNa currents in both SST and PV neurons but with opposite effects on neuronal output; SST neurons became hypoexcitable with a higher rheobase current and lower action potential (AP) firing frequency, whereas PV neurons became hyperexcitable with a lower rheobase current and higher AP firing frequency. Further neurophysiological and computational modeling experiments showed that the differential effects of the Kcnt1Y777H variant on SST and PV neurons are not likely due to inherent differences in these neuron types, but to an increased persistent sodium current in PV, but not SST, neurons. The Kcnt1Y777H variant also increased excitatory input onto, and chemical and electrical synaptic connectivity between, SST neurons. Together, these data suggest differential pathogenic mechanisms, both direct and compensatory, contribute to disease phenotypes, and provide a salient example of how a pathogenic ion channel variant can cause opposite functional effects in closely related neuron subtypes due to interactions with other ionic conductances.

Abstract Topological physics has revolutionized materials science, introducing topological phases of matter in diverse settings ranging from quantum to photonic and phononic systems. Herein, we present a family of topological systems, which we term “strain topological metamaterials”, whose topological properties are hidden and unveiled only under higher-order (strain) coordinate transformations. We firstly show that the canonical mass dimer, a model that can describe various settings such as electrical circuits and optics, among others, belongs to this family where strain coordinates reveal a topological nontriviality for the edge states at free boundaries. Subsequently, we introduce a mechanical analog of the Majorana-supporting Kitaev chain, which supports topological edge states for both fixed and free boundaries within the proposed framework. Thus, our findings not only extend the way topological edge states are identified, but also promote the fabrication of novel topological metamaterials in various fields, with more complex, tailored boundaries.

Abstract India is the world’s second largest producer of wheat, with more than 40% increase in production since 2000. Increasing temperatures raise concerns about wheat’s sensitivity to heat. Traditionally-grown sorghum is an alternative rabi (winter season) cereal, but area under sorghum production has declined more than 20% since 2000. We examine sensitivity of wheat and sorghum yields to historical temperature and compare water requirements in districts where both cereals are cultivated. Wheat yields are sensitive to increases in maximum daily temperature in multiple stages of the growing season, while sorghum does not display the same sensitivity. Crop water requirements (mm) are 1.4 times greater for wheat than sorghum, mainly due to extension of its growing season into summer. However, water footprints (m3 per ton) are approximately 15% less for wheat due to its higher yields. Sensitivity to future climate projections, without changes in management, suggests 5% decline in wheat yields and 12% increase in water footprints by 2040, compared with 4% increase in water footprint for sorghum. On balance, sorghum provides a climate-resilient alternative to wheat for expansion in rabi cereals. However, yields need to increase to make sorghum competitive for farmer profits and efficient use of land to provide nutrients.

The corrosion properties of engineering alloys are strongly influenced by local variations seen at the micro level. Here, the authors demonstrate how the metal grain orientation can influence the protectiveness of the oxide, which provides a possible route for improving corrosion performance.

Abstract Achieving food security in sub-Saharan Africa (SSA) is a multidimensional challenge. SSA reliance on food imports is expected to grow in the coming decades to meet the population's demand, projected to double to over 2 billion people by 2050. In addition, climate change is already affecting food production and supply chains across the region. Addressing these multiple food security challenges will necessitate rapid enhancements in agricultural productivity, which is influenced by a host of demographic, agronomic, and climatic factors. We use statistical approaches to examine rainfed maize in Kenya, where maize cultivation and consumption are widespread and central to livelihoods and national food security. We find that improving a suite of agronomic factors, such as applying fertilizer, planting certified seeds, and extension services, will have a greater effect on rainfed maize productivity than demographics and can offset the effects of climate change. These findings could also offer insights into similar challenges for other crops in Kenya and other SSA countries.

Summary: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with high metastasis and therapeutic resistance. Activating transcription factor 4 (ATF4), a master regulator of cellular stress, is exploited by cancer cells to survive. Prior research and data reported provide evidence that high ATF4 expression correlates with worse overall survival in PDAC. Tomatidine, a natural steroidal alkaloid, is associated with inhibition of ATF4 signaling in multiple diseases. Here, we discovered that in vitro and in vivo tomatidine treatment of PDAC cells inhibits tumor growth. Tomatidine inhibited nuclear translocation of ATF4 and reduced the transcriptional binding of ATF4 with downstream promoters. Tomatidine enhanced gemcitabine chemosensitivity in 3D ECM-hydrogels and in vivo. Tomatidine treatment was associated with induction of ferroptosis signaling validated by increased lipid peroxidation, mitochondrial biogenesis, and decreased GPX4 expression in PDAC cells. This study highlights a possible therapeutic approach utilizing a plant-derived metabolite, tomatidine, to target ATF4 activity in PDAC.

Water scarcity and related climate volatility are growing constraints on agricultural production landscapes around the world. While the adaptation options available are often dictated by system context, in many places broad-scale transformations are occurring in response to water-related pressures. We sought contributions from across regions, agricultural system types, and scientific disciplines to examine agricultural land use transitions driven by water scarcity, including the tradeoffs associated with alternative land uses; impacts on food production, environment, and society; innovations that can buffer risk; and considerations for planning and implementation. The research presented in this collection highlights the spectrum of policy and practice changes that are needed to facilitate beneficial land use transitions and system transformations, from quantifying risks, to evaluating multidimensional tradeoffs, to developing socio-technical policy bundles to maximize co-benefits.

Ant colonies regulate foraging in response to their collective hunger, yet the mechanism behind this distributed regulation remains unclear. Previously, by imaging food flow within ant colonies we showed that the frequency of foraging events declines linearly with colony satiation (Greenwald et al., 2018). Our analysis implied that as a forager distributes food in the nest, two factors affect her decision to exit for another foraging trip: her current food load and its rate of change. Sensing these variables can be attributed to the forager’s individual cognitive ability. Here, new analyses of the foragers’ trajectories within the nest imply a different way to achieve the observed regulation. Instead of an explicit decision to exit, foragers merely tend toward the depth of the nest when their food load is high and toward the nest exit when it is low. Thus, the colony shapes the forager’s trajectory by controlling her unloading rate, while she senses only her current food load. Using an agent-based model and mathematical analysis, we show that this simple mechanism robustly yields emergent regulation of foraging frequency. These findings demonstrate how the embedding of individuals in physical space can reduce their cognitive demands without compromising their computational role in the group.

Abstract Immune checkpoint inhibitors (ICI) with anti-PD-1/PD-L1 agents have improved the survival of patients with metastatic non-small cell lung cancer (mNSCLC). Tumor PD-L1 expression is an imperfect biomarker as it does not capture the complex interactions between constituents of the tumor microenvironment (TME). Using multiplex fluorescent immunohistochemistry (mfIHC), we modeled the TME to study the influence of cellular distribution and engagement on response to ICI in mNSCLC. We performed mfIHC on pretreatment tissue from patients with mNSCLC who received ICI. We used primary antibodies against CD3, CD8, CD163, PD-L1, pancytokeratin, and FOXP3; simple and complex phenotyping as well as spatial analyses was performed. We analyzed 68 distinct samples from 52 patients with mNSCLC. Patients were 39–79 years old (median 67); 44% were male and 75% had adenocarcinoma histology. The most used ICI was atezolizumab (48%). The percentage of PD-L1 positive epithelial tumor cells (EC), degree of cytotoxic T lymphocyte (CTL) engagement with EC, and degree of CTL engagement with helper T lymphocytes (HTL) were significantly lower in non-responders versus responders (p = 0.0163, p = 0.0026 and p = 0.0006, respectively). The combination of these 3 characteristics generated the best sensitivity and specificity to predict non-response to ICI and was also associated with shortened overall survival (p = 0.0271). The combination of low CTL engagement with EC and HTL along with low expression of EC PD-L1 represents a state of impaired endogenous immune reactivity. Together, they more precisely identified non-responders to ICI compared to PD-L1 alone and illustrate the importance of cellular interactions in the TME.

Abstract Spatial pattern modelling concepts are being increasingly used in capturing disease heterogeneity. Quantification of heterogeneity in the tumor microenvironment is extremely important in pancreatic ductal adenocarcinoma (PDAC), which has been shown to co-occur with other pancreatic diseases and neoplasms with certain attributes that make visual discrimination difficult. In this paper, we propose the GaWRDenMap framework, that utilizes the concepts of geographically weighted regression (GWR) and a density function-based classification model, and apply it to a cohort of multiplex immunofluorescence images from patients belonging to six different pancreatic diseases. We used an internal cohort of 228 patients comprised of 34 Chronic Pancreatitis (CP), 71 PDAC, 70 intraductal papillary mucinous neoplasm (IPMN), 16 mucinous cystic neoplasm (MCN), 29 pancreatic intraductal neoplasia (PanIN) and 8 IPMN-associated PDAC patients. We utilized GWR to model the relationship between epithelial cells and immune cells on a spatial grid. The GWR model estimates were used to generate density signatures which were used in subsequent pairwise classification models to distinguish between any two pairs of disease groups. Image-level, as well as subject-level analysis, were performed. When applied to this dataset, our classification model showed significant discrimination ability in multiple pairwise comparisons, in comparison to commonly used abundance-based metrics, like the Morisita-Horn index. The model was able to best discriminate between CP and PDAC at both the subject- and image-levels. It was also able to reasonably discriminate between PDAC and IPMN. These results point to a potential difference in the spatial arrangement of epithelial and immune cells between CP, PDAC and IPMN, that could be of high diagnostic significance. Further validation on a more comprehensive dataset would be warranted.

It has been suggested that the outer domains of bacterial flagellins are not needed for motility. Here, the authors show that flagellar filament outer domains from some bacteria have unique structures which can alter the motility of the bacteria.

Water scarcity associated with large-scale land acquisitions is exacerbated by adoption of water-intensive crops and expansion of irrigation, which in turn increases rival water uses.

IntroductionThe prevalence of Diabetic Kidney Disease (DKD) secondary to Type 2 Diabetes Mellitus (T2DM) is rising worldwide. However, real-world data linking glomerular function and albuminuria to the degree of multi-morbidity is lacking. We thus utilised the Discover dataset, to determine this association.MethodPatients with T2DM diagnosed prior to 1st January 2015 with no available biochemical evidence of CKD were included. Patients subsequently diagnosed and coded for CKD3a in 2015, were grouped by the degree of albuminuria. Baseline and 5-year co-morbidity was determined, as were prescribing practices with regards to prognostically beneficial medication.ResultsWe identified 56,261 patients with T2DM, of which 1082 had CKD stage 3a diagnosed in 2015 (224-CKD3aA1,154-CKD3aA2,93-CKD3aA1; 611 patients with CKD3a but no uACR available in 2015 were excluded from follow up). No statistically significant difference was observed in the degree of co-morbidities at baseline. A significant difference in the degree of hypertension, retinopathy, ischaemic heart disease and vascular disease from baseline compared to study end point was observed for all 3 study groups. Comparing co-morbidities developed at study end point, highlighted a statistical difference between CKD3aA1 Vs CKD3aA3 for retinopathy alone and for hypertension and heart failure between CKD3aA2 Vs CKD3aA3. 40.8% of patients with CKD3aA2 or A3 were prescribed Renin Angiotensin Aldosterone inhibitors (RAASi) therapy between June-December 2021. Survival analysis showed 15% of patients with CKD3aA3 developed CKD stage 5 within 5 years of diagnosis.DiscussionCKD3a secondary to DKD is associated with significant multimorbidity at baseline and 5 years post diagnosis, with CKD3aA3 most strongly associated with CKD progression to CKD 5, heart failure, hypertension and retinopathy compared to CKD3aA1 or CKD3aA2 at 5 years post diagnosis. The lack of uACR testing upon diagnosis and poor prescribing of RAASi, in those with CKD3aA2/A3, raises significant cause for concern.ConclusionDKD is associated with significant multimorbidity. Significant work is needed to be done to ensure patients undergo testing for uACR, to allow for future risk stratification and ability to be started on prognostically beneficial medication.

Protein activity regulated by phosphorylation can result in subcellular relocation. Here, the authors present a high throughput spatial phosphoproteomics approach to profile six subcellular compartments, providing insights into EGFR and stress signalling dynamics.

Summary: Absence seizures, manifested by spike-wave discharges (SWD) in the electroencephalogram, display synchronous reciprocal excitation between the neocortex and thalamus. Recent studies have revealed that inhibitory neurons in the reticular thalamic (RT) nucleus and excitatory thalamocortical (TC) neurons are two subcortical players in generating SWD. However, the signals that drive SWD-related activity remain elusive. Here, we show that SWD predominately occurs during wakefulness in several mouse models of absence epilepsy. In more focused studies of Gnb1 mutant mice, we found that sensory input regulates SWD. Using in vivo recording, we demonstrate that TC cells are activated prior to the onset of SWD and then inhibited during SWD. On the contrary, RT cells are slightly inhibited prior to SWD, but are strongly activated during SWD. Furthermore, chemogenetic activation of TC cells leads to the enhancement of SWD. Together, our results indicate that sensory input can regulate SWD by activating the thalamocortical pathway.

Mining for rare earth elements is rapidly increasing, driven by current and projected demands for information and energy technologies. Following China’s Central Government’s 2012 strategy to shift away from mining in favor of value-added processing, primary extraction has increased outside of China. Accordingly, changes in mineral exploitation in China and Myanmar have garnered considerable attention in the past decade. The prevailing assumption is that mining in China has decreased while mining in Myanmar has increased, but the dynamic in border regions is more complex. Our empirical study used Google Earth Engine (GEE) to characterize changes in mining surface footprints between 2005 and 2020 in two rare earth mines located on either side of the Myanmar–China border, within Kachin State in northern Myanmar and Nujiang Prefecture in Yunnan Province in China. Our results show that the extent of the mining activities increased by 130% on China’s side and 327% on Myanmar’s side during the study period. We extracted surface reflectance images from 2005 and 2010 from Landsat 5 TM and 2015 and 2020 images from Landsat 8 OLI. The Normalized Vegetation Index (NDVI) was applied to dense time-series imagery to enhance landcover categories. Random Forest was used to categorize landcover into mine and non-mine classes with an overall accuracy of 98% and a Kappa Coefficient of 0.98, revealing an increase in mining extent of 2.56 km2, covering the spatial mining footprint from 1.22 km2 to 3.78 km2 in 2005 and 2020, respectively, within the study area. We found a continuous decrease in non-mine cover, including vegetation. Both mines are located in areas important to ethnic minority groups, agrarian livelihoods, biodiversity conservation, and regional watersheds. The finding that mining surface areas increased on both sides of the border is significant because it shows that national-level generalizations do not align with local realities, particularly in socially and environmentally sensitive border regions. The quantification of such changes over time can help researchers and policymakers to better understand the shifting geographies and geopolitics of rare earth mining, the environmental dynamics in mining areas, and the particularities of mineral extraction in border regions.

Abstract Escherichia coli dynamics in urban watersheds are affected by a complex balance among external inputs, niche modulation and genetic variability. To explore the ecological processes influencing E. coli spatial patterns, we analyzed its abundance and phylogenetic structure in water samples from a stream network with heterogeneous urban infrastructure and environmental conditions. Our results showed that environmental and infrastructure variables, such as macrophyte coverage, DIN and sewerage density, mostly explained E. coli abundance. Moreover, main generalist phylogroups A and B1 were found in high proportion, which, together with an observed negative relationship between E. coli abundance and phylogroup diversity, suggests that their dominance might be due to competitive exclusion. Lower frequency phylogroups were associated with sites of higher ecological disturbance, mainly involving simplified habitats, higher drainage infrastructure and septic tank density. In addition to the strong negative relationship between phylogroup diversity and dominance, the occurrence of these phylogroups would be associated with increased facilitated dispersal. Nutrients also contributed to explaining phylogroup distribution. Our study proposes the differential contribution of distinct ecological processes to the patterns of E. coli in an urban watershed, which is useful for the monitoring and management of fecal pollution.

Summary: Cell-mediated immunity may contribute to providing protection against SARS-CoV-2 and its variants of concern (VOC). We developed COH04S1, a synthetic multiantigen modified vaccinia Ankara (MVA)-based COVID-19 vaccine that stimulated potent spike (S) and nucleocapsid (N) antigen-specific humoral and cellular immunity in a phase 1 clinical trial in healthy adults. Here, we show that individuals vaccinated with COH04S1 or mRNA vaccine BNT162b2 maintain robust cross-reactive cellular immunity for six or more months post-vaccination. Although neutralizing antibodies induced in COH04S1- and BNT162b2-vaccinees showed reduced activity against Delta and Omicron variants compared to ancestral SARS-CoV-2, S-specific T cells elicited in both COH04S1- and BNT162b2-vaccinees and N-specific T cells elicited in COH04S1-vaccinees demonstrated potent and equivalent cross-reactivity against ancestral SARS-CoV-2 and the major VOC. These results suggest that vaccine-induced T cells to S and N antigens may constitute a critical second line of defense to provide long-term protection against SARS-CoV-2 VOC.

Measurement(s) pitting potential • repassivation potential • Crevice corrosion potential • pitting temperature • Crevice corrosion temperature • corrosion current density • passive current density • corrosion rate • corrosion potential Technology Type(s) digital curation Factor Type(s) alloy composition • temperature • pH • chloride ion concentration • alloy microstructure • alloy heat treatment Machine-accessible metadata file describing the reported data: https://doi.org/10.6084/m9.figshare.13611623

Summary: Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder, in which an abnormal and toxic protein called progerin, accumulates in cell nuclei, leading to major cellular defects. Among them, chromatin remodeling drives gene expression changes, including miRNA dysregulation. In our study, we evaluated miRNA expression profiles in HGPS and control fibroblasts. We identified an enrichment of overexpressed miRNAs belonging to the 14q32.2-14q32.3 miRNA cluster. Using 3D FISH, we demonstrated that overexpression of these miRNAs is associated with chromatin remodeling at this specific locus in HGPS fibroblasts. We then focused on miR-376b-3p and miR-376a-3p, both overexpressed in HGPS fibroblasts. We demonstrated that their induced overexpression in control fibroblasts decreases cell proliferation and increases senescence, whereas their inhibition in HGPS fibroblasts rescues proliferation defects and senescence and decreases progerin accumulation. By targeting these major processes linked to premature aging, these two miRNAs may play a pivotal role in the pathophysiology of HGPS.

Genomic surveillance empowers agile responses to SARS-CoV-2 by enabling scientists and public health analysts to issue recommendations aimed at slowing transmission, prioritizing contact tracing, and building a robust genomic sequencing surveillance strategy. Since the start of the pandemic, real time RT-PCR diagnostic testing from upper respiratory specimens, such as nasopharyngeal (NP) swabs, has been the standard. Moreover, respiratory samples in viral transport media are the ideal specimen for SARS-CoV-2 whole-genome sequencing (WGS). In early 2021, many clinicians transitioned to antigen-based SARS-CoV-2 detection tests, which use anterior nasal swabs for SARS-CoV-2 antigen detection. Despite this shift in testing methods, the need for whole-genome sequence surveillance remains. Thus, we developed a workflow for whole-genome sequencing with antigen test-derived swabs as an input rather than nasopharyngeal swabs. In this study, we use excess clinical specimens processed using the BinaxNOW™ COVID-19 Ag Card. We demonstrate that whole-genome sequencing from antigen tests is feasible and yields similar results from RT-PCR-based assays utilizing a swab in viral transport media.

Agricultural large-scale land acquisitions have been linked with enhanced deforestation and land use change. Yet the extent to which transnational agricultural large-scale land acquisitions (TALSLAs) contribute to—or merely correlate with—deforestation, and the expected biodiversity impacts of the intended land use changes across ecosystems, remains unclear. We examine 178 georeferenced TALSLA locations in 40 countries to address this gap. While forest cover within TALSLAs decreased by 17% between 2000 and 2018 and became more fragmented, the spatio-temporal patterns of deforestation varied substantially across regions. While deforestation rates within initially forested TALSLAs were 1.5 (Asia) to 2 times (Africa) higher than immediately surrounding areas, we detected no such difference in Europe and Latin America. Our findings suggest that, whereas TALSLAs may have accelerated forest loss in Asia, a different mechanism might emerge in Africa where TALSLAs target areas already experiencing elevated deforestation. Regarding biodiversity (here focused on vertebrate species), we find that nearly all (91%) studied deals will likely experience substantial losses in relative species richness (−14.1% on average within each deal)—with mixed outcomes for relative abundance—due to the intended land use transitions. We also find that 39% of TALSLAs fall at least partially within biodiversity hotspots, placing these areas at heightened risk of biodiversity loss. Taken together, these findings suggest distinct regional differences in the nature of the association between TALSLAs and forest loss and provide new evidence of TALSLAs as an emerging threat to biodiversity in the Global South.

Abstract Carbapenem-resistance in Klebsiella pneumoniae (KP) sequence type ST258 is mediated by carbapenemases (e.g. KPC-2) and loss or modification of the major non-selective porins OmpK35 and OmpK36. However, the mechanism underpinning OmpK36-mediated resistance and consequences of these changes on pathogenicity remain unknown. By solving the crystal structure of a clinical ST258 OmpK36 variant we provide direct structural evidence of pore constriction, mediated by a di-amino acid (Gly115-Asp116) insertion into loop 3, restricting diffusion of both nutrients (e.g. lactose) and Carbapenems. In the presence of KPC-2 this results in a 16-fold increase in MIC to Meropenem. Additionally, the Gly-Asp insertion impairs bacterial growth in lactose-containing medium and confers a significant in vivo fitness cost in a murine model of ventilator-associated pneumonia. Our data suggests that the continuous selective pressure imposed by widespread Carbapenem utilisation in hospital settings drives the expansion of KP expressing Gly-Asp insertion mutants, despite an associated fitness cost.

Centrosomes drive mitotic spindle formation and chromosome segregation. Here, the authors show that centrosome stability is regulated by selective autophagic degradation of centriolar satellite components in a process they term doryphagy, connecting autophagy and chromosomal integrity.

Membraneless compartments have been theorized to be prebiotic micro-compartments as they spontaneously encapsulate RNA and proteins. Here, the authors report membraneless compartments can enhance RNA chemistries, affecting template directed RNA polymerization and stimulating nucleic acid enzymes.

IntroductionShigellosis, is a leading cause of moderate-to-severe diarrhoea and related mortality in young children in low and middle income countries (LMICs). Knowledge on naturally acquired immunity can support the development of Shigella candidate vaccines mostly needed in LMICs. We aimed to quantify Shigella-specific antibodies of maternal origin and those naturally acquired in Zambian infants.MethodsPlasma samples collected from infants at age 6, 14 and 52-weeks were tested for Shigella (S. sonnei and S. flexneri 2a) lipopolysaccharide (LPS) antigen specific immunoglobulin G (IgG) and A (IgA) by enzyme-linked immunosorbent assay.ResultsAt 6 weeks infant age, the IgG geometric mean titres (GMT) against S. sonnei (N = 159) and S. flexneri 2a (N = 135) LPS were 311 (95% CI 259-372) and 446 (95% CI 343-580) respectively. By 14 weeks, a decline in IgG GMT was observed for both S. sonnei to 104 (95% CI 88-124), and S. flexneri 2a to 183 (95% CI 147-230). Both S. sonnei and S. flexneri 2a specific IgG GMT continued to decrease by 52 weeks infant age when compared to 6 weeks. In 27% and 8% of infants a significant rise in titre (4 fold and greater) against S. flexneri 2a and S. sonnei LPS, respectively, was detected between the ages of 14 and 52 weeks. IgA levels against both species LPS were very low at 6 and 14 weeks and raised significantly against S. flexneri 2a and S. sonnei LPS in 29% and 10% of the infants, respectively.ConclusionIn our setting, transplacental IgG anti-Shigella LPS is present at high levels in early infancy, and begins to decrease by age 14 weeks. Our results are consistent with early exposure to Shigella and indicate naturally acquired IgG and IgA antibodies to S. flexneri 2a and S. sonnei LPS in part of infants between 14 and 52 weeks of age. These results suggest that a potential timing of vaccination would be after 14 and before 52 weeks of age to ensure early infant protection against shigellosis.

The evolutionary changes in immune profiles of triple negative breast cancer (TNBC) are not well understood, although it is known that immune checkpoint inhibitors have diminished activity in heavily pre-treated TNBC patients. This study was designed to characterize immune profile changes of longitudinal tumor specimens by studying immune subsets of tumor infiltrating lymphocytes (TILs) in paired primary and metastatic TNBC in a cohort of "poor outcome" (relapsed within 5 years) patients. Immune profiles of TNBCs in a cohort of "good outcome" (no relapse within 5 years) patients were also analyzed. Immune subsets were characterized for CD4, CD8, FOXP3, CD20, CD33, and PD1 using immuno-fluorescence staining in stroma, tumor, and combined stroma and tumor tissue. TIL subsets in "good outcome" versus "poor outcome" patients were also analyzed. Compared with primary, metastatic TNBCs had significantly lower TILs by hematoxylin and eosin (H&E) staining. Stromal TILs (sTILs), but not tumoral TILs (tTILs) had significantly reduced cytotoxic CD8+ T cells (CTLs), PD1+ CTLs, and total PD1+ TILs in metastatic compared with matched primary TNBCs. Higher PD1+ CTLs, PD1+CD4+ helper T cells (PD1+TCONV) and all PD1+ T cells in sTILs, tTILs and total stromal and tumor TILS (s+tTIL) were all associated with better prognosis. In summary, TIL subsets decrease significantly in metastatic TNBCs compared with matched primary. Higher PD1+ TILs are associated with better prognosis in early stage TNBCs. This finding supports the application of immune checkpoint inhibitors early in the treatment of TNBCs.

Infection of mice with Citrobacter rodentium is a common model of infection with attaching-and-effacing pathogens. Here, Connolly et al. analyse the transcriptome of C. rodentium during mouse infection, showing host-induced coordinated upregulation of virulence factors and 1,2-propanediol metabolism.

The concentration of divalent cations is well-known to influence molecular processes such as enzymatic activity and RNA folding. Here the authors attempt to reproduce physiological conditions for RNA folding in vitro and establish an important contribution of chelated magnesium ions in enhancing RNA function.

During the oocyte-to-embryo transition in C. elegans, multivesicular endosomes (MVEs) are formed rapidly. Using live imaging and electron microscopy, the authors here show that the ESCRT-III subunit Ist1 is required for normal MVE biogenesis in the early C. elegans embryo.

Lynch syndrome is characterized by predisposition to colorectal cancer and mutations in genes involved in mismatch repair. Here, the authors use whole genome sequencing and immunohistochemistry of mismatch repair proteins to show a high prevalence of Lynch syndrome in the Icelandic population.

The attributes of male acoustic advertisement displays are often related to a performer's age, breeding condition and motivation, but these relationships are particularly difficult to study in free-ranging marine mammals. For fin whale singers, we examined the relationships between a singer's swimming speed, song duration and amount of singing. We used a unique set of fin whale singing and swimming data collected in support of the US Navy's marine mammal monitoring programme associated with the Navy's Integrated Undersea Surveillance System. A goal of the programme is to improve understanding of the potential effects of anthropogenic sound sources on baleen whale behaviours and populations. We found that as whales swam faster, some continued to sing, while others did not. If swimming speed is an indication of male stamina, then singing while swimming faster could be a display by which females and/or other males assess a singer's physical fitness and potential reproductive quality. Results have implications for interpreting fin whale singing behaviour and the possible influences of anthropogenic sounds on fin whale mating strategies and breeding success.

The goal of this study is to characterize the genomic and immune profiles of metaplastic breast cancer (MpBC) and identify the association with survival through an analysis of archived tumor tissue. A next-generation sequencing-based mutational assay (Onco-48) was performed for 21 MpBC patients. Clinicopathologic characteristics were captured, including relapse free survival (RFS) and overall survival (OS). Immunohistochemistry (IHC) for CD3, CD4, CD8, and programmed death-ligand 1 (PD-L1) was also performed. Recurrence free survival (RFS) at 5 years was 57% (95% CI 0.34-0.75) and overall survival (OS) at 5 years was 66% (95% CI 0.41-0.82). The most commonly altered genes were TP53 (68.4%, 13/19), PIK3CA (42.1%, 8/19), and PTEN (15.8%, 3/19. For patients with PIK3CA mutations, RFS and OS were significantly worse than for those without (HR 5.6, 95% CI 1.33-23.1 and HR 8.0, 95% CI 1.53-41.7, respectively). Cox regression estimated that PD-L1 expression was associated with worse RFS and OS (HR 1.08, 95% CI 1.01-1.16 and HR 1.05, 95% CI 1.00-1.11, respectively, for an absolute increase in PD-L1 expression of 1%). In conclusion, PIK3CA mutation and PD-L1 expression confer poor prognosis in this cohort of patients with MpBC.

Traditional methods of assessing virtual water (VW) trade usually overestimate the foreign VW used by a country as the imported VW can also be re-exported and used by the countries other than the initial importer. Due to its ability to provide better estimates of transboundary resource use, the global value chain (GVC) method can provide more precise guidelines for the global policy debate over sustainable resource use. Here, we use GVC analysis based on multi-regional input-output tables to quantify the embodied trade of virtual blue and green water for China in the major agricultural sectors and agro-based industries. We find that China is a net importer of blue VW (2.9 billion m ^3 ) and green VW (57.9 billion m ^3 ) through agricultural commodities. Our results reveal that a large portion of imported blue (37%) and green (17%) VW is re-exported by China and consumed in other countries, representing the overestimated parts of China’s VW import use. These ratios are even higher for individual commodities, including 44% for blue VW for cotton and 22% for green VW for grains. This work demonstrates the importance of improved accounting in VW trade to achieve the sustainable use of global water resources and equitably share responsibility between producers, intermediates, and end-users.

Diets exercise great influence over both human and environmental health. While numerous efforts have sought to define and identify sustainable diets, there remains a poor understanding of the extent to which such shifts are feasible when taking into account local dietary preferences. Accounting for 40% of dietary calories and 46% of global cropland, cereals offer an important food group by which culturally appropriate dietary shifts may achieve large sustainability benefits. Here we combine country-specific information on dietary cereal supply with nutrient content values, CO _2 nutrient penalties, and environmental footprints to quantify the outcomes of adopting two feasible dietary shifts—maximizing the share of C4 cereals (e.g. maize, millet, sorghum) based on historical shares and increasing the share of whole grains. Our results show that increasing the share of whole grains can increase nutrient supply (+7% protein, +37% iron, +42% zinc) and overcome the nutrient-depleting effects of elevated CO _2 (eCO _2 ) and that maximizing the share of C4 cereals can substantially reduce environmental burden (−12% greenhouse gas emissions, −11% blue water demand), particularly in Africa and the Middle East. We also find that a combination of the two strategies would likely produce strong co-benefits between increased nutrient supply and reduced environmental impacts with mixed outcomes for offsetting the effects of eCO _2 . Such simultaneous improvements are particularly important for food insecure regions such as West Africa and Southeast Asia. These findings demonstrate important opportunities to identify sustainable diets that incorporate local preferences and cultural acceptability. Such considerations are essential when developing demand-side solutions to achieve more sustainable food systems.

Treatment responses of melanoma patients to MAPK pathway inhibitors are often limited. Here, the authors show that combining cardiac glycosides with MAPK inhibitors improves tumor regression by inducing intracellular acidification, mitochondrial calcium dysregulation, ATP depletion, and cell death.

Nearly one-sixth of U.S. river basins are unable to consistently meet societal water demands while also providing sufficient water for the environment. Water scarcity is expected to intensify and spread as populations increase, new water demands emerge, and climate changes. Improving water productivity by meeting realistic benchmarks for all water users could allow U.S. communities to expand economic activity and improve environmental flows. Here we utilize a spatially detailed database of water productivity to set realistic benchmarks for over 400 industries and products. We assess unrealized water savings achievable by each industry in each river basin within the conterminous U.S. by bringing all water users up to industry- and region-specific water productivity benchmarks. Some of the most water stressed areas throughout the U.S. West and South have the greatest potential for water savings, with around half of these water savings obtained by improving water productivity in the production of corn, cotton, and alfalfa. By incorporating benchmark-meeting water savings within a national hydrological model (WaSSI), we demonstrate that depletion of river flows across Western U.S. regions can be reduced on average by 6.2–23.2%, without reducing economic production. Lastly, we employ an environmentally extended input-output model to identify the U.S. industries and locations that can make the biggest impact by working with their suppliers to reduce water use ‘upstream’ in their supply chain. The agriculture and manufacturing sectors have the largest indirect water footprint due to their reliance on water-intensive inputs but these sectors also show the greatest capacity to reduce water consumption throughout their supply chains.

The type V intermediate filament lamins are the principal components of the nuclear matrix, including the nuclear lamina. Lamins are divided into A-type and B-type, which are encoded by three genes, LMNA, LMNB1, and LMNB2. The alternative splicing of LMNA produces two major A-type lamins, lamin A and lamin C. Previous studies have suggested that lamins are involved in cancer development and progression. A-type lamins have been proposed as biomarkers for cancer diagnosis, prognosis, and/or follow-up. The aim of the present study was to investigate lamins in cancer cells from metastatic pleural effusions using immunofluorescence, western blotting, and flow cytometry. In a sub-group of lung adenocarcinomas, we found reduced expression of lamin A but not of lamin C. The reduction in lamin A expression was correlated with the loss of epithelial membrane antigen (EMA)/MUC-1, an epithelial marker that is involved in the epithelial to mesenchymal transition (EMT). Finally, the lamin A expression was inversely correlated with the number of metastatic sites and the WHO Performance status, and association of pleural, bone and lung metastatic localizations was more frequent when lamin A expression was reduced. In conclusion, low lamin A but not lamin C expression in pleural metastatic cells could represent a major actor in the development of metastasis, associated with EMT and could account for a pejorative factor correlated with a poor Performance status.

Fluctuations in temperature and precipitation influence crop productivity across the planet. With episodes of extreme climate becoming increasingly frequent, buffering crop production against these stresses is a critical aspect of climate adaptation. In India, where grain production and diets are closely linked, national food supply is sensitive to the effect of climate variability on monsoon grain production. Here we quantitatively examine the historical (1966–2011) relationship between interannual variations in temperature and rainfall and rainfed yield variability for five monsoon crops—rice and four alternative grains (finger millet, maize, pearl millet, and sorghum). Compared to rice, we find that alternative grains are significantly less sensitive to climate variation and generally experience smaller declines in yield under climate extremes. However, maximizing harvested area allocations to coarse grains (i.e. holding maize production constant) reduced grain production by 12.0 Mtonnes (−17.2%) under drought conditions and 12.8 Mtonnes (−18.0%) during non-drought years (non-drought). Increasing the harvested area allocated to all alternative grains (i.e. including maize) can enhance production by +39.6% (drought) and by +37.0% (non-drought). These alternative grains therefore offer promise for reducing variations in Indian grain production in response to climate shocks, but avoiding grain production shortfalls from increased alternative grains will require yield improvements that do not compromise their superior climate resilience.

Racing pigeons are exposed to and act as carriers of diseases. Dietary protein requirement for their maintenance has not been determined experimentally despite their being domesticated for over 7000 years. A maintenance nitrogen (protein) requirement (MNR) for pigeons was determined in a balance study using diets containing 6, 10 and 14% crude protein (CP). Then, the effects of feeding the diets were investigated to determine whether they were adequate to sustain innate and acquired immune functions. Nitrogen intake from the 6% CP diet was sufficient to maintain nitrogen balance and body weight in pigeons. However, the immune functions of phagocytosis, oxidative burst and lymphocyte proliferation in pigeons fed this diet were reduced compared with those fed 10 and 14% CP diets. Pigeons given the 6 and 10% CP diets had lower antibody titres following inoculation against Newcastle disease (ND) than those on the 14% CP diet. A confounding factor found on autopsy was the presence of intestinal parasites in some of the pigeons given the 6 and 10% CP diets; however, none of the pigeons used to measure MNR or acquired immunity to ND were infested with parasites. In conclusion, neither the 6 nor 10% CP diets adequately sustained acquired immune function of pigeons.

The CyanoQ protein has been demonstrated to be a component of cyanobacterial Photosystem II (PS II), but there exist a number of outstanding questions concerning its physical association with the complex. CyanoQ is a lipoprotein; upon cleavage of its transit peptide by Signal Peptidase II, which targets delivery of the mature protein to the thylakoid lumenal space, the N-terminal cysteinyl residue is lipid-modified. This modification appears to tether this otherwise soluble component to the thylakoid membrane. To probe the functional significance of the lipid anchor, mutants of the CyanoQ protein have been generated in Synechocystis sp. PCC 6803 to eliminate the N-terminal cysteinyl residue, preventing lipid modification. Substitution of the N-terminal cysteinyl residue with serine (Q-C22S) resulted in a decrease in the amount of detectable CyanoQ protein to 17% that of the wild-type protein. Moreover, the physical properties of the accumulated Q-C22S protein were consistent with altered processing of the CyanoQ precursor. The Q-C22S protein was shifted to a higher apparent molecular mass and partitioned in the hydrophobic phase in TX-114 phase-partitioning experiments. These results suggest that the hydrophobic N-terminal 22 amino acids were not properly cleaved by a signal peptidase. Substitution of the entire CyanoQ transit peptide with the transit peptide of the soluble lumenal protein PsbO yielded the Q-SS mutant and resulted in no detectable accumulation of the modified CyanoQ protein. Finally, the CyanoQ protein was present at normal amounts in the PS II mutant strains ΔpsbB and ΔpsbO, indicating that an association with PS II was not a prerequisite for stable CyanoQ accumulation. Together these results indicate that CyanoQ accumulation in Synechocystis sp. PCC 6803 depends on the presence of the N-terminal lipid anchor, but not on the association of CyanoQ with the PS II complex.