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Book chapterGrob A, Enrico Bena C, Redwood-Sawyerr C, et al., 2024,
Simultaneous plate-reader characterization of promoter activity and cell growth in engineered mammalian cells
, Synthetic Promoters Methods and Protocols, Editors: Marchisio, Publisher: Humana, Pages: 85-96, ISBN: 9781071640623Automated high-throughput methods that support tracking of mammalian cell growth are currently needed to advance cell line characterization and identification of desired genetic components required for cell engineering. Here, we describe a high-throughput noninvasive assay based on plate reader measurements. The assay relies on the change in absorbance of the pH indicator phenol red. We show that its basic and acidic absorbance profiles can be converted into a cell growth index consistent with cell count profiles, and that, by adopting a computational pipeline and calibration measurements, it is possible to identify a conversion that enables prediction of cell numbers from plate measurements alone. The assay is suitable for growth characterization of both suspension and adherent cell lines when these are grown under different environmental conditions and treated with chemotherapeutic drugs. The method also supports characterization of stably engineered cell lines and identification of desired promoters based on fluorescence output.
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Journal articleRutter JW, Dekker L, Clare C, et al., 2024,
A bacteriocin expression platform for targeting pathogenic bacterial species
, NATURE COMMUNICATIONS, Vol: 15 -
Journal articleArmstrong E, Chiu MKL, Foo S, et al., 2024,
Combination of oncolytic Maraba virus with immune checkpoint blockade overcomes therapy resistance in an immunologically cold model of advanced melanoma with dysfunctional T-cell receptor signalling
, JOURNAL FOR IMMUNOTHERAPY OF CANCER, Vol: 12, Pages: 1-16 -
Journal articleGauci V, Pangala SR, Shenkin A, et al., 2024,
Global atmospheric methane uptake by upland tree woody surfaces
, Nature, Vol: 631, Pages: 796-800, ISSN: 0028-0836<jats:title>Abstract</jats:title><jats:p>Methane is an important greenhouse gas<jats:sup>1</jats:sup>, but the role of trees in the methane budget remains uncertain<jats:sup>2</jats:sup>. Although it has been shown that wetland and some upland trees can emit soil-derived methane at the stem base<jats:sup>3,4</jats:sup>, it has also been suggested that upland trees can serve as a net sink for atmospheric methane<jats:sup>5,6</jats:sup>. Here we examine in situ woody surface methane exchange of upland tropical, temperate and boreal forest trees. We find that methane uptake on woody surfaces, in particular at and above about 2 m above the forest floor, can dominate the net ecosystem contribution of trees, resulting in a net tree methane sink. Stable carbon isotope measurement of methane in woody surface chamber air and process-level investigations on extracted wood cores are consistent with methanotrophy, suggesting a microbially mediated drawdown of methane on and in tree woody surfaces and tissues. By applying terrestrial laser scanning-derived allometry to quantify global forest tree woody surface area, a preliminary first estimate suggests that trees may contribute 24.6–49.9 Tg of atmospheric methane uptake globally. Our findings indicate that the climate benefits of tropical and temperate forest protection and reforestation may be greater than previously assumed.</jats:p>
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Journal articleEwers RM, Orme CDL, Pearse WD, et al., 2024,
Thresholds for adding degraded tropical forest to the conservation estate
, Nature, Vol: 631, Pages: 808-813, ISSN: 0028-0836Logged and disturbed forests are often viewed as degraded and depauperate environments compared with primary forest. However, they are dynamic ecosystems1 that provide refugia for large amounts of biodiversity2,3, so we cannot afford to underestimate their conservation value4. Here we present empirically defined thresholds for categorizing the conservation value of logged forests, using one of the most comprehensive assessments of taxon responses to habitat degradation in any tropical forest environment. We analysed the impact of logging intensity on the individual occurrence patterns of 1,681 taxa belonging to 86 taxonomic orders and 126 functional groups in Sabah, Malaysia. Our results demonstrate the existence of two conservation-relevant thresholds. First, lightly logged forests (<29% biomass removal) retain high conservation value and a largely intact functional composition, and are therefore likely to recover their pre-logging values if allowed to undergo natural regeneration. Second, the most extreme impacts occur in heavily degraded forests with more than two-thirds (>68%) of their biomass removed, and these are likely to require more expensive measures to recover their biodiversity value. Overall, our data confirm that primary forests are irreplaceable5, but they also reinforce the message that logged forests retain considerable conservation value that should not be overlooked.
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Journal articleDavis D, Wizel A, Drier Y, 2024,
Accurate estimation of pathway activity in single cells for clustering and differential analysis.
, Genome Res, Vol: 34, Pages: 925-936Inferring which and how biological pathways and gene sets change is a key question in many studies that utilize single-cell RNA sequencing. Typically, these questions are addressed by quantifying the enrichment of known gene sets in lists of genes derived from global analysis. Here we offer SiPSiC, a new method to infer pathway activity in every single cell. This allows more sensitive differential analysis and utilization of pathway scores to cluster cells and compute UMAP or other similar projections. We apply our method to COVID-19, lung adenocarcinoma and glioma data sets, and demonstrate its utility. SiPSiC analysis results are consistent with findings reported in previous studies in many cases, but SiPSiC also reveals the differential activity of novel pathways, enabling us to suggest new mechanisms underlying the pathophysiology of these diseases and demonstrating SiPSiC's high accuracy and sensitivity in detecting biological function and traits. In addition, we demonstrate how it can be used to better classify cells based on activity of biological pathways instead of single genes and its ability to overcome patient-specific artifacts.
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Journal articleMartins LP, Stouffer DB, Blendinger PG, et al., 2024,
Birds optimize fruit size consumed near their geographic range limits
, SCIENCE, Vol: 385, Pages: 331-336, ISSN: 0036-8075 -
Journal articleGarner BH, Creedy TJ, Allan EL, et al., 2024,
The taxonomic composition and chronology of a museum collection of Coleoptera revealed through large-scale digitisation
, Frontiers in Ecology and Evolution, Vol: 12, ISSN: 2296-701XIntroduction: Historic museum collections hold a wealth of biodiversity data that are essential to our understanding of the rapidly changing natural world. Novel curatorial practices are needed to extract and digitise these data, especially for the innumerable pinned insects whose collecting information is held on small labels.Methods: We piloted semi-automated specimen imaging and digitisation of specimen labels for a collection of ~29,000 pinned insects of ground beetles (Carabidae: Lebiinae) held at the Natural History Museum, London. Raw transcription data were curated against literature sources and non-digital collection records. The primary data were subjected to statistical analyses to infer trends in collection activities and descriptive taxonomy over the past two centuries.Results: This work produced research-ready digitised records for 2,546 species (40% of known species of Lebiinae). Label information was available on geography in 91% of identified specimens, and the time of collection in 39.8% of specimens and could be approximated for nearly all specimens. Label data revealed the great age of this collection (average age 91.4 years) and the peak period of specimen acquisition between 1880 and 1930, with little differences among continents. Specimen acquisition declined greatly after about 1950. Early detected species generally were present in numerous specimens but were missing records from recent decades, while more recently acquired species (after 1950) were represented mostly by singleton specimens only. The slowing collection growth was mirrored by the decreasing rate of species description, which was affected by huge time lags of several decades to formal description after the initial specimen acquisition.Discussion: Historic label information provides a unique resource for assessing the state of biodiversity backwards to pre-industrial times. Many species held in historical collections especially from tropical super-diverse areas may not be discov
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Journal articleKourelis J, Schuster M, Demir F, et al., 2024,
Bioengineering secreted proteases converts divergent Rcr3 orthologs and paralogs into extracellular immune co-receptors
, PLANT CELL, Vol: 36, Pages: 3260-3276, ISSN: 1040-4651 -
Journal articleWan Y, Myall AC, Boonyasiri A, et al., 2024,
Integrated analysis of patient networks and plasmid genomes reveals a regional, multi-species outbreak of carbapenemase-producing Enterobacterales carrying both blaIMP and mcr-9 genes
, Journal of Infectious Diseases, Vol: 230, Pages: e159-e170, ISSN: 0022-1899BackgroundCarbapenemase-producing Enterobacterales (CPE) are challenging in healthcare, with resistance to multiple classes of antibiotics. This study describes the emergence of imipenemase (IMP)–encoding CPE among diverse Enterobacterales species between 2016 and 2019 across a London regional network.MethodsWe performed a network analysis of patient pathways, using electronic health records, to identify contacts between IMP-encoding CPE–positive patients. Genomes of IMP-encoding CPE isolates were overlaid with patient contacts to imply potential transmission events.ResultsGenomic analysis of 84 Enterobacterales isolates revealed diverse species (predominantly Klebsiella spp, Enterobacter spp, and Escherichia coli); 86% (72 of 84) harbored an IncHI2 plasmid carrying blaIMP and colistin resistance gene mcr-9 (68 of 72). Phylogenetic analysis of IncHI2 plasmids identified 3 lineages showing significant association with patient contacts and movements between 4 hospital sites and across medical specialties, which was missed in initial investigations.ConclusionsCombined, our patient network and plasmid analyses demonstrate an interspecies, plasmid-mediated outbreak of blaIMPCPE, which remained unidentified during standard investigations. With DNA sequencing and multimodal data incorporation, the outbreak investigation approach proposed here provides a framework for real-time identification of key factors causing pathogen spread. Plasmid-level outbreak analysis reveals that resistance spread may be wider than suspected, allowing more interventions to stop transmission within hospital networks.SummaryThis was an investigation, using integrated pathway networks and genomics methods, of the emergence of imipenemase-encoding carbapenemase-producing Enterobacterales among diverse Enterobacterales species between 2016 and 2019 in patients across a London regional hospital network, which was missed on routine investigations.
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Journal articleUkleja M, Kricks L, Torrens G, et al., 2024,
Flotillin-mediated stabilization of unfolded proteins in bacterial membrane microdomains
, Nature Communications, Vol: 15<jats:title>Abstract</jats:title><jats:p>The function of many bacterial processes depends on the formation of functional membrane microdomains (FMMs), which resemble the lipid rafts of eukaryotic cells. However, the mechanism and the biological function of these membrane microdomains remain unclear. Here, we show that FMMs in the pathogen methicillin-resistant <jats:italic>Staphylococcus aureus</jats:italic> (MRSA) are dedicated to confining and stabilizing proteins unfolded due to cellular stress. The FMM scaffold protein flotillin forms a clamp-shaped oligomer that holds unfolded proteins, stabilizing them and favoring their correct folding. This process does not impose a direct energy cost on the cell and is crucial to survival of ATP-depleted bacteria, and thus to pathogenesis. Consequently, FMM disassembling causes the accumulation of unfolded proteins, which compromise MRSA viability during infection and cause penicillin re-sensitization due to PBP2a unfolding. Thus, our results indicate that FMMs mediate ATP-independent stabilization of unfolded proteins, which is essential for bacterial viability during infection.</jats:p>
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Journal articleSaranholi BH, Franca FM, Vogler AP, et al., 2024,
Testing and optimizing metabarcoding of iDNA from dung beetles to sample mammals in the hyperdiverse Neotropics
, MOLECULAR ECOLOGY RESOURCES, Vol: 24, ISSN: 1755-098X -
Journal articleChia K-S, Kourelis J, Teulet A, et al., 2024,
The N-terminal domains of NLR immune receptors exhibit structural and functional similarities across divergent plant lineages
, The Plant Cell, Vol: 36, Pages: 2491-2511, ISSN: 1040-4651Nucleotide-binding domain and leucine-rich repeat (NLR) proteins are a prominent class of intracellular immune receptors in plants. However, our understanding of plant NLR structure and function is limited to the evolutionarily young flowering plant clade. Here, we describe an extended spectrum of NLR diversity across divergent plant lineages and demonstrate the structural and functional similarities of N-terminal domains that trigger immune responses. We show that the broadly distributed coiled-coil (CC) and toll/interleukin-1 receptor (TIR) domain families of non-flowering plants retain immune-related functions through trans-lineage activation of cell death in the angiosperm Nicotiana benthamiana. We further examined a CC subfamily specific to non-flowering lineages and uncovered an essential N-terminal MAEPL motif that is functionally comparable to motifs in resistosome-forming CC-NLRs. Consistent with a conserved role in immunity, the ectopic activation of CCMAEPL in the non-flowering liverwort Marchantia polymorpha led to profound growth inhibition, defense gene activation, and signatures of cell death. Moreover, comparative transcriptomic analyses of CCMAEPL activity delineated a common CC-mediated immune program shared across evolutionarily divergent non-flowering and flowering plants. Collectively, our findings highlight the ancestral nature of NLR-mediated immunity during plant evolution that dates its origin to at least ∼500 million years ago.
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Journal articleMoreno-Contreras I, Jokimaki J, Kaisanlahti-Jokimaki M-L, et al., 2024,
Disentangling the drivers of urban bird diversity in the non-breeding season: A general synthesis
, GLOBAL CHANGE BIOLOGY, Vol: 30, ISSN: 1354-1013 -
Journal articleHowes B, Gonzalez-Suarez M, Banks-Leite C, et al., 2024,
A global latitudinal gradient in the proportion of terrestrial vertebrate forest species
, GLOBAL ECOLOGY AND BIOGEOGRAPHY, Vol: 33, ISSN: 1466-822X -
Journal articleSimpson EG, Fraser I, Woolf H, et al., 2024,
Variation in near-surface soil temperature drives plant assemblage differentiation across aspect
, ECOLOGY AND EVOLUTION, Vol: 14, ISSN: 2045-7758 -
Journal articleDunning J, Sanchez-Tojar A, Girndt A, et al., 2024,
Extrapair paternity alongside social reproduction increases male lifetime fitness
, Animal Behaviour, Vol: 213, Pages: 117-123, ISSN: 0003-3472Within breeding years, male birds vary in their reproductive strategy. While some maintain monogamy with a social partner, others also engage with extrapair partners, while others forgo monogamy altogether in favour of exclusively seeking extrapair paternity. Although theory predicts that extrapair paternity is beneficial to males, which sire extrapair offspring without investing in costly parental care, empirical examples from wild populations are sparse. We used 17 years of data from a closed population of house sparrows, Passer domesticus, with a complete genetic pedigree, to test the hypothesis that extrapair paternity increases male lifetime reproductive success. We compared a mixed strategy of within-pair (or social) and extrapair paternity with total genetic monogamy and total extrapair paternity. We demonstrate that males who combine within-pair and extrapair paternity have increased reproductive success against the other two groups. Our results also suggest that males that exclusively seek extrapair paternity have the lowest lifetime fitness. Overall, we provide an empirical demonstration of the theory, showing that where males can sire extrapair offspring alongside within-pair offspring, extrapair paternity is beneficial to male lifetime fitness.
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Journal articleLi Z, Di Vagno L, Chawla H, et al., 2024,
Xylosyltransferase Bump-and-hole Engineering to Chemically Manipulate Proteoglycans in Mammalian Cells.
, bioRxivMammalian cells orchestrate signalling through interaction events on their surfaces. Proteoglycans are an intricate part of these interactions, carrying large glycosaminoglycan polysaccharides that recruit signalling molecules. Despite their importance in development, cancer and neurobiology, a relatively small number of proteoglycans have been identified. In addition to the complexity of glycan extension, biosynthetic redundancy in the first protein glycosylation step by two xylosyltransferase isoenzymes XT1 and XT2 complicates annotation of proteoglycans. Here, we develop a chemical genetic strategy that manipulates the glycan attachment site of cellular proteoglycans. By employing a tactic termed bump- and-hole engineering, we engineer the two isoenzymes XT1 and XT2 to specifically transfer a chemically modified xylose analogue to target proteins. The chemical modification contains a bioorthogonal tag, allowing the ability to visualise and profile target proteins modified by both transferases in mammalian cells. The versatility of our approach allows pinpointing glycosylation sites by tandem mass spectrometry, and exploiting the chemical handle to manufacture proteoglycans with defined GAG chains for cellular applications. Engineered XT enzymes permit a view into proteoglycan biology that is orthogonal to conventional techniques in biochemistry.
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Journal articleTica J, Chen H, LUO S, et al., 2024,
Engineering tunable, low latency spatial computation with dual input quorum sensing promoters
, ACS Synthetic Biology, Vol: 13, Pages: 1750-1761, ISSN: 2161-5063Quorum sensing signals have evolved for population-level signaling in bacterial communities and are versatile tools for engineering cell–cell signaling in synthetic biology projects. Here, we characterize the spatial diffusion of a palette of quorum sensing signals and find that their diffusion in agar can be predicted from their molecular weight with a simple power law. We also engineer novel dual- and multi-input promoters that respond to quorum-sensing diffusive signals for use in engineered genetic systems. We engineer a promoter scaffold that can be adapted for activation and repression by multiple diffusers simultaneously. Lastly, we combine the knowledge on diffusion dynamics with the novel genetic components to build a new generation of spatial, stripe-forming systems with a simplified design, improved robustness, tuneability, and response time.
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Journal articleEndres R, Matas-Gil A, 2024,
Unraveling biochemical spatial patterns: machine learning approaches to the inverse problem of stationary Turing patterns
, iScience, Vol: 27, ISSN: 2589-0042The diffusion-driven Turing instability is a potential mechanism for spatial pattern formation in numerous biological and chemical systems. However, engineering these patterns and demonstrating that they are produced by this mechanism is challenging. To address this, we aim to solve the inverse problem in artificial and experimental Turing patterns. This task is challenging since patterns are often corrupted by noise and slight changes in initial conditions can lead to different patterns. We used both least squares to explore the problem and physics-informed neural networks to build a noise-robust method. We elucidate the functionality of our network in scenarios mimicking biological noise levels and showcase its application using an experimentally obtained chemical pattern. The findings reveal the significant promise of machine learning in steering the creation of synthetic patterns in bioengineering, thereby advancing our grasp of morphological intricacies within biological systems while acknowledging existing limitations.
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Journal articleCollins ASP, Kurt H, Duggan C, et al., 2024,
Parallel, continuous monitoring and quantification of programmed cell death in plant tissue
, Advanced Science, Vol: 11, ISSN: 2198-3844Accurate quantification of hypersensitive response (HR) programmed cell death is imperative for understanding plant defense mechanisms and developing disease-resistant crop varieties. Here, a phenotyping platform for rapid, continuous-time, and quantitative assessment of HR is demonstrated: Parallel Automated Spectroscopy Tool for Electrolyte Leakage (PASTEL). Compared to traditional HR assays, PASTEL significantly improves temporal resolution and has high sensitivity, facilitating detection of microscopic levels of cell death. Validation is performed by transiently expressing the effector protein AVRblb2 in transgenic Nicotiana benthamiana (expressing the corresponding resistance protein Rpi-blb2) to reliably induce HR. Detection of cell death is achieved at microscopic intensities, where leaf tissue appears healthy to the naked eye one week after infiltration. PASTEL produces large amounts of frequency domain impedance data captured continuously. This data is used to develop supervised machine-learning (ML) models for classification of HR. Input data (inclusive of the entire tested concentration range) is classified as HR-positive or negative with 84.1% mean accuracy (F1 score = 0.75) at 1 h and with 87.8% mean accuracy (F1 score = 0.81) at 22 h. With PASTEL and the ML models produced in this work, it is possible to phenotype disease resistance in plants in hours instead of days to weeks.
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Journal articleJoyce M, Falconio FA, Blackhurst L, et al., 2024,
Divergent evolution of sleep in Drosophila species
, Nature Communications, Vol: 15, ISSN: 2041-1723Living organisms synchronize their biological activities with the earth’s rotation through the circadian clock, a molecular mechanism that regulates biology and behavior daily. This synchronization factually maximizes positive activities (e.g., social interactions, feeding) during safe periods, and minimizes exposure to dangers (e.g., predation, darkness) typically at night. Beyond basic circadian regulation, some behaviors like sleep have an additional layer of homeostatic control, ensuring those essential activities are fulfilled. While sleep is predominantly governed by the circadian clock, a secondary homeostatic regulator, though not well-understood, ensures adherence to necessary sleep amounts and hints at a fundamental biological function of sleep beyond simple energy conservation and safety. Here we explore sleep regulation across seven Drosophila species with diverse ecological niches, revealing that while circadian-driven sleep aspects are consistent, homeostatic regulation varies significantly. The findings suggest that in Drosophilids, sleep evolved primarily for circadian purposes. The more complex, homeostatically regulated functions of sleep appear to have evolved independently in a species-specific manner, and are not universally conserved. This laboratory model may reproduce and recapitulate primordial sleep evolution.
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Journal articleMiao A, Luo T, Hsieh B, et al., 2024,
Brain clearance is reduced during sleep and anesthesia (vol 27, pg 1046, 2024)
, NATURE NEUROSCIENCE, ISSN: 1097-6256 -
Journal articleNkumama IN, Ogwang R, Odera D, et al., 2024,
Breadth of Fc-mediated effector function correlates with clinical immunity following human malaria challenge
, IMMUNITY, Vol: 57, ISSN: 1074-7613 -
Journal articleHaber DA, Arien Y, Lamdan LB, et al., 2024,
Targeting mosquito X-chromosomes reveals complex transmission dynamics of sex ratio distorting gene drives
, NATURE COMMUNICATIONS, Vol: 15 -
Journal articleSmith T, Mishra S, Dorigatti I, et al., 2024,
Differential responses of SARS-CoV-2 variants to environmental drivers during their selective sweeps
, Scientific Reports, Vol: 14, ISSN: 2045-2322Previous work has shown that environmental variables affect SARS-CoV-2 transmission, but it is unclear whether different strains show similar environmental responses. Here we leverage genetic data on the transmission of three (Alpha, Delta and Omicron BA.1) variants of SARS-CoV-2 throughout England, to unpick the roles that climate and public-health interventions play in the circulation of this virus. We find evidence for enhanced transmission of the virus in colder conditions in the first variant selective sweep (of Alpha, in winter), but limited evidence of an impact of climate in either the second (of Delta, in the summer, when vaccines were prevalent) or third sweep (of Omicron, in the winter, during a successful booster-vaccination campaign). We argue that the results for Alpha are to be expected if the impact of climate is non-linear: we find evidence of an asymptotic impact of temperature on the alpha variant transmission rate. That is, at lower temperatures, the influence of temperature on transmission is much higher than at warmer temperatures. As with the initial spread of SARS-CoV-2, however, the overwhelming majority of variation in disease transmission is explained by the intrinsic biology of the virus and public-health mitigation measures. Specifically, when vaccination rates are high, a major driver of the spread of a new variant is it’s ability to evade immunity, and any climate effects are secondary (as evidenced for Delta and Omicron). Climate alone cannot describe the transmission dynamics of emerging SARS-CoV-2 variants.
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Journal articleBaxter JM, Hutchison CDM, Fadini A, et al., 2024,
Power Density Titration of Reversible Photoisomerization of a Fluorescent Protein Chromophore in the Presence of Thermally Driven Barrier Crossing Shown by Quantitative Millisecond Serial Synchrotron X-ray Crystallography
, JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, Vol: 146, Pages: 16394-16403, ISSN: 0002-7863 -
Journal articleFu Z, Ciais P, Wigneron J-P, et al., 2024,
Global critical soil moisture thresholds of plant water stress
, Nature Communications, Vol: 15, ISSN: 2041-1723During extensive periods without rain, known as dry-downs, decreasing soil moisture (SM) induces plant water stress at the point when it limits evapotranspiration, defining a critical SM threshold (θcrit). Better quantification of θcrit is needed for improving future projections of climate and water resources, food production, and ecosystem vulnerability. Here, we combine systematic satellite observations of the diurnal amplitude of land surface temperature (dLST) and SM during dry-downs, corroborated by in-situ data from flux towers, to generate the observation-based global map of θcrit. We find an average global θcrit of 0.19 m3/m3, varying from 0.12 m3/m3 in arid ecosystems to 0.26 m3/m3 in humid ecosystems. θcrit simulated by Earth System Models is overestimated in dry areas and underestimated in wet areas. The global observed pattern of θcrit reflects plant adaptation to soil available water and atmospheric demand. Using explainable machine learning, we show that aridity index, leaf area and soil texture are the most influential drivers. Moreover, we show that the annual fraction of days with water stress, when SM stays below θcrit, has increased in the past four decades. Our results have important implications for understanding the inception of water stress in models and identifying SM tipping points.
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Journal articleLane BJ, Ma Y, Yan N, et al., 2024,
Monitoring the conformational ensemble and lipid environment of a mechanosensitive channel under cyclodextrin-induced membrane tension
, STRUCTURE, Vol: 32, ISSN: 0969-2126 -
Journal articleCawood EE, Baker E, Edwards TA, et al., 2024,
Understanding β-strand mediated protein-protein interactions: tuning binding behaviour of intrinsically disordered sequences by backbone modification
, CHEMICAL SCIENCE, ISSN: 2041-6520
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