Browse Articles

Employing deep mutational scanning in the <i>Escherichia coli</i> periplasm to decode the thermodynamic landscape for amyloid formation

Deep mutational scanning (DMS) assays provide a powerful method to generate large-scale datasets essential for advancing AI-driven predictions in biology. The tripartite β-lactamase assay (TPBLA), in which a protein of interest is inserted between two domains of β-lactamase, has previously been reported as capable of detecting and quantitating the aggregation of proteins and biologics in the oxidizing periplasm of Escherichia coli and used as a platform for identifying small molecule inhibitors of aggregation. Here, we repurpose the TPBLA into a high-throughput DMS platform. We validate this format using a single-site saturation library of the intrinsically disordered peptide Aβ 42 , linked to Alzheimer’s disease, demonstrating strong agreement between observed variant fitness scores and variant behavior using our previously reported low-throughput TPBLA. The results of DMS revealed variant fitness scores that correlate with known amyloid-promoting regions. An in silico approach using FoldX-derived per-residue thermodynamic stability confirmed that the TPBLA reports on amyloid fibril stability. In vitro experiments support this finding, showing a strong correlation between variant fitness scores and the critical concentration of amyloid formation. Machine learning using the DMS dataset identified β‐sheet propensity and polarity as primary drivers of variant fitness scores. The derived model is also able to predict thermodynamically stabilizing regions in other amyloid systems, underscoring its generalizability. Collectively, our results demonstrate the TPBLA as a versatile platform for generating robust datasets to advance predictive modeling and to inform the design of aggregation‐resistant proteins.

Remembering ecologist Margaret Bryan Davis (1931–2024)

Margaret Bryan Davis, a prominent ecologist, died on May 22, 2024. She was a Regents Professor at the University of Minnesota and a member of the National Academy of Sciences. Her students remember her as a creative and clear-thinking scientist who combined the tools and perspectives of paleoecology with ideas and problems of ecology in order to understand long-term forest dynamics. She was a champion of diversity and equity in science.

Correction for Berenbaum, Political and biological reality checks

Phonons in electron crystals with Berry curvature

Recent advances in 2D materials featuring nonzero Berry curvature have inspired extensions of the Wigner crystallization paradigm. This paper derives a low-energy effective theory for such quantum crystals, including the anomalous Hall crystal (AHC) with nonzero Chern number. First, we show that the low frequency dispersion of phonons in AHC, despite the presence of Berry curvature, resembles that of the zero field (rather than finite magnetic field) Wigner crystal due to the commutation of translation generators. We explain how key parameters of the phonon theory such as elastic constants and effective mass can be extracted from microscopic models, and apply them to two families of models: the recently introduced λ -jellium model and a model of rhombohedral multilayer graphene (RMG). In the λ -jellium model, we explore the energy landscape as crystal geometry shifts, revealing that AHC can become “soft” under certain conditions. This causes transitions in lattice geometry, although the quantized Hall response remains unchanged. Surprisingly, the Berry curvature seems to enhance the effective mass, leading to a reduction in phonon speed. For the AHC in RMG, we obtain estimates of phonon speed and shear stiffness. We also identify a previously overlooked “kineo-elastic” term in the phonon effective action that is present in the symmetry setting of RMG, and leads to dramatic differences in phonon speeds in opposite directions. We numerically confirm these predictions of the effective actions by time-dependent Hartree–Fock calculations.

Correction for Loop Yao et al., Synergistic action of specialized metabolites from divergent biosynthesis in the human oral microbiome

Endothelial αvβ3 integrin induction during hypoxia protects blood–brain barrier integrity

The blood–brain barrier (BBB) is critical for maintaining cerebral homeostasis, and its deterioration with age is an important pathogenic factor in the etiology of vascular dementia. Extracellular matrix–integrin interactions play a central role in regulating vascular stability. The αvβ3 integrin is not expressed by brain endothelial cells under stable conditions but is strongly induced by hypoxia. However, it is currently unclear whether αvβ3 integrin exerts a destructive or protective influence on BBB integrity. In young (8 to 10 wk) and aged (20 mo) mice, we examined the impact of a function-blocking β3 integrin antibody as well as the inhibitory peptide cilengitide on BBB disruption during exposure to CMH (8% O 2 ). Hypoxic induction of brain endothelial β3 integrin was much stronger in aged mice. In both young and aged mice, β3 integrin inhibition greatly amplified hypoxia-induced BBB disruption, correlating with loss of tight junction proteins and induction of the leaky BBB marker mouse endothelial cell antigen (MECA)-32. Consistent with this, β3 integrin null mice showed increased levels of hypoxia-induced BBB disruption and MECA-32 expression. Cilengitide also reduced the integrity of a brain endothelial monolayer in vitro, prevented β3 integrin localization to focal adhesions, and reduced expression of vascular endothelial (VE)-cadherin and tight junction proteins. These observations suggest that hypoxic induction of endothelial αvβ3 integrin enhances BBB integrity by stabilizing endothelial adhesion. This raises the interesting possibility that pharmacological upregulation of endothelial αvβ3 integrin in the aged brain might hold therapeutic promise for vascular dementia.

Flow cytometry protocol for cell death analysis in glioblastoma organoids: A technical note

Tumor organoid models have emerged as a promising tool in cancer research. By preserving intra- and intertumoral heterogeneity and structural integrity they provide a physiologically relevant platform for drug-response studies. However, valid methodological approaches for cell death analyses applying flow cytometry, particularly in complex, large organoids, are lacking. Using glioblastoma organoids (GBOs), we developed a flow cytometry protocol to quantify cell death as an important readout in cancer research. Human GBOs were generated out of tumor material from six patients. Temozolomide (TMZ) and lomustine (CCNU) were used as cytotoxic agents commonly employed in glioblastoma therapy. After treatment for 144 and 288 hours, single cell suspensions from densely-packed GBOs were generated through a combined approach of enzymatic and mechanical dissociation. Cells were permeabilized with Triton X and subsequently stained with propidium iodide (PI). PI staining labels fragmented nuclear DNA, yielding a hypodiploid sub-G1 peak in flow cytometry that markes cell death. After treatment for 288 hours with physiologically-relevant concentrations of TMZ and CCNU cell death rates reached up to 63% in our GBO model. Across three GBO populations, the impact of CCNU at the given concentration was more pronounced compared to that observed with TMZ and the cell death rates of treatment for 288 hours surpassed that of the 144-hour treatment. Both biological and technical replicates showed low variability. Hoechst 33258 staining on the same samples confirmed trends in cell death rates obtained from PI-based analysis. We further validated the treatment-induced effect using a plate-based lactate dehydrogenase release assay and measurements of GBO diameter. Our single-stain flow-cytometry protocol scales to large, dense organoids and provides a practical balance of performance, hands-on time, cost, specificity, and throughput. This protocol could support development and evaluation of subtype-specific therapeutic strategies in translational cancer research.

Pseudouridine prevalence in Kaposi’s sarcoma–associated herpesvirus transcriptome reveals an essential mechanism for viral replication

Pseudouridylation is a prevalent RNA modification occurring in transfer RNAs (tRNAs), ribosomal RNAs (rRNAs), small non-coding RNAs (snoRNAs), and has been most recently identified in mRNAs and lncRNAs. Emerging evidence suggests that this dynamic RNA modification is implicated in altering gene expression by regulating RNA stability, modulating translation elongation, and modifying amino acid substitution rates. However, the role of pseudouridylation in infection is poorly understood. Herein, we demonstrate that Kaposi’s sarcoma–associated herpesvirus (KSHV) manipulates the pseudouridylation pathway to enhance replication. We show that the pseudouridine synthases (PUS), PUS1, and PUS7 are essential for efficient KSHV lytic replication, supported by their redistribution to viral replication and transcription complexes. We present a comprehensive analysis of KSHV RNA pseudouridylation, revealing hundreds of modified RNAs at single-nucleotide resolution. Notably, we demonstrate that pseudouridylation of the KSHV-encoded polyadenylated nuclear RNA (PAN) plays a significant role in the expression of PAN RNA. These findings reveal an essential role of pseudouridine modification in the KSHV replication cycle.

The polarization of literary censorship in the U.S

Literary censorship is not new, but the recent resurgence in the U.S. differs from earlier post-war controversies, many of which were bipartisan and focused on protecting children from exposure to sexual and graphic material. Current censorship controversies in the U.S. appear to be much more partisan and ideological, focused on protecting children from politically offensive ideas. Anecdotally, the political right is portrayed as attacking literature in the name of conservative values like preserving the traditional family from alternative expressions of sexuality and sexual preference, with the left targeting material regarded as contrary to progressive values like tolerance of diverse cultural identities. However, these ideological rationales largely reflect the rhetoric of political activists and have not been empirically tested among the broader public. We conducted two studies designed to measure the polarization of support for literary censorship among the voting-age population of the U.S. Surprisingly, both studies cast doubt on the ideological divisions apparent in activist rhetoric. The survey findings from Study 1 indicate widespread opposition to literary censorship that spans ideological divisions, but both liberals and conservatives were more inclined to support censorship of materials that deviated from their respective values and beliefs. The experiment in Study 2 revealed differences in participants’ responses to liberal and conservative criticisms but little difference in the attitudes of liberal and conservative participants. However, liberals were marginally more likely than conservatives to agree with ideologically aligned literary criticism.

<i>Bacillus subtilis</i> σ ^A and <i>Escherichia coli</i> σ ⁷⁰ lacking σ region 1.1 are not released during transcription initiation and elongation

A “σ cycle” in which the initiation factor σ associates with RNA polymerase (RNAP) core enzyme to permit transcription initiation and dissociates from RNAP core enzyme to permit transcription elongation, has been proposed to occur and to be an essential step for σ-exchange, with all principal σ factors from all bacteria. These proposals were based on studies of the principal σ factor of Escherichia coli , σ 70 , which generally, albeit not obligatorily, is released from RNAP upon the transition from transcription initiation to elongation. Here, we show that, in contrast to E. coli σ 70 , the Bacillus subtilis principal σ factor, σ A , is not released and is retained on RNAP core throughout transcription elongation. We further show that a mutant E. coli σ 70 derivative lacking σ region 1.1 (σ R1.1) is not released and is retained on RNAP core throughout transcription elongation. We also observe that B. subtilis σ A and the mutant E. coli σ 70 derivative lacking σ R1.1 interact much more stably with RNAP than full-length E. coli σ 70 . Our results indicate that the σ cycle is not a universal phenomenon in bacteria.

Dynamics and diversity in adolescents’ experienced barriers and facilitators for physical activity maintenance

Despite the well-documented health benefits of physical activity (PA), over 80% of adolescents worldwide fail to meet recommended daily levels. This study identified experienced barriers and facilitators for PA maintenance among Dutch adolescents, examine how they form barrier and facilitator profiles, and explore how barriers and facilitators evolve over time. First, we conducted 21 interviews with adolescents (13–16 years) to uncover relevant barriers and facilitators. Then, we developed and applied a card sorting task based on Q methodology, and examined barrier and facilitator configurations with 30 adolescents (aged 13–18 years) who had maintained a physical activity for ≥2 years, followed by interviews. Factor analysis revealed five facilitator profiles: Mental and physical health benefits, A way to be myself around others, Pursuing health goals, Developing confidence and strength, and Developing along my own path. Four barrier profiles emerged: Low motivation and energy, Not a good fit for me, Balancing act, and Proximity, possibility and perception. Facilitator profiles ranged from immediate characteristics of the activity, such as enjoyment, social connection, and mental well being, to more future-oriented drivers such as autonomy and self-development. Barriers profiles varied from predominantly internal (e.g., low motivation) to external (e.g., distance, weather) or mixed influences linking life demands to reduced personal resources. Across participants, enjoyment was the most consistent facilitator, but perceived influences often shifted with time, from immediate, activity-based facilitators related to competence and relatedness, toward motivations tied to autonomy, identity and coping with growing responsibilities characteristic of this life stage. These results highlight the diversity and dynamic nature of barriers and facilitators in adolescent PA maintenance. Tailoring PA promotion programs to adolescents’ evolving motivations and constraints can increase their effectiveness, supporting sustained active lifestyles into adulthood.

Distinct timescales dissociate spontaneous thought dimensions

Our spontaneous thoughts encompass various dimensions, such as task-relatedness (off vs. on task), and thought orientation (internal vs. external). However, their distinction remains unclear. Our study addresses this issue by focusing on their timescales at both the behavioral level (using fast and slow finger tapping) and the neural level (using EEG) using two independent datasets (N = 84 and 35). Behavioral results revealed a double dissociation: Task-relatedness was linked to fast tapping only, whereas thought orientation was associated with slow tapping only. At the neural level, we assessed topographic similarity in EEG to quantify the temporal influence of past neural activity on current ones. Task-relatedness modulated topographic similarity only during fast tapping, while thought orientation did so only during slow tapping. Critically, topographic similarity was phase-based, as shown by its correlation with phase-locking value and the loss of associations with thought after phase-shuffling. This indicates that the neural signatures of both thought dimensions are strongly phase-dependent. Finally, we demonstrate that nonlinearity plays a distinct role mediating the impact of different timescales (slow and fast finger tapping) on spontaneous thoughts at both behavioral (precision error) and neural levels (topographic similarity). Overall, these results demonstrate that task-relatedness is associated with short timescales, whereas thought orientation is associated with long timescales. This highlights how distinct temporal dynamics shape different spontaneous thought dimensions on both their behavioral and neural features.

Correction: Genetic variations associated with adaptation in Acrocomia palms: A comparative study across the Neotropics for crop improvement

Discovering neural elastoplasticity from kinematic observations

Inferring accurate and precise material models necessary for high-fidelity predictions has been a central challenge in constitutive modeling. Both traditional regression methods and modern machine-learning approaches require specialized data labels, which often cannot be sufficiently obtained from experiments. This data demand makes many sophisticated models impractical for real-world problems. Improvements in digital image correlation techniques have enabled accurate measurements of displacement data, providing an alternative kinematics-based approach for model identification. However, for materials undergoing plastic deformation, fracture, and damage, the corresponding inverse problem could be inherently challenging due to the dependence on loading history. We overcome this by formulating an inverse problem to discover interpretable plasticity models parameterized by neural networks (NN) from kinematic observations, leveraging a differentiable simulator with a smooth constitutive update that enables backpropagation for the NN training. The ability to use kinematic observations to infer complex material models may pave the way for a massive generation of material models that can be game-changing for emerging applications such as the design of metamaterials, response surface analyses, and the design of experiments.

Exploration of association rule mining between lost-linking features and modes of loan customers using the FP-growth algorithm for risk warning strategies

In the new model of China’s dual-circulation economy, the opening-up and deepening of financial markets have imposed higher requirements on the risk management capacity of financial institutions, with the issue of loan customers losing contact and defaulting becoming an urgent concern. Based on desensitized samples of lost-linking customers (with multidimensional features such as communication behavior and loan qualifications), this study uses the FP-Growth algorithm to systematically mine association rules between loss-of-contact features and three modes: “Hide and Seek”, “Flee with the Money”, and “False Disappearance”, providing effective risk management strategies for financial institutions. Through association rule mining, this study reveals significant correlations between some feature combinations and lost-linking modes. The results reveal substantial variations in correlation strength among different feature combinations and lost-linking modes, and the association strength increases significantly with the prolongation of overdue time. The results provide banks with quantitative early warning signs based on feature combinations, which can be applied to risk-grading monitoring systems. The research emphasizes the requirement for combined analysis of multidimensional features and dynamic monitoring in precise risk control.

Characterization of <i>Sr</i> UGT76G4 reveals a key residue for regioselectivity and efficient Reb M synthesis

Steviol glycosides (SGs) from Stevia rebaudiana are prized as noncaloric sweeteners, with rebaudioside M (Reb M)—a next-generation SG known for its sucrose-like sweetness and lack of off-tastes—standing out for its superior sensory profile. However, Reb M’s limited natural abundance impedes its commercial production. Here, we report the identification of a glucosyltransferase, UGT76G4 that efficiently catalyzes the conversion of Reb D to Reb M with a strong preference for C19 glycosylation. Structural and functional analyses, including X-ray crystallography, molecular dynamics simulations, and mutagenesis, revealed key residues in UGT76G4 that dictate its regioselectivity, with residue 200 playing a pivotal role. Engineered UGT76G4 variants, including Q199I/G200Y and H155S/Q199I/G200Y, enhanced Reb E and Reb D conversion efficiency by 1.46-fold and 23-fold, respectively, compared to UGT76G1. The engineered variants offer a promising pathway for increasing Reb M production, advancing biotechnological strategies for steviol glycoside biosynthesis and optimizing plant metabolic engineering approaches. Our findings deepen the understanding of SG biosynthesis and provide a basis for sustainable production of high-value sweeteners.

Time trends and persistence of the return difference between growth and value investment strategies

This paper examines the dynamic disequilibrium between value investing and growth strategies, focusing on the structural changes induced by the COVID-19 pandemic. Using fractional integration and Markov-switching dynamic regression (MS-DR) models, we analyze persistence and regime shifts. The results reveal that, prior to March 2020, the return difference was in a regime of high persistence and no reversion to the mean, making the deviations long-lasting. After the pandemic, the system shifted to a regime of moderate persistence with reversion to the mean, indicating that the return differences now tend to correct over time. This regime shift, confirmed by the Markov switching model, highlights a permanent change in the dynamics of value and growth strategies, which significantly affects their long-term equilibrium.

Correction for Leprevost et al., A widespread family of ribosomal peptide metallophores involved in bacterial adaptation to metal stress

Influence of environmental and anthropogenic factors on forest patch composition and structure in North Wollo Zone, Amhara region, Ethiopia

Ethiopia’s vegetation is increasingly threatened by anthropogenic activities and natural factors, leading to forest degradation and fragmentation. This study analyzed the composition and structure of woody species in the Gerado, Micha, and Mekelet forest patches in the North Wollo Zone, Ethiopia. Data were collected from 95 systematically laid plots (20 x 20 m²) along elevation gradients, sampling trees and shrubs with a diameter at breast height (DBH) ≥ 2.5 cm and height ≥ 2 m. Species diversity and richness were assessed using the Shannon-Wiener diversity index, while plant communities and their relationships with environmental variables were analyzed using hierarchical clustering and Redundancy Analysis (RDA) in R software. A total of 55 woody species, from 46 genera and 31 families, were recorded. Fabaceae was the most species-rich family (12 species). Four distinct plant community types were identified (R = 0.4703, p ≤ 0.001), with altitude, slope, and tree cutting significantly influencing community composition (p ≤ 0.05). The dominant species across all patches were Dodonaea viscosa subsp. angustifolia, Olea europaea L. subsp. cuspidata, and Vachellia sieberiana. Anthropogenic disturbances, such as tree cutting, firewood collection, charcoal production, and grazing, were key factors affecting vegetation structure. The findings highlight the need for community-based conservation strategies tailored to the unique ecological and socio-economic conditions of each forest patch to improve ecosystem resilience and sustainability.

Experimental evidence for the continuous transition between elastic and elastoinertial turbulence

Elastic turbulence (ET) and elastoinertial turbulence (EIT) of viscoelastic fluids are unique flow states with features distinct from the inertial turbulence of Newtonian fluids. Whether these two states are connected or entirely decoupled remains controversial. We here resolve this controversy by providing experimental evidence of a continuous transition between ET and EIT in Taylor-Couette flow. Through experimentally quantifying the roles of elasticity and inertia in flow stability, we find that elasticity is the primary driving mechanism for both elastic and elastoinertial instabilities, and inertia plays a secondary role in the latter. Remarkably, the critical condition for these instabilities can be described by a unified function derived from stability analysis, revealing that the transition between elastic instability to elastoinertial instability is continuous. Moreover, we show that the flow structures and the energy spectrum evolve seamlessly from ET to elasticity-dominated EIT, transitional EIT, and inertia-modulated EIT, with inertia playing an increasingly important role in the last three regimes. Our results offer insights into the fundamental nature of turbulence in viscoelastic flows and would have implications for applications involving drag reduction and polymer processing.