bluuebunny/crossref_metadata_2025 huggingface dataset:
Prediction of Hemolytic Peptides and their Hemolytic Concentration (HC50)
Authors: Rathore Anand Singh , Kumar Nishant , Choudhury Shubham , Mehta Naman Kumar , Raghava Gajendra P. S.
Published: July 2024

Several peptide-based drugs fail in clinical trials due to their toxicity or hemolytic activity against red blood cells (RBCs). Existing methods predict hemolytic peptides but not the concentration (HC50) required to lyse 50% of RBCs. In this study, we developed a classification model and regression model to identify and quantify the hemolytic activity of peptides. Our models were trained and validated on 1924 peptides with experimentally determined HC50 against mammalian RBCs. Analysis indicates that hydrophobic and positively charged residues were associated with higher hemolytic activity. Our classification models achieved a maximum AUC of 0.909 using a hybrid model of ESM-2 and a motif-based approach. Regression models using compositional features achieved R of 0.739 with R² of 0.543. Our models outperform existing methods and are implemented in the web-based platform HemoPI2 and standalone software for designing hemolytic peptides with desired HC50 values (http://webs.iiitd.edu.in/raghava/hemopi2/).

Highlights

Developed classification and regression models to predict hemolytic activity and HC50 values of peptides.

A hybrid model combining machine learning and motif prediction excels in accuracy.

Benchmarking of the existing classification methods on independent datasets.

Web server, standalone software, and pip package for hemolytic activity prediction of peptides/proteins.
DNA Methylation Age Acceleration Mediates the Relationship between Systemic Inflammation and Cognitive Impairment
Authors: Tejera César Higgins , Zhu Peiyao , Ware Erin B. , Hicken Margaret T. , Zawistowski Matthew , Kobayashi Lindsay C. , Seblova Dominika , Manly Jennifer , Mukherjee Bhramar , Bakulski Kelly M.
Published: July 2024

Background

Chronic inflammation and DNA methylation are potential mechanisms in dementia etiology. The linkage between inflammation and DNA methylation age acceleration in shaping dementia risk is understudied. We explored the association of inflammatory cytokines with cognitive impairment and whether DNA methylation age acceleration mediates this relationship.

Methods

In a subset of the 2016 wave of the Health and Retirement Study (n=3,346, age>50), we employed logistic regression to estimate the associations between each inflammatory cytokine (interleukin-6 (IL-6), C-reactive protein (CRP), and insulin-like growth factor-1 (IGF-1)), and both Langa-Weir classified cognitive impairment non-dementia and dementia, respectively. We calculated DNA methylation age acceleration residuals by regressing GrimAge on chronologic age. We tested if DNA methylation age acceleration mediated the relationship between systemic inflammation and cognitive impairment, adjusting for sociodemographic, behavioral factors, chronic conditions, and cell type proportions.

Results

The prevalence of cognitive impairment was 16%. In the fully-adjusted model, participants with a doubling of IL-6 levels had 1.12 (95% CI: 1.02-1.22) times higher odds of cognitive impairment. Similar associations were found for CRP and IGF-1. Participants with a doubling of IL-6 levels had 0.77 (95% CI: 0.64, 0.90) years of GrimAge acceleration. In mediation analyses with each cytokine as predictor separately, 17.7% (95% CI: 7.0%, 50.9%) of the effect of IL-6 on cognitive impairment was mediated through DNA methylation age acceleration. Comparable mediated estimates were found for CRP and IGF-1.

Conclusions

Systemic inflammation is associated with cognitive impairment, with suggestive evidence that this relationship is partially mediated through DNA methylation age acceleration.
Negatively charged α-Synuclein condensate modulates partitioning of molecules
Authors: Yang Qingqing , Chen Shunfa , Zhang Pengfei , Lu Zhonghua , Chang Shuwen , Wong Leo E.
Published: July 2024

α-Synuclein (αSyn) aggregation via liquid-liquid phase separation has recently emerged as a crucial mechanism underlying amyloid fibril formation implicated in Parkinson’s disease. However, comprehensive investigations of the physico-chemical properties of αSyn condensate remains incomplete. Here, we showed that αSyn condensate possesses a highly negative electrostatic potential that spans the whole condensate. This property causes differential partitioning of dye-labeled αSyn as well as fluorescent molecules by an order of magnitude depending on their net charges. Consistent with this, the phase separation propensity of αSyn is governed by a delicate balance between self-association of αSyn and electrostatic repulsion, hence is antagonized by excess negative charge. We further demonstrated that, in differentiated neuron-like SH-SY5Y cells, αSyn also forms negatively charged condensate. Our results highlighted the significant impact of αSyn condensate’s electrostatic potential on molecular partitioning, hence calling for close examination of the electrostatic property of other biomolecular condensates.
Stress relaxation rates of myocardium from failing and non-failing hearts
Authors: Gionet-Gonzales Marissa , Gathman Gianna , Rosas Jonah , Kunisaki Kyle Y. , Inocencio Dominique Gabriele P. , Hakami Niki , Milburn Gregory N. , Pitenis Angela A. , Campbell Kenneth S. , Pruitt Beth L. , Stowers Ryan S.
Published: July 2024

The heart is a dynamic pump whose function is influenced by its mechanical properties. The viscoelastic properties of the heart, i.e. its ability to exhibit both elastic and viscous characteristics upon deformation, influence cardiac function. Viscoelastic properties change during heart failure (HF), but, direct measurements of failing and non-failing myocardial tissue stress relaxation under constant displacement are lacking. Further, how consequences of tissue remodeling, such as fibrosis and fat accumulation, alter the stress relaxation remains unknown. To address this gap, we conducted stress relaxation tests on porcine myocardial tissue to establish baseline properties of cardiac tissue. We found porcine myocardial tissue to be fast relaxing, characterized by stress relaxation tests on both a rheometer and microindenter. We then measured human left ventricle (LV) epicardium and endocardium human tissue from non-failing, ischemic HF, and non-ischemic HF patients by microindentation. We found that the ischemic HF had slower stress relaxation than non-failing endocardium; and that slower stress relaxing tissues were correlated with increased collagen deposition and increased α-smooth muscle actin (α-SMA) stress fibers, a marker of fibrosis and cardiac fibroblast activation, respectively. In the epicardium, we found that ischemic HF had faster stress relaxation than non-ischemic HF and non-failing; and that faster stress relaxation correlated with Oil Red O staining, a marker for adipose tissue. These data show that changes in stress relaxation vary across the different layers of the heart during ischemic vs. non-ischemic HF. These findings reveal how the viscoelasticity of the heart changes, which will lead to better modeling of cardiac mechanics for in vitro and in silico HF models.
Enhancing the Reverse Transcriptase Function in Taq Polymerase via AI-driven Multiparametric Rational Design
Authors: Tomilova Yulia E. , Russkikh Nikolay E. , Yi Igor M. , Shaburova Elizaveta V. , Tomilov Viktor N. , Pyrinova Galina B. , Brezhneva Svetlana O. , Tikhonyuk Olga S. , Gololobova Nadezhda S. , Popichenko Dmitriy V. , Arkhipov Maxim O. , Bryzgalov Leonid O. , Brenner Evgeny V. , Artyukh Anastasia A. , Shtokalo Dmitry N. , Antonets Denis V. , Ivanov Mikhail K.
Published: July 2024

Modification of natural enzymes to introduce new properties and enhance existing ones is a central challenge in bioengineering. This study is focused on the development of Taq polymerase mutants that show enhanced reverse transcriptase (RTase) activity while retaining other desirable properties such as fidelity, 5′-3′ exonuclease activity, effective deoxyuracil incorporation, and tolerance to locked nucleic acid (LNA)-containing substrates. Our objective was to use AI-driven rational design combined with multiparametric wet-lab analysis to identify and validate Taq polymerase mutants with an optimal combination of these properties. The experimental procedure was conducted in several stages: 1) On the basis of a foundational paper, we selected 18 candidate mutations known to affect RTase activity across six sites. These candidates, along with the wild type, were assessed in the wet lab for multiple properties to establish an initial training dataset. 2) A ridge regression model was trained on this dataset to predict the enzymes’ properties. This model enabled us to select 14 new candidates for further experimental testing. 3) We refined our predictive model using Gaussian process regression and trained it on an expanded dataset now including 33 data points. 4) Leveraging the refined model, we screenedin silicoover 27 million potential mutations, thus selecting 16 for detailed wet-lab evaluation. Through this iterative data-driven approach, we identified 18 enzymes that not only manifested considerably enhanced RTase activity but also retained a balance of other required properties. These enhancements were generally accompanied by lower Kd, moderately reduced fidelity, and greater tolerance to noncanonical substrates, thereby illustrating a strong interdependence among these traits. Several enzymes validated via this procedure were effective in single-enzyme real-time reverse-transcription PCR setups, implying their utility for the development of new tools for real-time reverse-transcription PCR technologies, such as pathogen RNA detection and gene expression analysis. This study illustrates how AI can be effectively integrated with experimental bioengineering to enhance enzyme functionality systematically. Our approach offers a robust framework for designing enzyme mutants tailored to specific biotechnological applications. The results of our biological activity predictions for mutated Taq polymerases can be accessed athttps://huggingface.co/datasets/nerusskikh/taqpol_insilico_dms.
Rice JASMONIC ACID OXIDASES (OsJAO) control resting jasmonate metabolism to promote development and repress basal immune responses
Authors: Ndecky Simon , Malherbe Ludivine , Villette Claire , Chalvon Véronique , Meusnier Isabelle , Beltran-Valencia Dennisse , Baumberger Nicolas , Riemann Michael , Kroj Thomas , Champion Antony , Heitz Thierry
Published: July 2024

Recent research has established that catabolic conversions within the jasmonate pathway have significant consequences on hormone signaling output. In dicotyledonous plants, the jasmonic acid oxidase (JAO) catabolic route is endowed with a regulatory function by diverting jasmonic acid (JA) towards hydroxylation, at the expense of its conjugation into the bioactive jasmonoyl-isoleucine (JA-Ile) hormone. Here we functionally characterized the JAO pathway in rice (Oryza sativa) and demonstrate its prevalent function in promoting growth and attenuating JA responses in vegetative tissues. The rice genome contains four JAO-related homologs of which three generated hydroxy-JAin vitroand reverted the high defense phenotype when expressed in the Arabidopsisjao2-2mutant. By generating and analyzing a series of single to quadruple ricejaomutants, we show the incremental effect of gradual JAO depletion on JA metabolism, basal defense levels, growth inhibition, fitness and global metabolic reprogramming. JAO-deficient lines were significantly growth-retarded at the juvenile stage, while recovering a near wild-type vegetative development after three months, where they exhibited a enhanced resistance to virulent and avirulent strains ofMagnaporthe oryzae, the causal agent of fungal blast disease. Our findings identify the JAO pathway as an integral component of rice JA homeostasis and an important determinant of the growth-defense tradeoff. They demonstrate its conserved regulatory function in monocots and open possibilities for modulating selectively basal JA responses in a major cereal crop. Natural variation in JAO activity could also be explored as a mechanism underlying varying levels of JA signaling output in rice.
Differential responses in the core, active site and peripheral regions of cytochrome c peroxidase to extreme pressure and temperature
Authors: Zawistowski Rebecca K. , Crane Brian R.
Published: July 2024

In consideration of life in extreme environments, the effects of hydrostatic pressure on proteins at the atomic level have drawn substantial interest. Large deviations of temperature and pressure from ambient conditions can shift the free energy landscape of proteins to reveal otherwise lowly populated structural states and even promote unfolding. We report the crystal structure of the heme-containing peroxidase, cytochrome c peroxidase (CcP) at 1.5 and 3.0 kbar and make comparisons to structures determined at 1.0 bar and cryo-temperatures (100 K). Compressibility plateaus after 1.5 kbar and pressure produces anisotropic changes in CcP. CcP responds to pressure with volume declines at the periphery of the protein where B-factors are relatively high but maintains nearly intransient core structure and active site channels. Compression at the surface affects neither alternate side-chain conformers nor B-factors. Thus, packing in the core, which resembles a crystalline solid, limits motion and protects the active site, whereas looser packing at the surface preserves side-chain dynamics. Changes in active-site solvation and heme ligation reveal pressure sensitivity to protein-ligand interactions and reveal a potential docking site for the substrate peroxide.
Adaptation to water salinity changes: phenotypic plasticity meets and moulds carry-over effects in sea rock-pool mosquitoes
Authors: Cordeschi Giulia , Bisconti Roberta , Mastrantonio Valentina , Canestrelli Daniele , Porretta Daniele
Published: July 2024

Organisms living in highly variable environments are expected to exhibit a strong phenotypic variability within populations due to plastic responses to environmental fluctuations. Yet, this phenomenon is not commonly seen in nature, since compensatory mechanisms can promote phenotypic stability. Here, we hypothesized a potential compensatory effect that could result from the interaction between behavioural and morphological trait plasticity. These traits are highly responsive to environmental factors, and compelling research has demonstrated that behavioural plasticity can impact individual life history traits. In microcosm experiments, we analysed plasticity in body size, larval and pupal behaviour and carry-over effect across developmental stages in the sea rock-pool mosquitoAedes mariaein response to water salinity changes. Analyzing morphological traits, we found that larvae raised under increasing salinity were smaller than those raised under constant conditions. Additionally, a smaller body size at the pupal stage was observed, which indicated the presence of carry-over effects from the larval to pupal stages. However, no differences were observed in adult sizes between the two salinity conditions. When examining behavioural traits, we observed that higher salinity promoted plastic changes in larval activity behaviour and pupal diving behaviour. In particular, pupae exposed to increased salinity spent 20.6% less time underwater and performed fewer abdominal contractions than those under constant conditions. Since pupal energy expenditure is proportional to the time spent underwater, we suggest that the plastic pupal behaviour promoted compensatory energy allocation to growth, thereby reducing the carry-over effects from the pupal to the adult stage. This study emphasizes that plasticity at multiple traits during development can impact carry-over effects, ultimately leading to the convergence of adult phenotypes.
A family of bacterial Josephin-like deubiquitinases with an unusual cleavage mode
Authors: Hermanns Thomas , Kolek Susanne , Uthoff Matthias , de Heiden Richard A. , Mulder Monique P.C. , Baumann Ulrich , Hofmann Kay
Published: July 2024

Many intracellular bacteria secrete deubiquitinase (DUB) effectors into eukaryotic host cells to keep the bacterial surface or the enclosing vesicle membrane free of ubiquitin marks. Here, we describe a new family of bacterial DUBs that is structurally related to eukaryotic Josephins, but contains members that catalyze a unique destructive substrate deubiquitination. These ubiquitin C-terminal clippases (UCCs) cleave ubiquitin before the C-terminal diGly motif, thereby truncating the modifier and leaving a remnant on the substrate. By comparing the crystal structures of substrate-bound clippases and a closely related conventional DUB, we identified the factors causing the shift and found them conserved in other clippases, including one highly specific for M1-linked ubiquitin chains. This new enzyme class has great potential as tools to study the ubiquitin system, in particular aspects involving branched chains.
Satellite glial contact enhances differentiation and maturation of human iPSC-derived sensory neurons
Authors: LeBlang Chelsey J. , Pazyra-Murphy Maria F. , Silagi Elizabeth S. , Dasgupta Srestha , Tsolias Marianna , Miller Toussaint , Petrova Veselina , Zhen Shannon , Jovanovic Vukasin , Castellano David , Gerrish Kathryn , Ormanoglu Pinar , Tristan Carlos , Singeç Ilyas , Woolf Clifford J. , Tasdemir-Yilmaz Ozge , Segal Rosalind A.
Published: July 2024

Sensory neurons generated from induced pluripotent stem cells (iSNs) are used to model human peripheral neuropathies, however current differentiation protocols produce sensory neurons with an embryonic phenotype. Peripheral glial cells contact sensory neurons early in development and contribute to formation of the canonical pseudounipolar morphology, but these signals are not encompassed in current iSN differentiation protocols. Here, we show that terminal differentiation of iSNs in co-culture with rodent Dorsal Root Ganglion satellite glia (rSG) advances their differentiation and maturation. Co-cultured iSNs develop a pseudounipolar morphology through contact with rSGs. This transition depends on semaphorin-plexin guidance cues and on glial gap junction signaling. In addition to morphological changes, iSNs terminally differentiated in co-culture exhibit enhanced spontaneous action potential firing, more mature gene expression, and increased susceptibility to paclitaxel induced axonal degeneration. Thus, iSNs differentiated in coculture with rSGs provide a better model for investigating human peripheral neuropathies.
Reliable inference of phylogenomic relationship via assembly-based strategy accommodating raw reads and proteins
Authors: Li Yunlong , Liu Xu , Chen Chong , Qiu Jian-Wen , Kocot Kevin , Sun Jin
Published: July 2024

Phylogenomics has emerged as a transformative approach in systematics, conservation biology, and biomedicine, enabling the inference of evolutionary relationships by leveraging hundreds to thousands of genes from genomic or transcriptomic data. However, acquiring high-quality genomes and transcriptomes necessitates samples with intact DNA and RNA, substantial sequencing investments, and extensive bioinformatic processing, such as genome/transcriptome assembly and annotation. This challenge is particularly pronounced for rare or difficult-to-collect species, such as those inhabiting the deep sea, where only fragmented DNA reads are often available due to environmental degradation or suboptimal preservation conditions. To address these limitations, we introduce VEHoP (Versatile, Easy-to-use Homology-based Phylogenomic pipeline), a tool designed to infer protein-coding regions from diverse inputs, including raw reads (short and long), draft genomes, transcriptomes, and annotated genomes. VEHoP automates the generation of orthologous sequence alignments, concatenated matrices, and phylogenetic trees, streamlining phylogenomic analyses for researchers across disciplines. The tool aims to (1) expand taxonomic sampling by accommodating a wide range of input data types and (2) simplify phylogenomic workflows, making them accessible to researchers with varying levels of bioinformatic expertise. We evaluated VEHoP’s performance using datasets from oysters, catfish, and insects, demonstrating its ability to produce robust phylogenetic trees with strong bootstrap support, outperforming assembly-free methods. Additionally, we applied VEHoP to reconstruct the phylogeny of the enigmatic deep-sea gastropod order Neomphalida, successfully resolving a well-supported phylogenetic backbone for this poorly understood group. VEHoP is freely available on GitHub (https://github.com/ylify/VEHoP), with dependencies easily installable via Bioconda.
Structural diversity and distribution of NMCP-class nuclear lamina proteins in streptophytic algae
Authors: Kosztyo Brendan S. , Richards Eric J.
Published: July 2024

Nuclear Matrix Constituent Proteins (NMCPs) in plants function like animal lamins, providing the structural foundation of the nuclear lamina and regulating nuclear organization and morphology. Although they are well-characterized in angiosperms, the presence and structure of NMCPs in more distantly related species, such as streptophytic algae, are relatively unknown. The rapid evolution of NMCPs throughout the plant lineage has caused a divergence in protein sequence that makes similarity-based searches less effective. Structural features are more likely to be conserved compared to primary amino acid sequence; therefore, we developed a filtration protocol to search for diverged NMCPs based on four physical characteristics: intrinsically disordered content, isoelectric point, number of amino acids, and the presence of a central coiled-coil domain. By setting parameters to recognize the properties of bona fide NMCP proteins in angiosperms, we filtered eight complete proteomes from streptophytic algae species and identified strong NMCP candidates in six taxa in the Classes Zygnematophyceae, Charophyceae, and Klebsormidophyceae. Through analysis of these proteins, we observed structural variance in domain size between NMCPs in algae and land plants, as well as a single block of amino acid conservation. Our analysis indicates that NMCPs are absent in the Mesostigmatophyceae. The presence versus absence of NMCP proteins does not correlate with the distribution of different forms of mitosis (e.g., closed/semi-closed/open) but does correspond to the transition from unicellularity to multicellularity in the streptophytic algae, suggesting that an NMCP-based nucleoskeleton plays important roles in supporting cell-to-cell interactions.

SIGNIFICANCE STATEMENT

All eukaryotic organisms contain a membrane-bound nucleus, which holds a cell’s DNA, but plants and animals have distinct sets of proteins that make up the mesh-like scaffold inside the nucleus (i.e., nuclear lamina) that is essential for structural integrity and organization. Nonetheless, the major nuclear lamina proteins in plants and animals share structural features, and we exploited these characteristics to identify a key class of nuclear lamina proteins in streptophytic algae, allowing us to chart the distribution of these proteins across photosynthetic organisms and to gain insight into the evolution of nuclear organization. Our results indicate that the major class of nuclear lamina protein in plants evolved independently from structurally similar nuclear lamina proteins in animals.
A novel method for integrating genomic and Tn-Seq data to identify commonin vivofitness mechanisms across multiple bacterial species
Authors: Fouts Derrick E. , Clarke Thomas H. , Brown Aric N. , Ottosen Elizabeth N. , Holmes Caitlyn L. , Moricz Bridget S. , Mason Sophia , Sinha Ritam , Severin Geoffrey B. , Anderson Mark T. , DiRita Victor , Bachman Michael A. , Mobley Harry L. T.
Published: July 2024

Sepsis is life-threatening organ dysfunction due to an unregulated immune response to infection. Bacteremia is a leading cause of sepsis, and members of theEnterobacteralescause nearly half of bacteremia cases annually. While previous Tn-Seq studies to identify novel bacteremia-fitness genes have provided valuable insight into virulence mechanisms, evidence for common pathways across species was lacking. To identify common fitness pathways in five bacteremia-causedEnterobacteralesspecies, we utilized the JCVI pan-genome pipeline to integrate Tn-Seq fitness data with multiple available functional data types. Core genes from species pan-genomes were used to construct a multi-species core pan-genome, producing 2,850 core gene clusters found in four out of the five species. Integration of Tn-Seq fitness data enabled identification of 373 protein clusters that were conserved in all five species. A scoring rubric was applied to these clusters, which incorporated Tn-Seq fitness defects, operon localization, and antibiotic susceptibility data to identify seven common bacteremia-fitness pathways. Mutations intatCshowed reduced fitnessin vivoand increased susceptibility to beta-lactams that were restored followingtatCcomplementationin trans. By integrating known operon structures and antibiotic susceptibility with Tn-Seq fitness data, common genes within the core pan-genome emerged and revealed mechanisms that are essential for colonization of, or survival in, the mammalian bloodstream. Our prediction and validation oftatCas a common bacteremia fitness factor and contributor of antibiotic resistance supports the utility of this bioinformatic approach. This study represents a major step forward to identify novel targets of therapy against these deadly widespread sepsis infections.
Flooding patterns shape microbial community in mangrove sediments
Authors: Vázquez-Rosas-Landa Mirna , Pérez-Ceballos Rosela , Zaldívar-Jiménez Arturo , Hereira-Pacheco Stephanie E , Pérez-González Leonardo D. , Prieto‑Davó Alejandra , Celis-Hernández Omar , Canales-Delgadillo Julio C.
Published: July 2024

Background

Mangrove ecosystems located in the tropics and subtropics, are crucial for regulating global weather patterns and sequestering carbon. However, they face threats from human activities like altered water flow and deforestation. While the symbiotic relationship between mangrove trees and surrounding microbes are essential for their survival, the impact of human activity on these microbial communities remains incompletely understood. We investigated how microbial communities change in degraded mangrove ecosystems due to loss of hydrologic connectivity, aiming to elucidate consequences and inform restoration strategies.

Methods

Employing 16S rRNA sequencing, we analyzed samples of sediment cores from conserved, moderately degraded, and degraded mangrove sites across dry and flood seasons at three sediment depths.

Results

Our analysis identified 11,469 Amplicon Single Variant (ASVs), revealing diversity loss correlated with degradation levels. Notably, we observed shifts in microbial diversity within sediment layers, with conserved sites dominated by Vibrionaceae in upper layers, potentially indicating urban contamination. In moderate-degradation sites, seasonal patterns emerged, with Halomonas and Marinomonas dominating the dry season and Exiguobacterium thriving during flooding. Interestingly, a community mainly composed of Firmicutes persisted across all degradation scenarios in deeper sediment layers, suggesting potential for ecosystem restoration. Our findings provide insights into microbial responses to human-induced stressors and highlight the role of core microbial communities in guiding restoration efforts for degraded mangrove ecosystems.
Illuminating microbial mat assembly: Cyanobacteria and Chloroflexota cooperate to structure light-responsive biofilms
Authors: Bunbury Freddy , Rivas Carlos , Calatrava Victoria , Malkovskiy Andrey , Joubert Lydia-Marie , Parvate Amar D. , Evans James E. , Grossman Arthur , Bhaya Devaki
Published: July 2024

Microbial mats are stratified communities often dominated by unicellular and filamentous phototrophs within an exopolymer matrix. It is challenging to quantify the dynamic responses of community membersin situas they experience steep gradients and rapid fluctuations of light. To address this, we developed a binary consortium using two representative isolates from hot spring mats: the unicellular oxygenic phototrophic cyanobacteriumSynechococcusOS-B’ (Syn OS-B’) and the filamentous anoxygenic phototrophChloroflexusMS-CIW-1 (Chfl MS-1). We quantified the motility of individual cells and entire colonies and demonstrated that Chfl MS-1 formed bundles of filaments that moved in all directions with no directional bias to light. Syn OS- B’ was slightly less motile but exhibited positive phototaxis. This binary consortium displayed cooperative behavior by moving further than either species alone and formed ordered arrays where both species aligned with the light source. No cooperative motility was observed when a non-motilepilBmutant of Syn OS-B’ was used instead of Syn OS-B’. The binary consortium also produced more adherent biofilm than individual species, consistent with the close interspecies association revealed by electron microscopy. We propose that cyanobacteria and Chloroflexota cooperate in forming natural microbial mats, by colonizing new niches and building robust biofilms.

Significance

Microbial mats are dense, layered communities with ancient origins and widespread occurrence, but how they assemble is not well understood. To investigate how microbial motility, physical interactions, and responses to light affect mat assembly, we developed a binary consortium from representative hot spring mat isolates. Individually, the Cyanobacteria and Chloroflexota isolates displayed significant differences in motility and biofilm formation. When combined, the consortium exhibited enhanced motility towards light and formed more robust biofilms. This model consortium approach complementsin situstudies by directly testing the role of motility and physical cooperation in shaping microbial mats, and could inform biofilm applications in industrial settings.
Leaky ribosomal scanning enables tunable translation of bicistronic ORFs in green algae
Authors: Duenas Marco A. , Craig Rory J. , Gallaher Sean D. , Moseley Jeffrey L. , Merchant Sabeeha S.
Published: July 2024

Advances in sequencing technology have unveiled examples of nucleus-encoded polycistronic genes, once considered rare. Exclusively polycistronic transcripts are prevalent in green algae, although the mechanism by which multiple polypeptides are translated from a single transcript is unknown. Here, we used bioinformatic and in vivo mutational analyses to evaluate competing mechanistic models for polycistronic expression in green algae. High-confidence manually curated datasets of bicistronic loci from two divergent green algae,Chlamydomonas reinhardtiiandAuxenochlorella protothecoides, revealed 1) a preference for weak Kozak-like sequences for ORF 1 and 2) an underrepresentation of potential initiation codons before ORF 2, which are suitable conditions for leaky scanning to allow ORF 2 translation. We used mutational analysis inAuxenochlorella protothecoidesto test the mechanism. In vivo manipulation of the ORF 1 Kozak-like sequence and start codon altered reporter expression at ORF 2, with a weaker Kozak-like sequence enhancing expression and a stronger one diminishing it. A synthetic bicistronic dual reporter demonstrated inversely adjustable activity of green fluorescent protein expressed from ORF 1 and luciferase from ORF 2, depending on the strength of the ORF 1 Kozak-like sequence. Our findings demonstrate that translation of multiple ORFs in green algal bicistronic transcripts is consistent with episodic leaky ribosome scanning of ORF 1 to allow translation at ORF 2. This work has implications for the potential functionality of upstream open reading frames found across eukaryotic genomes and for transgene expression in synthetic biology applications.

Significance Statement

Textbook dogma states that nucleus-encoded genes are monocistronic, producing transcripts with a single translated open reading frame. However, highly conserved bicistronic loci are pervasive in green algae that are separated by several hundred million years of evolution, speaking to their ancestral origins and functions within the Chlorophyte lineage. A combination of bioinformatic analysis and in vivo gene manipulation supports leaky ribosomal scanning as the primary mechanism for translation of multiple ORFs from bicistronic transcripts. We have successfully tuned synthesis levels of two proteins encoded on one mRNA by modifying the ORF 1 Kozak-like sequence. These findings may have broad applications in synthetic biology.
Cortico-subcortical dynamics in primate working memory
Authors: von Nicolai Constantin , Siegel Markus
Published: July 2024

Working memory is essential for cognition, facilitating the temporary maintenance and manipulation of information to produce goal-directed behavior. While both cortical and subcortical structures are involved, their precise roles and interactions are not fully understood. To investigate this, we simultaneously recorded neural activity from the frontal and parietal cortex, higher-order thalamic nuclei, and core basal ganglia structures during color and spatial working memory tasks in non-human primates. We found widespread yet differential encoding of color and spatial information, marked by area-specific temporal dynamics and modulation according to task demands. Both cortical and subcortical information increased towards working memory-dependent actions, suggesting a task-specific reloading of information. Directed interactions between cortical and subcortical regions were extensive and reciprocal, with dominant directions of information flow, especially from frontoparietal areas. These interactions were dynamically modulated and partially task specific. Our findings provide comprehensive insights into the large-scale circuit dynamics underlying primate working memory and suggest that flexible goal-directed behavior relies on the selective processing of task-relevant memory information within information-specific cortico-subcortical networks.
Does early-life food shortage alter the effect of elevated temperature on female life history?
Authors: Chung Meng-Han Joseph , Zang Chenke , Moura-Campos Diego , Jennions Michael D. , Head Megan L.
Published: July 2024

Global warming is reducing prey availability in many aquatic systems, raising questions about the combined effects of higher temperatures and lower food availability on fish life histories and reproductive output.

In ectotherms, higher temperatures accelerate growth and promote an earlier onset of reproduction. However, when fish have less food during development, resource depletion might constrain these temperature-driven processes.

We manipulated water temperature (24 or 28°C) and early-life food availability (control or restricted) for female guppies (Poecilia reticulata). We measured how both factors affected key life history traits (growth, reproduction, survival, self-maintenance).

Higher temperature significantly affected female life histories. Females at 28°C matured at a larger size, but then grew more slowly and produced fewer, smaller offspring than females at 24°C. The effect of temperature on reproduction persisted even after controlling for body size, suggesting there was a shift in the fecundity-size relationship.

Adult mortality was greater at 28°C. Higher temperature also resulted in a longer gut, potentially enhancing resource acquisition, but a higher temperature did not affect immunity or telomere length of the surviving females.

Early-life food shortage affected very few traits, except for a weak interaction with temperature that affected total fecundity. At 28°C, females that experienced early-life food restriction produced fewer offspring than females with continual food supply. No such diet effect occurred at 24°C.

Our results suggest that tropical fish may be severely impacted by increased temperatures (i.e., decreased reproduction with increased morality), but are likely to be resilient to brief periods of food limitations during early development.

Interestingly, early-life food shortage caused a reduction in total offspring number but only at 28°C, suggesting that global prey decline might exacerbate the negative effects of a warming climate on stock recruitment of tropical fish.
Self-Assembled Chambered Cardiac Organoids for Modeling Cardiac Chamber Formation and Cardiotoxicity Assessment
Authors: Zou Xinle , Wang Fanwen , Zheng Huilin , Kong Tianci , Pei Duanqing
Published: July 2024

The human heart, crucial for health and longevity, is a primary focus of medical research. Despite advancements in cardiac organoid technology, replicating early cardiac chamber formation stages remains challenging. Here, we develop chambered cardiac organoids (CCOs) by orchestrating cardiac signaling pathways, including the synergistic modulation of FGF and Wnt signaling via the HAND1 transcription factor. These CCOs exhibit stable chambers with self-organized outer myocardial and inner endocardial layers, and express developmental markers such as NKX2.5, TNNT2, and NAPPA, demonstrating physiological functionality including spontaneous contractions and calcium transients. Immunofluorescence and single-cell RNA sequencing confirmed the presence and stability of cardiomyocytes within CCOs. Further validation showed CCOs’ involvement in critical processes like endothelial-mesenchymal transition (EndoMT) and valvulogenesis. Ultrastructural and electrophysiological analysis revealed organized myofibrils and atrial-like action potentials. Importantly, CCOs proved effective in assessing cardiotoxicity through observable morphological changes, demonstrating specific cellular responses to established cardiotoxic compounds, thereby highlighting their potential for drug testing and safety evaluations. Our findings offer insights into cardiac chamber formation mechanisms and establish a robust platform for drug testing, disease modeling, and personalized medicine, representing a significant advancement in functional cardiac organoid development for both basic research and translational applications.
How to reward animals based on their subjective percepts: A Bayesian approach to online estimation of perceptual biases
Authors: Dong Yelin , Lengyel Gabor , Shivkumar Sabyasachi , Anzai Akiyuki , DiRisio Grace F. , Haefner Ralf M. , DeAngelis Gregory C.
Published: July 2024

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Elucidating the neural basis of perceptual biases, such as those produced by visual illusions, can provide powerful insights into the neural mechanisms of perceptual inference. However, studying the subjective percepts of animals poses a fundamental challenge: unlike human participants, animals cannot be verbally instructed to report what they see, hear, or feel. Instead, they must be trained to perform a task for reward, and researchers must infer from their responses what the animal perceived. However, animals’ responses are shaped by reward feedback, thus raising the major concern that the reward regimen may alter the animal’s decision strategy or even intrinsic perceptual biases. We developed a method that estimates perceptual bias during task performance and then computes the reward for each trial based on the evolving estimate of the animal’s perceptual bias. Our approach makes use of multiple stimulus contexts to dissociate perceptual biases from decision-related biases. Starting with an informative prior, our Bayesian method updates a posterior over the perceptual bias after each trial. The prior can be specified based on data from past sessions, thus reducing the variability of the online estimates and allowing it to converge to a stable estimate over a small number of trials. After validating our method on synthetic data, we apply it to estimate perceptual biases of monkeys in a motion direction discrimination task in which varying background optic flow induces robust perceptual biases. This method overcomes an important challenge to understanding the neural basis of subjective percepts.
Co-recognition of histone acetylation and H3K4 trimethylation by GTE4-EML complex in Arabidopsis
Authors: Qian Feng , Zhao Qiang-Qiang , Zhou Jin-Xing , Yuan Dan-Yang , Su Yin-Na , Li Lin , Chen She , He Xin-Jian
Published: July 2024

Although histone acetylation and H3K4 trimethylation (H3K4me3) are well-known histone marks associated with active transcription, how they cooperate to regulate transcription remains largely unclear in plants. Our study revealed that the Bromodomain and Extra-terminal (BET) protein GTE4 binds to acetylated histone and forms a complex with the redundant H3K4me3-binding EMSY-Likeproteins EML1 or EML2 (EML1/2) inArabidopsis thaliana. Theeml1 eml2(eml1/2) double mutant exhibited a morphological phenotype similar to thegte4mutant, and most of thegte4-mediated differentially expressed genes were co-regulated in theeml1/2mutant. Through chromatin immunoprecipitation followed by deep sequencing (ChIP-seq), we found that GTE4 and EML2 co-occupy protein-coding genes enriched with both histone acetylation and H3K4me3, exhibiting a synergistic effect on the association of the GTE4-EML complex with chromatin. The association of GTE4 with chromatin requires both the Bromodomain and the EML-interacting domain. This study identified a previously uncharacterized complex and uncovered how it cooperatively recognizes histone acetylation and H3K4me3 to facilitate gene transcription at the whole-genome level in Arabidopsis.
Characterization of radiations-induced genomic structural variations inArabidopsis thaliana
Authors: Sall Salimata Ousmane , Alioua Abdelmalek , Staerck Sébastien , Graindorge Stéfanie , Pellicioli Michel , Schuler Jacky , Raffy Quentin , Rousseau Marc , Molinier Jean
Published: July 2024

DNA, is assaulted by endogenous and exogenous agents that lead to the formation of damage. In order to maintain genome integrity DNA repair pathways must be efficiently activated to prevent mutations and deleterious chromosomal rearrangements. Conversely, genome flexibility is also necessary to allow genetic diversity and evolution. The antagonist interaction between maintenance of genome integrity and flexibility determines genome shape and organization. Therefore, it is of great interest to understand how the whole linear genome structure behaves upon formation and repair of DNA damage. For this, we used long reads sequencing technology to identify and to characterize genomic structural variations (SV) of wild-typeArabidopsis thalianasomatic cells exposed either to UV-B, to UV-C or to protons irradiations. We found that genomic regions located in heterochromatin a more prone to form SVs than those located in euchromatin, highlighting that genome stability and flexibility differs along the chromosome. This holds true in Arabidopsis plants deficient for the expression of master regulators of the DNA Damage Response (DDR), ATM and ATR, suggesting that independent and alternative surveillance processes exist to maintain integrity in genic regions. Finally, the analysis of the radiations-induced deleted regions allowed determining that exposure UV-B, UV-C and protons induced the Microhomology-mediated end joining mechanism (MMEJ) and that both ATM and ATR repress this repair pathway.
The deubiquitinating enzyme CYLD impairs NF-κB- and STAT1-dependent macrophage intrinsic immunity toStaphylococcus aureus
Authors: Schmidt Christina , Harit Kunjan , Traidl Stephan , Werfel Thomas , Rösner Lennart , Nishanth Gopala , Schlüter Dirk
Published: July 2024

In atopic dermatitis (AD), lesional skin is frequently colonized byStaphylococcus(S.)aureus, contributing to the severity and clinical symptoms of the disease. The inflammatory milieu in the skin is characterized by a type 2 inflammatory signature, including M2 macrophages, which cannot eradicate S. aureus. Since S. aureus is effectively controlled in macrophages activated by pattern recognition receptor (PRR)-induced NF-κB and interferon (IFN)-γ-induced STAT1 stimulation, we hypothesized that pre-treatment with LPS as a PRR/TLR4-activating agent and IFN-γ would induce effective control of S. aureus in monocyte-derived macrophages (MDMs) of AD patients. Our data show that the deubiquitinating enzyme CYLD is strongly expressed in skin macrophages and MDMs from AD patients compared to healthy controls and impairs the anti-staphylococcal activity of PRR-activated and IFN-treated MDMs. Functionally, CYLD impaired M1 macrophage polarization, as evidenced by reduced expression of CD80, TNF, and IL-6 upon LPS- and IFN-γ treatment in CYLD-deficient as compared to wild-type (WT) MDM/THP1 macrophages. Mechanistically, CYLD inhibited IFN-γ-induced STAT1 phosphorylation and activation by binding to STAT1 and inducing its K63 deubiquitination. CYLD also interacted with TRAF6 and NEMO/IKKγ in the MYD88 signaling pathway and with RIPK2 in the NOD2 pathway, leading to impaired activation of NF-κB. Inhibition of both STAT1 by siRNA and NF-κB by IKK inhibitor treatment, respectively, independently abolished the control ofS. aureusin both CYLD-deficient and WT THP1 macrophages, which harbored identical high numbers of the pathogen. The in vivo inhibitory function of CYLD on the control ofS. aureuswas also observed upon infection ofCyld-deficient and WT mice. Collectively, these data illustrate that the increased expression of CYLD in macrophages of AD patients is a factor contributing to the ineffective control ofS. aureus, diminishes M1 macrophage polarization, and that CYLD deletion unleashes the break on effective STAT1 and NF-κB-dependent control ofS. aureus.
Single-molecule detection of transient dimerization of opioid receptors 2: Heterodimer blockage reduces morphine tolerance
Authors: Zhou Peng , Kasai Rinshi S. , Fujita Wakako , Tsunoyama Taka A. , Ueda Hiroshi , Pigolotti Simone , Fujiwara Takahiro K. , Kusumi Akihiro
Published: July 2024

Heterodimerization of opioid receptors (ORs), MOR, KOR, and DOR, is implied in their functional regulation and diversification, and thus its understanding is crucial for developing better analgesic treatments. However, our knowledge on OR heterodimerization/heterodimers remains limited. Here, using single-molecule imaging and functional analysis, we found that MOR, the main morphine receptor, repeatedly formstransient (≈250 ms) heterodimerswith DOR every 1-10 seconds, but not with KOR, whereas DOR and KOR also form transient heterodimers. We obtained all the heterodimer-monomer equilibrium constants and rate constants with/without agonists. We identified the critical heterodimer binding sites in the extracellular domains, in addition to the less-specific transmembrane domains, and developed soluble peptide blockers for MOR-DOR and DOR-KOR heterodimerization, using amino-acid sequences mimicking the extracellular binding sites. With these peptide blockers, we dissected the monomer/dimer roles in OR internalization and signaling. The soluble MOR-DOR heterodimer blocker reduced the development of long-term morphine tolerance in mice.