bluuebunny/crossref_metadata_2025 huggingface dataset:
Revealing a Correlation between Structure andin vitroactivity of mRNA Lipid Nanoparticles
Authors: Chen Xiaoxia , Li Mengrong , Jiang Fan , Hong Liang , Liu Zhuo
Published: September 2024

Messenger RNA-containing lipid nanoparticles (mRNA-LNPs) represent a promising platform for disease prevention, cancer immunotherapy, and gene editing, etc. Despite the clinical success of mRNA-LNPs based vaccines in preventing infectious diseases, the relationship between their structure and efficacy remains poorly understood, primarily due to their complex chemical composition. In this study, we generated a series of mRNA-LNPs with varied structural properties andin vitrocellular activities by altering their processing and storage conditions to investigate the structure-activity relationship (SAR) of these nanoparticles. Our findings revealed a moderate anticorrelation between particle size distribution andin vitroactivity, while nanoparticle size and morphology exhibited only weak correlations with efficacy. Notably, the intensity of a characteristic peak, as detected by small angle X-ray scattering (SAXS), demonstrated a strong correlation within vitroactivity. Additionally, the peak width and area were moderately correlated and anticorrelated with activity, respectively. These observations suggest that a more ordered internal structure, as identified by SAXS, is likely associated with enhancedin vitroactivity of mRNA-LNPs. Further analysis using31P nuclear magnetic resonance indicated that lyophilization may induce phase separation of mRNA and lipids within the LNPs, leading to a diminished SAXS peak and reducedin vitroactivity. Overall, our study establishes an SAR for mRNA-LNPs, highlighting that a more ordered internal structure correlates with higher efficacy, which could be instrumental in high-throughput screening of LNP libraries for mRNA loading and monitoring efficacy during storage.

Significance

In addition to the two mRNA-based vaccines developed for COVID-19, the FDA approved a third mRNA-based vaccine for respiratory syncytial virus in 2024. These vaccines rely on mRNA-containing lipid nanoparticles (mRNA-LNPs), which are also being explored for cancer treatment, gene editing, and antibody production. Despite their promise, the structure-activity relationship of mRNA-LNPs remains inadequately understood, limiting the optimization of lipid formulations, long-term storage, and mechanistic insights. Our study suggests that an ordered internal structure correlates with enhancedin vitroefficacy, consistent with previous findings. We identified the intensity of a peak at ∼0.13 Å-1in small-angle X-ray scattering (SAXS) as a strong indicator of the efficacy. Furthermore, we demonstrated that this correlation aroused from phase separation between mRNA and lipids inside LNPs in the lyophilization process. These insights offer a straightforward method to predictin vitroactivity of mRNA-LNPs with certain formulation and shed light on the mechanisms underlying the loss of mRNA-LNPs’ activity during lyophilization.

Table of Content
Nucleocytoviricotaviral factories are transient organelles made by liquid-liquid phase separation
Authors: Rigou Sofia , Schmitt Alain , Moreno Anaísa B , Lartigue Audrey , Danner Lucile , Giry Claire , Trabelsi Feres , Belmudes Lucid , Olivero-Deibe Natalia , Guenno Hugo Le , Couté Yohann , Berois Mabel , Legendre Matthieu , Jeudy Sandra , Abergel Chantal , Bisio Hugo
Published: September 2024

Phase separation is a widespread mechanism in viral processes, mediating replication, host manipulation and virion morphogenesis. The phylumNucleocytoviricotaencompasses diverse and ubiquitous viruses, includingPoxviridae,the climate-modulatingEmiliania huxleyivirus and other so-called Giant Viruses. Cytoplasmic members of this phylum form viral factories but their nature has remained unresolved. Here, we demonstrate that these viral factories are formed by liquid-liquid phase separation. We prove that mimivirus viral factories are formed by multilayered phase separation, orchestrated by at least two scaffold proteins. To extend these findings across the phylumNucleocytoviricota, we developed a bioinformatic pipeline to predict scaffold proteins based on a conserved molecular grammar, despite major primary sequence variability. Scaffold candidates were validated inMarseilleviridaeandPoxviridae, highlighting a role of H5 as a scaffold protein in the vaccinia virus. Finally, we provide a repertoire of client proteins of the nucleus-like viral factory of mimivirus and demonstrate important sub-compartmentalization of functions, including those related to the central dogma. Overall, we reveal a new mechanism for an organelle to deploy nuclear-like functions entirely based on phase separation and re-classified phylumNucleocytoviricotaviral factories as biomolecular condensates.
How do socioeconomic trajectories experienced during early adulthood contribute to the development of cardiometabolic health in young adults?
Authors: Winpenny Eleanor , Stochl Jan , Hughes Alun , Tilling Kate , Howe Laura D
Published: September 2024

Introduction

Socioeconomic position has been strongly associated with cardiovascular health. However, little is known about the short-term health impacts of socioeconomic exposures during early adulthood. In this study we describe distinct socioeconomic trajectories of early adulthood (age 16-24y), and assess associations of these trajectories with measures of cardiometabolic health at age 24y.

Methods

Participants of the Avon Longitudinal Study of Parents and Children (ALSPAC), with data across age 16y to 24y (2007-2017) were included (n=7,568). Longitudinal latent class analysis identified socioeconomic trajectories, based on education and employment data across ages 16–24y. Cardiometabolic outcomes at age 24y comprised anthropometric, vascular, metabolic and cardiovascular structure and function measures. We modelled differences in cardiometabolic outcomes at age 24y across the socioeconomic trajectory classes, adjusting for childhood socioeconomic position, adolescent health behaviours and adolescent health.

Results

Four early adulthood socioeconomic trajectories were identified: (1) Higher Education (41% of the population), (2) Extended Education (9%), (3) Part-Time Employment (21%), and (4) Early Employment (29%). Associations between socioeconomic trajectory and cardiometabolic outcomes differed by sex. Among males, the Higher Education and Extended Education classes showed a healthier cardiometabolic profile, and the Part-time Employment class the least healthy. Among females there was less clear distinction between the classes, and the pattern across different outcomes was not consistent.

Conclusion

The newly identified ‘Part-time Employment’ class showed the least healthy cardiometabolic profile, and further research should focus on this group to understand the exposures contributing to poor cardiometabolic health in this sector of the population.
Understanding Tooth Agenesis: A Multi-omics Insight into MicroRNA Regulation
Authors: Ranjan Prashant , Devi Chandra , Verma Neha , Bansal Rajesh , Srivastava Vinay Kumar , Das Parimal
Published: September 2024

This study reveals novel microRNAs (miRNAs) implicated in congenital tooth agenesis (CTA), a common dental anomaly with a complex genetic basis. Through a multi-omics approach combining bioinformatics, whole exome sequencing, metabolite analysis, and gene expression profiling, we identified eight key miRNAs potentially involved in tooth development. Among these, four miRNAs viz. miR-218-5p, miR-15b-5p, miR-200b-3p, and let-7a-3p were validated as significant regulators in CTA, marking their first investigation in blood samples from CTA patients. Our analysis revealed that these miRNAs play critical roles in odontogenesis, influencing essential signaling pathways, includingWnt, FGF, andPI3kinase pathways. Among these four, miR-218-5p and let-7a-3p emerged as key players in dental tissue morphogenesis, each contributing to previously unidentified networks crucial for tooth development.

This study highlights the potential of these miRNAs as non-invasive biomarkers for early CTA diagnosis and therapeutic targets. This is the first comprehensive investigation of these specific miRNAs in CTA, utilizing a multi-omics approach to offer fresh insights into miRNA-mediated mechanisms and their role in regulating dental anomalies. Our findings not only advance the understanding of the genetic regulation of tooth development but also pave the way for personalized approaches in managing dental anomalies. Further research is needed to validate these results and explore their clinical applications.

Graphical Abstract
Generation of motor program diversity and variability through inhibitory circuit motifs in theDrosophilalarval locomotor system
Authors: Francis Jacob , Gibeily Caius R. , Smith William V. , Petropoulos Isabel S. , Anderson Michael , Heitler William J. , Prinz Astrid A. , Pulver Stefan R.
Published: September 2024

How do neural networks generate and regulate diversity and variability in motor outputs with finite cellular components? Here we examine this problem by exploring the role that inhibitory neuron motifs play in generating mixtures of motor programs in the segmentally organisedDrosophilalarval locomotor system. We developed a computational model that is constrained by experimental calcium imaging data. The model comprises single-compartment cells with a single voltage-gated calcium current, which are interconnected by graded excitatory and inhibitory synapses. Local excitatory and inhibitory neurons form conditional oscillators in each hemisegment. Surrounding architecture reflects key aspects of inter- and intrasegmental connectivity motifs identified in the literature. The model generates metachronal waves of activity that recapitulate key features of fictive forwards and backwards locomotion, as well as bilaterally asymmetric activity in anterior regions that represents fictive head sweeps. The statistics of inputs to competing command-like motifs, coupled with inhibitory motifs that detect activity across multiple segments generate network states that promote diversity in motor outputs, while at the same time preventing maladaptive overlap in motor programs. Overall, the model generates testable predictions for connectomics and physiological studies while providing a platform for uncovering how inhibitory circuit motifs underpin generation of diversity and variability in motor systems.
PPARγ, a key modulator of metabolic reprogramming, stemness and chemoresistance associated with retrodifferentiation in human hepatocellular carcinomas
Authors: Daniel Yoann , Rauch Claudine , Moutaux Lucille , Fekir Karim , Desquilles Lise , Cano Luis , Catheline Daniel , Pierre Servane , Burel Agnès , Savary Camille , Ribault Catherine , Bendavid Claude , Clément Bruno , Aninat Caroline , Rioux Vincent , Musso Orlando , Fromenty Bernard , Cabillic Florian , Corlu Anne
Published: September 2024

Human hepatocellular carcinomas (HCCs) with cancer stem cell (CSC) features are a subclass of therapeutically challenging cancers. We recently showed that retrodifferentiation of hepatic cancer cells into CSC-like cells leads to metabolic reprogramming and chemoresistance. The molecular mechanisms whereby differentiated cancer cells switch towards a CSC phenotype are poorly understood. By studying metabolic reprogramming associated with HCC cell plasticity, we identified an unsuspected role of peroxisome proliferator-activated receptor (PPAR)γ in hepatic CSC phenotype acquisition.

Gene expression and metabolic analyses performed throughout cell differentiation/retrodifferentiation process of human HepaRG and HBG-BC2 HCC cells show that metabolic reprogramming in hepatic CSCs is associated with fragmented mitochondrial network, decreased respiration,de novolipogenesis, fatty acid oxidation, but increased glycolysis and lipid storage. Mitochondrial genes downregulated in HepaRG-CSCs are also downregulated in the STEM HCC subclass. While PPARα is the main isoform in differentiated hepatic cells, we find high PPARγ expression in hepatic CSCs. Accordingly, nuclear localization of PPARγ is detected in human HCC tumors and PPARγhigh/PPARαlowexpression is associated with the STEM HCC subclass and a poor outcome in human HCC cohorts. PPARγ silencing or/and inhibition of its target gene pyruvate dehydrogenase kinase 4 reactivates cell respiration, increases reactive oxygen species production and sensitizes hepatic CSCs to chemotherapy. Conversely, PPARα activation synergizes with chemotherapy to induce cell death.

Targeting PPARγ, a key regulator of metabolic reprogramming and stemness in hepatic CSCs, or modulating the PPARγ/PPARα balance that finely tunes the differentiation/retrodifferentiation process in HCC deserves further investigation for antitumor therapy.

Implications heading and statement

PPARγ, a key regulator of metabolic reprogramming and stemness in hepatic CSC, reduces oxidative phosphorylation and reactive oxygen species production, therefore contributing to HCC chemoresistance.
Molecular insights into dynamic protein structures by high-contrast crosslinking mass spectrometry
Authors: Chen Zhuo Angel , Absmeier Eva , Stowell James , Sinn Ludwig Roman , Shakeel Shabih , Sijacki Tamara , Njo Kendra , Stahl Kolja , Rullmann Edward , O’Reilly Francis J. , Passmore Lori A. , Rappsilber Juri
Published: September 2024

Proteins are comprised of structured domains and dynamic regions, and both are essential for biological function. However, studying dynamic regions is challenging using most structural biology methods, including crosslinking mass spectrometry. Here, we dramatically improve the usefulness of distance restraints from crosslinking MS by taking advantage of short-lived reactive species generated from diazirine-based photo-crosslinking. This leads to a clear view of complex topologies and conformational changes, including in dynamic regions. We demonstrate that photo-crosslinking MS data can be used to model flexible regions and conformational changes in the DNA repair complexes; Fanconi Anemia core complex and FANCD2-FANCI. In addition, we obtain new insights into the architecture and arrangement of the highly flexible CCR4-NOT mRNA deadenylation complex. The improved contrast of photo-crosslinking empowers structural biology by providing clearer structural insights into dynamic biological systems that have eluded other structural biology approaches.
Yeast-MetaTwin for Systematically Exploring Yeast Metabolism through Retrobiosynthesis and Deep Learning
Authors: Wu Ke , Liu Haohao , Sun Manda , Mao Runze , Jiang Yindi , Kerkhoven Eduard J. , Chen Yu , Nielsen Jens , Li Feiran
Published: September 2024

Underground metabolism plays a crucial role in understanding enzyme promiscuity, cellular metabolism, and biological evolution, yet experimental exploration of underground metabolism is often sparse. Even though yeast genome-scale metabolic models have been reconstructed and curated for over 20 years, more than 90% of the yeast metabolome is still not covered by these models. To address this gap, we have developed a workflow based on retrobiosynthesis and deep learning methods to comprehensively explore yeast underground metabolism. We integrated the predicted underground network into the yeast consensus genome-scale model, Yeast8, to reconstruct the yeast metabolic twin model, Yeast-MetaTwin, covering 16,244 metabolites (92% of the total yeast metabolome), 2,057 metabolic genes and 59,914 reactions. We revealed thatKmparameters differ between the known and underground network, identified hub molecules connecting the underground network and pinpointed the underground percentages for yeast metabolic pathways. Moreover, the Yeast-MetaTwin can predict the by-products of chemicals produced in yeast, offering valuable insights to guide metabolic engineering designs.
Nanoscale visualization ofDrosophilaE-cadherin ectodomain fragments and their interactions using DNA origami nanoblocks
Authors: Oda Hiroki , Nishiguchi Shigetaka , Song Chihong , Murata Kazuyoshi , Uchihashi Takayuki , Suzuki Yuki
Published: September 2024

The adhesive function of cell surface proteins can be visually assessed through direct observation; however, the underlying structures that mediate adhesion typically remain invisible at the nanoscale level. This hinders knowledge on the diversity of molecular architectures responsible for cell-cell adhesion.DrosophilaE-cadherin (DE-cadherin), a classical cadherin with a unique domain structure, demonstrates adhesive function; however, it lacks a structural model that explains its adhesion mechanism. In this study, we present a novel application of DNA origami technology to create a cell-free, flat environment in which full DE-cadherin ectodomains are anchored using SNAP-tags and biotin-streptavidin interactions. DNA origami was assembled into a 120 nm long block, bearing 5 or 14 biotin:streptavidin sites that were evenly spaced on one lateral face. DE-cadherin ectodomain fragments were attached via biotinylated SNAP-tags. These decorated DNA origami nanoblocks were subjected to transmission electron and high-speed atomic force microscopy, which revealed a hinge-like site that separated the membrane-distal and -proximal portions of the DE-cadherin ectodomain, suggesting a role in mechanical flexibility. We also observed interactions between DE-cadherin ectodomains via their membrane-distal portions on single DNA origami nanoblocks. We reconstituted an adhesion-like process via pairing DNA origami nanoblocks using DE-cadherin ectodomain interactions. Homophilic associations of functional DE-cadherin ectodomains between the paired DNA origami nanoblocks were visualized at the nanoscale, displaying strand-like molecular configurations, likely representing the extracellular cadherin repeats without regular arrays of structural elements. This study introduces a DNA origami-based platform for reconstituting and visualizing cadherin ectodomain interactions, with potential applications for a broader range of adhesion molecules.

Highlights

DNA origami technology was applied to perform a structure-function study of cadherin.

DNA origami nanoblocks decorated with DE-cadherin ectodomains were observed by TEM/HS-AFM.

A hinge-like site that separated the membrane-distal and -proximal portions of the DE-cadherin ectodomain was revealed.

An adhesion-like process was mimicked via pairing two nanoblocks using DE-cadherin ectodomain interactions.

Homophilic associations of DE-cadherin ectodomains between the nanoblocks were visualized at the nanoscale level.
eProbe: a capture probe design toolkit for genetic diversity reconstructions from ancient environmental DNA
Authors: Huang Zihao , Gu Zhengquan , Cai Yuanyang , Macleod Ruairidh , Xue Zhe , Dong Haoran , Overballe-Petersen Søren , Liu Shanlin , Gao Yu , Li Hao , Tang Sha , Diao Xianmin , Joergensen Morten Egevang , Dockter Christoph , Vinner Lasse , Willerslev Eske , Chen Fahu , Wang Hongru , Wang Yucheng
Published: September 2024

Ancient environmental DNA (aeDNA) is now commonly used in paleoecology and evolutionary ecology, yet due to difficulties in gaining sufficient genome coverage on individual species from metagenome data, its genetic perspectives remain largely uninvestigated. Hybridization capture has proven as an effective approach for enriching the DNA of target species, thus increasing the genome coverage of sequencing data and enabling population and evolutionary genetics analysis. However, to date there is no tool available for designing capture probe sets tailored for aeDNA based population genetics. Here we presenteProbe, an efficient, flexible and easy-to-use program toolkit that provides a complete workflow for capture probe design, assessment and validation. By benchmarking a probe set for foxtail millet, an annual grass, made by theeProbeworkflow, we demonstrate a remarkable increase of capturing efficiency, with the target taxa recovery rate improved by 577-fold, and the genome coverage achieved by soil capture-sequencing data even higher than data directly shotgun sequenced from the plant tissues. Probes that underwent our filtering panels show notably higher efficiency. The capture sequencing data enabled accurate population and evolutionary genetic analysis, by effectively inferring the fine-scale genetic structures and patterns, as well as the genotypes on functional genes.
Bacteriophages Phi 8 and Phi 12 host infection are inhibited by OMVs and LPS purified fromP. pseudoalcaligenesstrain: East River Isolate A
Authors: Robinson Charles F. , Khayat Reza
Published: September 2024

Cystoviridaeis a family of double stranded RNA (dsRNA) phage that infects various strains ofPseudomonas syringae, a Gram-negative soil bacteria known to infect various crops. Surrounding the icosahedral capsids of these phages is a bacterial derived phospholipid membrane. Embedded within this membrane is a multi-component protein complex, referred to as the spike complex. The spike complex is responsible for host recognition and membrane fusion. We studied the ability of two members of theCystivirdaefamily to infect cells in the presence of purified outer membrane vesicles (OMVs) and lipopolysaccharide (LPS) derived from distinct sources. In this study we determined that OMVs from the hostPseudomonas pseudoalcaligenesstrain: East River isolate A (ERA) inhibit Phi 8 and Phi 12 host infection. These OMVs range in size from 30 to 60 nm and bind to Phi 8 and Phi 12. However, OMV purified fromP. syringaepv. phaseolicola LM2691 andE. coliΔyciBΔdcrBdid not inhibit Phi 8 or Phi 12 host infection. However, LPS derived from ERA and LM2691 inhibited Phi 8 and Phi 12 infection, demonstrating that LPS is the receptor for these two viruses, and that OMV biogenesis is selective of LPS. LPS derived from other non-CystoviridaeGram-negative bacteria, did not inhibit infection. We confirmed that host proteins are not required for Phi 8 or Phi 12 host interaction. Our results also suggest that differences in lipid A and the core polysaccharide in LPS may influence Phi 8 and Phi 12 host binding.

IMPORTANCE

Most phage families studied to date use a tailed appendage, composed of a multitude of proteins, for cellular recognition, membrane penetration, and genome injection. This contrasts with members of theCystoviridaefamily which possess a phospholipid membrane bilayer with embedded proteins responsible for cellular recognition and membrane fusion. Thus, theCystoviridaeare akin to enveloped viruses which also use protein complexes embedded into their membrane for cellular recognition and membrane fusion. Examples of such viruses include theRetroviridae, Coronoviridae, Herpesviridae, andOrthomyxoviridaefamilies. The binding specifics ofCystoviridaeto the host outer membrane are unknown. UsingCystoviridae-OMV interaction we began to uncover the host requirements for bindingCystoviridae. The results presented determine that only lipid A and the core polysaccharide of LPS are required forCystoviridaeouter membrane binding.
Identification of nuclear pore proteins at plasmodesmata
Authors: Schladt T. Moritz , Miras Manuel , Ejike Jona Obinna , Pottier Mathieu , Xi Lin , Restrepo-Escobar Andrea , Nakamura Masayoshi , Pütz Niklas , Hänsch Sebastian , Gao Chen , Engelhorn Julia , Dickmanns Marcel , Davis Gwendolyn V. , Dalal Ahan , Gombos Sven , Lange Ronja , Simon Rüdiger , Schulze Waltraud X. , Frommer Wolf B.
Published: September 2024

Plasmodesmata (PD) exchange small molecules, RNAs and proteins between plant cells with an exclusion limit for passive, non-specific transport. PD also facilitate the transport of specific cargo that may require chaperones or carriers for transport. The mechanisms underlying PD transport are not understood. PD and nuclear pore complexes (NPC), are nanometer size micropores with similar transport properties. The permeability barrier in NPCs is a phase separation domain formed by FG-nucleoporins (FG-NUP). Here we used bioinformatics, proteomics and fluorescence imaging to identify proteins with similarities to phase separating FG-NUPs at PD. We identified 20 NUPs in PD fractions, and validated dual localization to NPC and PD for 7 NUPs. Structured illumination microscopy detected the transmembrane anchor NUP CPR5 at both orifices of PD.cpr5mutants showed reduced intercellular transport of SHR from stele to endodermis. The identification of FG-NUPs at PD is consistent with the recruitment of NUPs in the green lineage to form a PD pore gating complex consistent with liquid phase separation domains as diffusion barriers at PD.
Hypothermia protects against ventilator-induced lung injury by limiting IL-1β release and NETs formation
Authors: Nosaka Nobuyuki , Borges Vanessa , Martinon Daisy , Crother Timothy R , Arditi Moshe , Shimada Kenichi
Published: September 2024

Although mechanical ventilation is a critical intervention for acute respiratory distress syndrome (ARDS), it can trigger an IL-1β-associated complication known as ventilator-induced lung injury. In mice, we found that LPS and high-volume ventilation, LPS-HVV, leads to hypoxemia with neutrophil extracellular traps (NETs) formation in the alveoli. Furthermore,Il1r1-/-LPS-HVV mice did not develop hypoxemia and had reduced NETs, indicating that IL-1R1 signaling is important for NETs formation and hypoxemia. Therapeutic hypothermia (TH) is known to reduce the release of inflammatory mediators. In LPS-HVV mice, TH (32 °C body temperature) prevented hypoxemia development, reducing albumin leakage, IL-1β, gasdermin D (GSDMD) and NETs formation. We also observed that LPS-primed macrophages, when stimulated at 32°C with ATP or nigericin, release less IL-1β associated with reduced GSDMD cleavage. Thus, hypothermia is an important modulating factor in the NLRP3 inflammasome activation, IL-1β release and NETs formation, preventing LPS-HVV-induced acute respiratory failure.
Design of an Artificial Natural Killer Cell Mimicking System to Target Tumour Cells
Authors: Chugh Vaishali , Krishna K. Vijaya , Quandt Dagmar , Kelly Suainibhe , King Damien , Jensen Lasse D. , Simpson Jeremy C , Pandit Abhay
Published: September 2024

NK cell mimics are assemblies of a cell membrane and a template that replicate biomimetic features and physicochemical properties, respectively. For the reported design, we used the cell membrane from human NK cell (KHYG-1) line and gelatin microspheres as a template. The gelatin microspheres were reinforced via DMTMM cross-linking in a water-in-oil emulsion to exhibit tunable Young’s modulus. These engineered NK cell mimics were found to be non-toxic, non-inflammatory, and capable of evading macrophage detection when tested with differentiated THP-1 cells.In vitrostudies showed significant interaction/proximity of the mimics with cancer cells when tested in 2D cultures of breast cancer cells (MDA-MB-231), 3D spheroids of liver (HepG2) and colon (HT-29) cancer cell models, and a zebrafish breast cancer xenograft (MDA-MB-231) model. The NK cell mimics also evaded macrophage detection in a Kdrl:EGFP Spil: Ds Red zebrafish model. In a pilot assessment, loading and release of the sialyltransferase inhibitor (STI, 3Fax-Peracetyl Neu5Ac) using NK cell mimics significantly reduced α-2,6 sialylation in 2D cultures of MDA-MB-231 cells, demonstrating the STI’s intact functionality in inhibiting sialylation. These findings collectively underscore the promising potential of engineered NK cell mimics as versatile tools in cancer research and therapeutic applications.
ETTIN-mediated auxin signalling is an angiosperm-specific neofunctionalization for carpel development
Authors: Ang Aaron Chun Hou , Mutte Sumanth , Weijers Dolf , Østergaard Lars
Published: September 2024

The phytohormone auxin affects processes throughout plant growth and development. While auxin signalling has been mainly attributed to a repressor degradation-based pathway, numerous alternative mechanisms for how auxin mediates its effect have been revealed in recent years. One such mechanism involves a direct auxin-induced switch in the transcriptional regulatory activity of the Auxin Response Factor (ARF) ETTIN (ETT). ETT lacks a conserved C-terminus domain involved in canonical pathway interactions but contains a middle region domain mediating auxin binding. As the ETT clade only exists in the angiosperms, it remains unknown when the pathway evolved. Here we provide evidence for a two-step origin of the ETT clade and its neofunctionalisation through the gain of auxin perception in gynoecium patterning. Phylogenetic analyses reveal the structural divergence of ETT and its paralogue ARF4 after their duplication from an ancestral euphyllophyte ARF3/4 clade. Auxin sensitivity was identified as an ETT-specific innovation that likely originated in the last common angiosperm ancestor. Furthermore,in plantacomplementation experiments demonstrated the full genetic redundancy of ETT and ARF4 in leaf and ovary development, but a specialised role for the ETT-mediated auxin signalling pathway in style development. Our work thus provides evidence that ETT was recruited from an ancestral role in leaf development and subsequently underwent neofunctionalisation through the acquisition of direct auxin sensing for a novel role in gynoecium patterning.
The speed limits for tau pathology progression in Alzheimer’s disease
Authors: Hoenig Merle C. , Dzialas Verena , Doering Elena , Bischof Gérard N. , van Eimeren Thilo , Drzezga Alexander ,   Alzheimer’s Disease Neuroimaging Initiative
Published: September 2024

Objective

To examine interactive effects of modifiable factors, genetic determinants and load-dependent pathology effects on tau pathology progression.

Methods

Data of 162 amyloid-positive individuals were included, for whom longitudinal [18F]AV-1451-PET scans, baseline information on global amyloid load, ApoE4 status, body-mass-index (BMI), hypertension, education, neuropsychiatric symptom severity and demographic information were available in ADNI. All [18F]AV-1451 PETs were intensity-standardized (reference: inferior cerebellum), z-transformed (control sample: 147 amyloid-negative subjects) and subsequently thresholded (z-score > 1.96) and converted to volume-maps. Based on these volume-maps, tau-changes over time were assessed in terms of 1)tau-speed(i.e. newly affected volume at follow-up), and 2)tau-level-rise(i.e. tau increase in previously affected volume). These two measures were entered as dependent variables in separate linear mixed effects models including four baseline risk factors (BMI, education, hypertension, neuropsychiatric symptom severity), baseline amyloid, tau-volume or tau burden, ApoE4 status, clinical stage, sex, and age as predictors. Next, we tested the interactive effects between baseline amyloid or tau burden with the four modifiable factors on either tau-speed or tau-level-rise, respectively.

Results

Faster tau-speed was linked to higher BMI, female sex, ApoE4-status, and baseline tau-volume. The effect of baseline tau-volume on tau-speed was driven by greater global amyloid burden. In terms of tau-level-rise, we observed that lower hypertension and BMI were linked to a slower increase in tau burden. A load-dependent effect of baseline amyloid and tau burden was found. Higher amyloid and BMI as well as lower education and higher tau burden were linked to greater tau-level-rise.

Conclusion

Education, BMI and hypertension differentially influence tau speed and level rise by its interaction with initial pathological burden. Timely modification of these factors may overall slow tau’s progression.
Biases in volumetric versus surface analyses in population receptive field mapping
Authors: Linhardt David , Woletz Michael , Paz-Alonso Pedro M. , Windischberger Christian , Lerma-Usabiaga Garikoitz
Published: September 2024

Population receptive field (pRF) mapping is a quantitative fMRI analysis method that links visual field positions with specific locations in the visual cortex. A common preprocessing step in pRF analyses involves projecting volumetric fMRI data onto the cortical surface, often leading to upsampling of the data. This process may introduce biases in the resulting pRF parameters. To investigate this, we present CON-pRF, a fully containerized pipeline for pRF mapping analysis that is designed to maximize reproducibility in pRF mapping studies. Using this pipeline, we compared pRF maps generated from original volumetric with those from upsampled surface data. Our results show substantial increases in pRF coverage in the central visual field of upsampled data sets. These effects were consistent across early visual cortex areas V1-3. Further analysis indicates that this bias is primarily driven by the non-linear relationship between cortical distance and visual field eccentricity, known as cortical magnification. Our results demonstrate that reproducible analysis pipelines enable the detection of potential biases introduced by varying processing steps, particularly when comparing across differently processed datasets.

Key Points

Spatial upsampling increases pRF coverage in the fovea due to enhanced CNR and cortical magnification.

The study highlights the need for careful consideration of data processing steps.

CON-pRF is a containerized pipeline for enhanced reproducibility in pRF analysis.
Early Fibrin Biofilm Development in Cardiovascular Infections
Authors: Oukrich Safae , Hong Jane , Leon-Grooters Mariël , van Cappellen Wiggert , Slotman Johan A. , Koenderink Gijsje H. , van Wamel Willem J.B. , de Maat Moniek P.M. , Kooiman Klazina , Lattwein Kirby R.
Published: September 2024

The single most common microbe causing cardiovascular infections isStaphylococcus aureus(S. aureus).S. aureusproduces coagulase that converts fibrinogen to fibrin, which is incorporated into biofilms. This process aids in adherence to intravascular structures, defense against the host immune system, and resistance to antimicrobial treatment. Despite its significance, fibrin formation inS. aureusbiofilms remains poorly understood. Therefore, this study aimed to elucidate the early development of cardiovascular biofilms. Clinically isolated coagulase-positiveS. aureusand coagulase-negativeStreptococcus gordonii(S. gordonii) from patients with cardiovascular infections, and a coagulase mutantS. aureusΔcoa, were grown in tryptic soy broth (TSB), Iscove’s Modified Dulbecco’s Medium (IMDM), and pooled human plasma, with or without porcine heart valves. Bacterial growth, metabolic activity, and bacterial fibrinogen utilization were measured over 24 hr at 37 °C. Time-lapse confocal microscopy was used to visualize and track biofilm development.S. aureusexhibited more growth in TSB and human plasma thanS. gordoniiandS. aureusΔcoa, but showed similar growth asS. aureusΔcoa in IMDM. Peak metabolic activity for all isolates was highest in TSB and lowest in human plasma. The presence of porcine valves caused strain-dependent alterations in time to peak metabolic activity. Confocal imaging revealed fibrin-based biofilm development exclusively in the coagulase-producingS. aureusstrains. Between 2 and 6 hr of biofilm development, 74.9% (p=0.034) of the fibrinogen from the medium was converted to fibrin. Variations in fibrin network porosity and density were observed among different coagulase-producingS. aureusstrains. Fibrin formation is mediated byS. aureuscoagulase and first strands occurred within 3 hr for clinical strains after exposure to human plasma. This study stresses the importance of experimental design given the bacterial changes due to different media and substrates and provides insights into the early pathogenesis ofS. aureuscardiovascular biofilms.

Highlights

Bacterial growth and activity are medium and substrate dependent

Coagulase is necessary forStaphylococcus aureusfibrin biofilm development

Fibrin strands begin forming inStaphylococcus aureusbiofilms within 3 hours
A newBetaflexivirusspecies inFerraria crispaplants
Authors: Dekker Rob J. , de Leeuw Wim C. , van Olst Marina , Ensink Wim A. , van Leeuwen Selina , Breit Timo M. , Jonker Martijs J.
Published: September 2024

The global virome is still largely unknown. In this study we describe the discovery of a newBetaflexivirusspecies inFerraria crispaplants. The plant samples were collected in an experiment of 25Asparagalesplants, obtained from an urban botanic garden in the Netherlands, and analyzed by smallRNA-seq as well as RNA-seq. The newBetaflexivirusonly occurred in fourFerraria cripsaplants and was tentatively named Ferraria Betaflexivirus 1 (FerBfV-1). ThisBetaflexivirusshowed an RNA genome structure characteristic for the genusCapillovirus, a single-stranded (+) RNA virus. The closest knownCapillovirusesin GenBank showed a protein similarity less than 49%, which is well below the proposed demarcation value of species in this genus. Further comparison analysis of theCapillovirusgenus revealed the possible presence of two groups.
Mitochondrial Deletions and Healthspan: Lessons from the Common Repeat
Authors: Shamanskiy Victor , Gunbin Konstantin , Tretiakov Evgenii O. , Mazunin Ilya , Skripskaya Viktoria , Mikhailova Alina A. , Mikhailova Alina G. , Ree Natalia , Timonina Valeriia , Knorre Dmitry , Kunz Wolfram S. , Fiorellino Nathan , Reymond Alexandre , Bazykin Georgii A. , Fellay Jacques , Tanaka Masashi , Khrapko Konstantin , Popadin Konstantin
Published: September 2024

Aging, characterized by a series of functional declines correlated with advancing chronological age, has a significant mitochondrial DNA (mtDNA) component, with somatic mtDNA deletions playing a central role. In post-mitotic or slow-dividing cells like neurons and skeletal muscles, selfish mtDNA deletions clonally expand within a cell, ultimately leading to the deterioration and death of host cells and appearence of age-related phenotypes. Thus reducing the burden of somatic deletions could have far- reaching systemic benefits for the entire human body. Given the crucial role of direct nucleotide repeats in the formation of mitochondrial deletions, we hypothesize that minimizing these repeats in the human mitochondrial genome could enhance healthspan by decreasing somatic deletions. To investigate this hypothesis, we focus on the “common repeat”, a 13-base pair perfect direct repeat sequence (ACCTCCCTCACCA) located at positions 8470-8482 and 13447-13459, respectively. This perfect repeat: (i) is highly prevalent, with its potential deleterious consequences affecting the majority of humans; (ii) represents one of the most fragile sites, highly prone to forming deletions; (iii) when disrupted, is associated with a decreased somatic deletion load and enhanced human healthspan; (iv) is likely to experience positive selection in the present or near future due to indirect fitness effects, such as the “grandmother effect”, and direct fitness effects, such as (v) a decreased mutation rate. These observations support the argument that reducing the mtDNA somatic deletion load through targeted disruption of these repeats, or by using naturally occurring polymorphisms with disrupted repeats in mitochondrial medicine, could be an effective approach to increasing human longevity.
Aneuploidy of Specific Chromosomes is Beneficial to Cells Lacking Spindle Checkpoint Protein Bub3
Authors: Gadgil Pallavi , Ballew Olivia , Sullivan Timothy J. , Lacefield Soni
Published: September 2024

Aneuploidy typically poses challenges for cell survival and growth. However, recent studies have identified exceptions where aneuploidy is beneficial for cells with mutations in certain regulatory genes. Our research reveals that cells lacking the spindle checkpoint geneBUB3exhibit aneuploidy of select chromosomes. While the spindle checkpoint is not essential in budding yeast, the loss ofBUB3andBUB1increases the probability of chromosome missegregation compared to wildtype cells. Contrary to the prevailing assumption that the aneuploid cells would be outcompeted due to growth defects, our findings demonstrate thatbub3Δ cells consistently maintained aneuploidy of specific chromosomes over many generations. We investigated whether the persistence of these additional chromosomes inbub3Δ cells resulted from the beneficial elevated expression of certain genes, or mere tolerance. We identified several genes involved in chromosome segregation and cell cycle regulation that confer an advantage to Bub3-depleted cells. Overall, our results suggest that the upregulation of specific genes through aneuploidy may provide a survival and growth advantage to strains with poor chromosome segregation fidelity.

AUTHOR SUMMARY

Accurate chromosome segregation is crucial for the proper development of all living organisms. Errors in chromosome segregation can lead to aneuploidy, characterized by an abnormal number of chromosomes, which generally impairs cell survival and growth. However, under certain stress conditions, such as in various cancers, cells with specific mutations and extra copies of advantageous chromosomes exhibit improved survival and proliferation. In our study, we discovered that cells lacking the spindle checkpoint protein Bub3 became aneuploid, retaining specific chromosomes. This finding was unexpected because althoughbub3Δ cells have a higher rate of chromosome mis-segregation, they were not thought to maintain an aneuploid karyotype. We investigated whether the increased copy number of specific genes on these acquired chromosomes offered a benefit to Bub3-deficient cells. Our results revealed that several genes involved in chromosome segregation and cell cycle regulation prevented the gain of chromosomes upon Bub3-depletion, suggesting that these genes confer a survival advantage. Overall, our study demonstrates that cells lacking Bub3 selectively retain specific chromosomes to increase the copy number of genes that promote proper chromosome segregation.
The contributions of biological maturity and experience to fine motor development in adolescence
Authors: Berencsi Andrea , Gombos Ferenc , Fehér Lili Julia , Gerván Patrícia , Utczás Katinka , Oláh Gyöngyi , Tróznai Zsófia , Kovács Ilona
Published: September 2024

Fine motor function develops into adulthood, but little is known about the differential effects of biological maturation and experience on speed and complex sequential performance of the hand. To determine maturity levels, ultrasonic bone age (BA) was assessed in 225 adolescents (123 females; BA range: 9.9 to 17.9 years). The role of experience was evaluated based on chronological age (CA, range: 11.1 to 16.5 years), musical instrumental experience, and handedness. Multiple linear regression modeling showed that BA is the strongest predictor of sequential motor performance, while CA influenced motor speed when no musical instrumental experience was present. When present, the amount of highly specific musical instrumental experience becomes the main predictor of sequential performance.
A FUCCI sensor reveals complex cell cycle organization ofToxoplasmaendodyogeny
Authors: Batra Mrinalini , Marsilia Clem , Awshah Danya , Hawkins Lauren M. , Wang Chengqi , Chaput Dale , Naumova Daria A. , Suvorova Elena S.
Published: September 2024

In this study, we report the atypical cell cycle organization of the unicellular eukaryotic pathogenToxoplasma gondii. The remarkably flexible cell division ofT. gondiiand other apicomplexan parasites differs considerably from the cell division modes employed by other model eukaryotes. Additionally, there is a lack of recognizable cell cycle regulators, which have contributed to the difficulties in deciphering the order of events in the apicomplexan cell cycle. To aid in studies of the cell cycle organization of theT. gondiitachyzoite, we have created the Fluorescent Ubiquitination-based Cell Cycle Indicator (FUCCI) probes,ToxoFUCCISandToxoFUCCISC. We introduced a DNA replication factor TgPCNA1 tagged with NeonGreen that can be used alone or in conjunction with an mCherry-tagged budding indicator TgIMC3 in the auxin-induced degradation (AID) parental strain. The varied localization and dynamic cell cycle oscillation have confirmed TgPCNA1 to be a suitableT. gondiiFUCCI probe. TheToxoFUCCISanalysis showed that tachyzoite DNA replication starts at or near centromeric regions, has a bell-shaped dynamic and a significant degree of the cell cycle asynchrony within the vacuoles. Quantitative live and immunofluorescence microscopy analyses ofToxoFUCCISand its derivatives co-expressing epitope-tagged cell cycle markers have revealed an unusual composite cell cycle phase that incorporates overlapping S, G2, mitosis and cytokinesis (budding). We identified five intervals of the composite phase and their approximate duration: S (19%), S/G2/C (3%), S/M/C (9%), M/C (18%) and C/G1(<1%). TheToxoFUCCISprobe efficiently detected G2/M and Spindle Assembly Checkpoints, as well as the SB505124-induced TgMAPK1 dependent block. Altogether, our findings showed an unprecedented complexity of the cell cycle in apicomplexan parasites.
Design of linked-domain protein inhibitors of UBE2D as tools to study cellular ubiquitination
Authors: Bukhari Zara , Gu Li , Nederstigt Anneroos E. , Cope Logan J. , Bolhuis Derek L. , Harvey Kim , Allen Tristan , Hill Spencer , Yang Yujie , Lawson Guy , Lu Cai , Tran Tommy , Pineda Leah , Low Leanne , Chiang Andrew , Song Jason , Fong Michelle V. , Rangel Vanessa M. , Chan William K. , Kleiger Gary , Goldfarb Dennis , Vierra Craig A. , Brown Nicholas G. , Harrison Joseph S.
Published: September 2024

Ubiquitin (Ub) is a post-translational modification that largely controls proteostasis through mechanisms spanning transcription, translation, and notably, protein degradation. Ub conjugation occurs through a hierarchical cascade of three enzyme classes (E1, E2, and E3s) involving >1000 proteins that regulate the ubiquitination of proteins. The E2 Ub-conjugating enzymes are the midpoint, yet their cellular roles remain under-characterized, partly due to a lack of inhibitors. For example, the cellular roles of the promiscuous E2 UBE2D/UBCH5 are not well described. Here, we develop a highly selective, multivalent, engineered protein inhibitor for the UBE2D family that simultaneously targets the RING- and backside-binding sites. In HeLa cells, these inhibitors phenocopy knockdown of UBE2D by reducing the IC50to cisplatin and whole-cell proteomics reveal an increased abundance of ∼20% of the identified proteins, consistent with reduced Ub degradation and proteotoxic stress. These precision tools will enable new studies probing UBE2D’s central role in proteome management.
Phage-mediated intercellular CRISPRi for biocomputation in bacterial consortia
Authors: Pujar Abhinav , Pathania Amit , Hopper Corbin , Pandi Amir , Calderon Cristian Ruiz , Fugger Matthias , Nowak Thomas , Kushwaha Manish
Published: September 2024

Coordinated actions of cells in microbial communities and multicellular organisms enable them to perform complex tasks otherwise difficult for single cells. This has inspired biological engineers to build cellular consortia for larger circuits with improved functionalities, while implementing communication systems for coordination among cells. Here, we investigate the signalling dynamics of a phage-mediated synthetic DNA messaging system, and couple it with CRISPR interference to build distributed circuits that perform logic gate operations in multicellular bacterial consortia. We find that growth phases of both sender and receiver cells, as well as resource competition between them, shape communication outcomes. Leveraging the easy programmability of DNA messages, we build 8 orthogonal signals and demonstrate that intercellular CRISPRi (i-CRISPRi) regulates gene expression across cells. Finally, we multiplex the i-CRISPRi system to implement several multicellular logic gates that involve up to 7 cells and take up to 3 inputs simultaneously, with single- and dual-rail encoding: NOT, YES, AND, and AND-AND-NOT. The communication system developed here lays the groundwork for implementing complex biological circuits in engineered bacterial communities, using phage signals for communication.