bluuebunny/crossref_metadata_2025_split huggingface dataset:
Giant virus creates subcellular environment to overcome codon– tRNA mismatch
Authors: Zhang Ruixuan , Mayer Lotte , Hikida Hiroyuki , Shichino Yuichi , Mito Mari , Willemsen Anouk , Iwasaki Shintaro , Ogata Hiroyuki 
Published: October 2024

AbstractCodon usage consonant with the cellular tRNA pool is important for efficient translation. However, many eukaryotic viruses, including amoeba-infecting mimiviruses, have codon usage that is largely deviated from that of their host, despite using the host machinery for translation. This raises the question of how these viruses cope with the mismatch between tRNA supply and demand. Here we show that Acanthamoeba castellanii mimivirus generates a subcellular area in the host cells to translate virus mRNAs. A combination of genome-wide ribosome profiling and RNA sequencing showed that ribosomes traversed along viral mRNAs with fewer pausing events than were observed on amoeba mRNAs. Frequently used codons in viral mRNAs had higher tRNA accessibility than the same type of codons in amoeba mRNAs. tRNA sequencing showed that the tRNA pool was not greatly altered during the infection even though the virus encodes tRNA genes. Instead, byin situlabelling, we found that viral mRNAs and newly synthesized proteins were localized at the periphery region of the viral factory, likely creating a unique environment to facilitate viral translation. Our data provide a perspective on how local translation assists the virus in overcoming the mismatch between tRNA supply and demand.
Modular cloning of multigene vectors for the baculovirus system and yeast
Authors: Lai Zhihao , Flanigan Sarena F. , Boudes Marion , Davidovich Chen 
Published: October 2024

AbstractRecombinant macromolecular complexes are often produced by the baculovirus system, using multigene expression vectors. Yet, the construction of baculovirus-compatible multigene expression vectors is complicated and time-consuming. Furthermore, while the baculovirus and yeast are popular protein expression systems, no single method for multigene vector construction is compatible with both. Here we present the modular cloning (MoClo) Baculo toolkit for constructing multigene expression vectors for the baculovirus system and, through compatibility with the MoClo Yeast toolkit, also for yeast. Vector construction by MoClo Baculo utilises Golden Gate assembly, which does not require PCR, primers or the sequencing of intermediate products. As a proof of principle, MoClo Baculo was used to construct baculovirus and yeast multigene vectors expressing the four- and five-subunit human Polycomb Repressive Complex 2. We show that MoClo Baculo simplifies and expedites the construction of multigene expression vectors for the baculovirus system and provides compatibility with yeast as an alternative expression system.
Cardiomyopathic mutations of lamin A perturb mutual interactions of lamin, nuclear membrane, and chromatin leading to LLPS
Authors: Nath Subhradip , Dutta Shuvadip , Sarkar Shreyasi Dey , Sengupta Duhita , Mitra Mithun K. , Sengupta Kaushik 
Published: October 2024

AbstractLamins are nuclear intermediate filaments constituting the nuclear lamina which maintains the structural integrity of the nucleus and play a key role in the spatial organisation of the genome. Mutations in the lamin protein have been associated with diverse diseases collectively known as laminopathies. In this study, we focused on two lamin A mutants - E161K and K97E - associated with dilated cardiomyopathy (DCM). Through confocal imaging, we established that these mutations cause large scale disruption of the peripheral lamin and consequent heterochromatin organisation, along with the formation of lamin aggregates inside the nucleoplasm. Using coarse-grained polymer simulations, we uncovered the role of lamin-lamin, lamin-membrane and lamin-chromatin interactions in maintaining wild-type lamin and chromatin organisation and showed that disruptions in these interactions can reproduce the experimental observations in the lamin mutants. These predictions were verified using 3D-FISH experiments to quantify the reorganisation of chromosome territories in these mutants. Using advanced imaging methods, we characterised the dynamical properties of the lamin aggregates in the mutants to show for the first time a liquid-like state of the lamin aggregates through a liquid-liquid phase separation. The altered lamin and chromatin interactions in the mutants thus manifest as liquid-like aggregates in the nucleoplasm leading to disruption of the spatial organisation of the genome in these laminopathy-associated mutants.
Mutators can increase both the rate of occurrence and specificity of nitrofurantoin resistance mutations
Authors: Kettlewell Riannah , Forsyth Jessica H. , Gifford Danna R. 
Published: October 2024

AbstractAntimicrobial resistance is a significant global health crisis, with many antibiotics losing effectiveness within a decade of their introduction. The antibiotic nitrofurantoin, however, counters this trend, as it has sustained low resistance rates despite prolonged and widespread use. A key factor behind nitrofurantoin’s success is that resistance requires two independent inactivating mutations in separate genes,nfsAandnfsB. However, this inherent safeguard may be undermined by elevated mutation rates, a risk that remains unquantified for this antibiotic. Here, we investigated how mutation rates influence nitrofurantoin resistance inEscherichia coliusing experimental evolution with both low-level and high-level mutator strains, alongside genomic analyses of uropathogenic nitrofurantoin-resistant clinical isolates. Under experimental conditions, increasing mutation rates increased the development of nitrofurantoin resistnace. Moreover, changes in the mutations that occurred also shifted the spectrum of resistance mutations from broad-impact frameshifts and indels to specific amino acid substitutions in the proteins’ active sites. Of the nitrofurantoin-resistant clinical isolates we analysed, nearly 40% harboured disruptive variants in DNA replication fidelity and repair genes—a prevalence at the higher end of what is typically observed in uropathogenicE. coli. These findings indicate that elevated mutation rates pose a risk to the continued efficacy of nitrofurantoin, highlighting the need for increased monitoring of mutator-driven resistance.
Distinct Virulence Mechanisms ofBurkholderia gladioliin Onion Foliar and Bulb Scale Tissues
Authors: Paudel Sujan , Franco Yaritza , Zhao Mei , Dutta Bhabesh , Kvitko Brian H. 
Published: October 2024

AbstractSlippery skin of onion caused byBurkholderia gladiolipv.alliicola(Bga) is a common bacterial disease reported from onion growing regions around the world. Despite the increasing attention in recent years, our understanding of the virulence mechanisms of this pathogen remains limited. In this study, we characterized the predicted genetic determinants of virulence in Bga strain 20GA0385 using reverse genetics approach. Using the closely related rice pathogen,B. glumaeas a reference, comparative genomics analysis was performed to identify Bga candidate virulence factors and regulators. Marked and unmarked deletion mutants were generated using allelic exchange and the mutants were functionally validated usingin vitroandin vivoassays. The role of mutants in pathogenic phenotypes was analyzed using onion foliar/seedling necrosis assays, the Red Scale Necrosis (RSN) assay andin plantabacterial population counts. The phytotoxin toxoflavin was a major contributor to foliar necrosis and bacterial populations whereas the type II and type III secretion system (T2SS/T3SS) were dispensable for foliar symptoms. In onion scale tissue, the T2SS single mutantgspCand its double and triple mutant derivatives all contributed to scale lesion area. Neither the lipocyclopeptide icosalide, toxoflavin, nor T3SS were required for scale symptoms. Our results suggest the quorum sensingtofIMRsystem in Bga regulates, toxoflavin, T2SS, and T3SS, contributing to onion symptom production. We show different virulence factors contribute to onion tissue-specific virulence patterns in Bga and that decreases in scale symptoms often do not result in decreased Bga populations in onion tissue.
Development of Clinically Viable Non-Muscle Myosin II Small Molecule Inhibitors with Broad Therapeutic Potential
Authors: Radnai Laszlo , Young Erica J. , Kikuti Carlos , Hafenbreidel Madalyn , Stremel Rebecca F. , Lin Li , Toth Katalin , Pasetto Paolo , Jin Xiaomin , Patel Aagam , Conlon Michael , Briggs Sherri , Heidsieck Leïla , Sweeney H. Lee , Sellers James , Krieger-Burke Teresa , Martin William H. , Sisco Jay , Young Steven , Pearson Paul , Rumbaugh Gavin , Araldi Gian Luca , Duddy Steven K. , Cameron Michael D. , Surman Matthew , Houdusse Anne , Griffin Patrick R. , Kamenecka Theodore M. , Miller Courtney A. 
Published: October 2024

ABSTRACTNon-muscle myosin II (NMII), a molecular motor that regulates critical processes such as cytokinesis and neuronal synaptic plasticity, has substantial therapeutic potential. However, translating this potential toin vivouse has been hampered by the lack of selective tools. The most prototypical non-selective inhibitor, blebbistatin inactivates both NMII and cardiac myosin II (CMII), a key regulator of heart function. Using rational drug design, we developed a series of NMII inhibitors that improve tolerability by selectively targeting NMII over CMII, including MT-228, which has excellent properties such as high brain penetration and efficacy in preclinical models of stimulant use disorder, which has no current FDA-approved therapies. The structure of MT-228 bound to myosin II provides insight into its 17-fold selectivity for NMII over CMII. MT-228’s broad therapeutic window opens the door to new disease treatments and provides valuable tools for the scientific community, along with promising leads for future medication development.HighlightsResearch suggests numerous indications, from axon regeneration and cancer, would benefit from a small molecule inhibitor of non-muscle myosin II, a molecular motor that regulates the actin cytoskeleton.Current chemical probe options are very limited and lack sufficient safety forin vivostudies, which we show is primarily due to potent inhibition of cardiac myosin II.Rational design that focused on improving target selectivity over the pan-myosin II inhibitor, blebbistatin, led to the identification of MT-228, a small molecule inhibitor with a wide therapeutic window.High-resolution structure of MT-228 bound to myosin II reveals that selectivity results from a different positioning compared to blebbistatin and an important sequence difference between cardiac and non-muscle myosin II in the inhibitor binding pocket.A single administration of MT-228 shows long-lasting efficacy in animal models of stimulant use disorder, a current unmet and rapidly escalating need with no FDA-approved treatments.
Local cortical inhibitory subnetworks are shaped by pyramidal neuron progenitor type
Authors: Gothard Gemma , Mahfooz Kashif , Newey Sarah E. , Tinelli Sasha , Buchan Matthew J. , Burman Richard J. , Akerman Colin J. 
Published: October 2024

AbstractThe degree to which cortical neurons share inhibitory synaptic input determines their co-activity within a network. However, the principles by which inhibition is shared between neurons are not known. Here we combinein uterolabeling within vivotwo-photon targeted patch-clamp recordings in mature cortex to reveal that a layer 2/3 (L2/3) pyramidal neuron’s local inhibitory input reflects the embryonic progenitor type from which the neuron is born. In contrast to neighboring neurons, pyramidal neurons derived from intermediate progenitors receive synaptic inhibition that is weakly coupled to local network activity. The underlying mechanisms do not depend upon the amount of inhibitory input received from different interneuron subclasses. Rather, progenitor type defines how much inhibitory input a neuron shares with its neighbors, which is reflected in how individual interneurons target pyramidal neurons according to progenitor type. These findings reveal new significance for progenitor diversity and identify ontogenetic origins of fine-scale inhibitory cortical subnetworks.
GenomicGapID: Leveraging Spatial Distribution of Conserved Genomic Sites for Broad-Spectrum Microbial Identification
Authors: Asthana Vishwaratn , Bugga Pallavi , Smith Clara Elaine , Wellman Catherine , Dwight Zachary , Ranjan Piyush , Nieves Erika Martínez , Dickson Robert P. , VanEpps J. Scott 
Published: October 2024

ABSTRACTBacterial detection and identification methods can be broadly classified as either untargeted with expansive taxonomic coverage or targeted with narrow taxonomic focus. Untargeted approaches, such as culture and sequencing, are often time-consuming and/or costly, whereas targeted methods, such as PCR, can offer faster and more cost-effective results but require a priori knowledge of the likely pathogen to select the appropriate assay. GenomicGapID, a novel approach that leverages the spatial distribution of conserved genetic regions across microbial genomes, represents a significant advancement in the field of microbial identification. This technique has the potential to provide the taxonomic breadth of culture and sequencing, while maintaining the speed, simplicity, and cost-effectiveness of PCR. By leveraging the conservation and relative positioning of highly conserved coding regions across different species, GenomicGapID enables the development of universal primer sets that amplify the non-conserved gaps between these regions. This creates a unique electrophoretic signature that facilitates rapid and accurate target agnostic microbial identification. In this study, we apply the principles of GenomicGapID to the critical task of identifying clinical pathogens. We focus on expanding the coverage of a previously developed universal bacterial identification system, which initially targeted the 16s-23s internal transcribed spacer (ITS) region and was capable of discerning 45 pathogens. To enhance this system, we assembled a comprehensive database of 189 clinically relevant bacterial species. We then identified conserved primer binding sites that produce unique amplicon size signatures for each species. While we found that the use of amplicon size signatures alone would require an impractical number of universal primer sets, we demonstrate that this challenge can be effectively mitigated through concurrent melt analysis. Ultimately, we show that just three universal primer sets, guided by the GenomicGapID framework, are sufficient to cover 189 clinical bacterial pathogens, representing a majority of microbes identified in positive cultures in a clinical microbiology setting, with experimental validation of a subset of these pathogens. This study not only enhances the existing universal bacterial identification system but also establishes GenomicGapID as a versatile and powerful tool in microbial diagnostics and beyond with the potential to open new areas of investigation in genomics, with significant implications for molecular biology, clinical practice, and public health.
Small heat shock proteins (sHSPs) identified in nodules of tropical woody legumes
Authors: Flynn Cara G. , Fakih Rayan , Gehring Kalle , Soper Fiona M. 
Published: October 2024

AbstractBiological nitrogen fixation (BNF) is a primary input of nitrogen to natural and agricultural systems globally. BNF is a temperature-dependant enzymatic process and can be conducted by microbes (includingRhizobia) hosted symbiotically in root nodules of some plants. Heat shock proteins (Hsps) have been implicated in the process of acquiring thermotolerance or acclimating to elevated temperature, as they play a vital role in maintaining cell integrity and homeostasis during heat stress. Although the BNF response to temperature may crucially impact future ecosystem productivity in the face of global climate change, little is known about Hsp expression in nodules of N-fixing non-agricultural species, such as tropical N-fixing trees in theFabaceaefamily. This project aimed to characterize small (15-20 kDa) Hsp (sHsp) expression in nodule tissue to examine the biochemical mechanisms of heat response in these tissues. To first identify Hsps in nodule tissues,Vachellia farnesianaandAcacia confusanodules were excised, heat shock was induced, and protein content was isolated via chemical treatment before separation of protein species and analysis with SDS-PAGE. Two polyacrylamide gels yielded bands in the 15-20 kDa region that displayed differential Coomassie staining, which were sent for further characterization by HPLC-MS analysis for protein sequencing. Ten rhizobial sHsps were detected in these samples in addition to sevenAcaciasHsps when compared independently to reference rhizobial and plant proteome databases. In an attempt quantify relative expression of Hsps in nodule and root tissue, we performed western blot experiments using Anti-Hsp20 antibodies raised against human and mouse Hsp proteins, with anti-beta actin loading control. While nonuniform beta-actin expression across tissue types (A. confusanodules versus root control) prevented quantitative analysis, the experiments validated that Hsp20s are expressed inAcacianodules as well as in root tissue. These experiments provide a foundation for future studies to determine variation in responses to key stressors predicted to increase with global climate change and help determine the implications of warming across the tropics and beyond. Proteomics data are available via ProteomeXchange with identifier PXD055599.
Vocal communication is seasonal in social groups of wild, free-living house mice
Authors: Jourjine N , Goedecker C , König B , Lindholm AK 
Published: October 2024

AbstractHouse mice (Mus musculus domesticus) are among the most widely studied laboratory models of mammalian social behavior, yet we know relatively little about the ecology of their behaviors in natural environments. Here, we address this gap using radiotelemetry to track social interactions in a population of wild mice over 10 years, from 2013 to 2023, and interpret these interactions in the context of passive acoustic monitoring data collected from August 2022 until November 2023. Using deep convolutional neural networks, we identify 1.3 million individual vocalizations and align them in time with continuously collected telemetry data recording social interactions between individually identifiable mice. We find that vocalization is seasonal and correlated with long-term dynamics in features of social groups. In addition, we find that vocalization is closely associated in time with entrances to and exits from those groups, occurs most often in the presence of pups, and is correlated with how much time pairs of mice spend together. This work identifies seasonal patterns in the vocalizations of wild mice and lays a foundation to investigate the social role of acoustic communication in wild populations of a classic laboratory model organism.
Metastatic organotropism in small cell lung cancer
Authors: Krishnamurthy Manan , Dhall Anjali , Sahoo Sarthak , Schultz Christopher W. , Baird Michelle A. , Desai Parth , Odell Jacob , Takahashi Nobuyuki , Nirula Michael , Zhuang Sophie , Huang Yue , Schroeder Brett , Zhang Yang , Thomas Maria Sebastian , Redon Christophe , Robinson Christina , Thang Lai , Ileva Lilia , Patel Nimit L. , Kalen Joseph D. , Varlet Alice-Anaïs , Zuela-Sopilniak Noam , Jha Ankita , Wangsa Darawalee , Butcher Donna , Morgan Tamara , Afzal Alyah N. , Chari Raj , Baktiar Karim , Kumar Suresh , Pongor Lorinc , Difilippantonio Simone , Aladjem Mirit I. , Pommier Yves , Jolly Mohit Kumar , Lammerding Jan , Sharma Ajit Kumar , Thomas Anish 
Published: October 2024

AbstractMetastasis is the leading cause of cancer-related deaths, yet its regulatory mechanisms are not fully understood. Small-cell lung cancer (SCLC) is the most metastatic form of lung cancer, with most patients presenting with widespread disease, making it an ideal model for studying metastasis. However, the lack of suitable preclinical models has limited such studies. We utilized rapid autopsy-derived tumors to develop xenograft models that mimic key features of SCLC, including histopathology, rapid and widespread development of metastasis to the liver, brain, adrenal, bone marrow, and kidneys within weeks, and response to chemotherapy. By integrating in vivo lineage selection with comprehensive bulk and single cell multiomic profiling of transcriptomes and chromatin accessibility, we identified critical cellular programs driving metastatic organotropism to the liver and brain, the most common sites of SCLC metastasis. Our findings reveal the key role of nuclear-cytoskeletal interactions in SCLC liver metastasis. Specifically, the loss of the nuclear envelope protein lamin A/C, encoded by theLMNAgene, increased nuclear deformability and significantly increased the incidence of liver metastasis. Human liver metastases exhibited reducedLMNAexpression compared to other metastatic sites, correlating with poorer patient outcomes and increased mortality. This study introduces novel preclinical models for SCLC metastasis and highlights pathways critical for organ-specific metastasis, offering new avenues for the development of targeted therapies to prevent or treat metastatic disease.
Reduced methane emissions in transgenic rice genotypes are associated with altered rhizosphere microbial hydrogen cycling
Authors: Shi Ling-Dong , Ercoli Maria Florencia , Kim Jack , de Araujo Junior Artur Teixeira , Soni Subah , Weitz Tracy Satomi , Shigenaga Alexandra M. , Dukovski Ilija , Sachdeva Rohan , Turumtay Halbay , Louie Katherine B. , Bowen Benjamin P. , Scheller Henrik V. , Segrè Daniel , Northen Trent R. , Ronald Pamela C. , Banfield Jillian F. 
Published: October 2024

AbstractRice paddies contribute substantially to atmospheric methane (CH4) and these emissions are expected to increase as the need to feed the human population grows. Here, we show that two independent rice genotypes overexpressing genes forPLANT PEPTIDES CONTAINING SULFATED TYROSINE(PSY) reduced cumulative CH4emissions by 38% (PSY1) and 58% (PSY2) over the growth period compared with controls. Genome-resolved metatranscriptomic data from rhizosphere soils reveal lower ratios of gene activities for CH4production versus consumption, decrease in activity of H2-producing genes, and increase in bacterial H2oxidation pathways in the PSY genotypes. Metabolic modeling using metagenomic and metabolomic data predicts elevated levels of H2oxidation and suppressed H2production in the PSY rhizosphere. The H2-oxidizing bacteria have more genes for utilization of gluconeogenic acids than H2-producing counterparts, and their activities were likely stimulated by the observed enrichment of gluconeogenic acids (mostly amino acids) in PSY root exudates. Together these results suggest that decreased CH4emission is due to the reduction of H2available for hydrogenotrophic methanogenesis. The combination of rice phenotypic characterization, microbiome multi-omic analysis, and metabolic modeling described here provides a powerful strategy to discover the mechanisms by which specific plant genotypes can alter biogeochemical cycles to reduce CH4emissions.