bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genome-wide analysis of the class III peroxidase gene family inPhyscomitrium patensand a search for clues to ancient class III peroxidase functions
Authors: Aparato Vincent P. M. , Rabbi Fazle , Madarash Taylor , Brisbourne Wyllie A. , Barker Elizabeth I. , Suh Dae-Yeon 
Published: January 2024

AbstractPlant class III peroxidases (PRX) catalyze generation of reactive oxygen species and oxidation of various compounds including lignin precursors. PRXs function in cell wall metabolism, defense and stress responses. However, gene redundancy and catalytic versatility have impeded detailed functional characterization ofPRXgenes. The genome of the model mossPhyscomitrium patensharbors a relatively small number (49) ofPRXgenes. Conserved architecture of four exons and three ‘001’ introns, found in some algalPRXgenes and in thePpPRXfamily, suggests that this architecture predated divergence of the green algal and land plant lineages. ThePpPRXfamily expanded mainly through whole-genome duplications. All duplicated pairs but one were under purifying selection and generally exhibited similar expression profiles. An expanded phylogenetic tree revealed a conserved land plant-wide clade that contained PRXs implicated in stress responses in non-lignifying cells, providing a clue to ancient functions of land plant PRXs. Functional clustering was not observed, suggesting convergent evolution of specific PRX functions (e.g., lignification) in different plant lineages. With its small complement of PRXs, P. patensmay be useful for functional characterization of land plant PRXs. Several PpPRXs were proposed for further study, includingPpPRX34andPpPRX39in the ancient land plant-wide clade.
k-nonical space: sketching with reverse complements
Authors: Marçais Guillaume , Elder C.S. , Kingsford Carl 
Published: January 2024

AbstractSequences equivalent to their reverse complements (i.e., double-stranded DNA) have no analogue in text analysis and non-biological string algorithms. Despite this striking difference, algorithms designed for computational biology (e.g., sketching algorithms) are designed and tested in the same way as classical string algorithms. Then, as a post-processing step, these algorithms are adapted to work with genomic sequences by folding ak-mer and its reverse complement into a single sequence: the canonical representation (k-nonical space). The effect of using the canonical representation with sketching methods is understudied and not understood. As a first step, we use context-free sketching methods to illustrate the potentially detrimental effects of using canonicalk-mers with string algorithms not designed to accommodate for them. In particular, we show that large stretches of the genome (“sketching deserts”) are undersampled or entirely skipped by context-free sketching methods, effectively making these genomic regions invisible to subsequent algorithms using these sketches. We provide empirical data showing these effects and develop a theoretical framework explaining the appearance of sketching deserts. Finally, we propose two schemes to accommodate for these effects: (1) a new procedure that adapts existing sketching methods tok-nonical space and (2) an optimization procedure to directly design new sketching methods fork-nonical space.The code used in this analysis is freely available athttps://github.com/Kingsford-Group/mdsscope.
Predicting microbial genome-scale metabolic networks directly from 16S rRNA gene sequences
Authors: Øyås Ove , Kobel Carl M. , Vik Jon Olav , Pope Phillip B. 
Published: January 2024

AbstractGenome-scale metabolic models are key biotechnology tools that can predict metabolic capabilities and growth for an organism. In particular, these models have become indispensable for metabolic analysis of microbial species and communities such as the gut microbiomes of humans and other animals. Accurate microbial models can be built automatically from genomes, but many microbes have only been observed through sequencing of marker genes such as 16S rRNA and thus remain inaccessible to genome-scale modeling. To extend the scope of genome-scale metabolic models to microbes that lack genomic information, we trained an artificial neural network to build microbial models from numeric representations of 16S rRNA gene sequences. Specifically, we built models and extracted 16S rRNA gene sequences from more than 15,000 reference and representative microbial genomes, computed multiple sequence alignments and large language model embeddings for the 16S rRNA gene sequences, and trained the neural network to predict metabolic reaction probabilities from sequences, alignments, or embeddings. Training was fast on a single graphics processing unit and trained networks predicted reaction probabilities accurately for unseen archaeal and bacterial sequences and species. This makes it possible to reconstruct microbial genome-scale metabolic networks from any 16S rRNA gene sequence and enables simulation of metabolism and growth for all observed microbial life.
Paraoxonase-like APMAP maintains endoplasmic reticulum-associated lipid and lipoprotein homeostasis
Authors: Paul Blessy , Merta Holly , Ugrankar-Banerjee Rupali , Hensley Monica , Tran Son , Dias do Vale Goncalo , McDonald Jeffrey G , Farber Steven A. , Henne W Mike 
Published: January 2024

SummaryOxidative stress perturbs lipid homeostasis and contributes to metabolic diseases. Though ignored compared to mitochondrial oxidation, the endoplasmic reticulum (ER) generates reactive oxygen species requiring antioxidant quality control. Using multi-organismal profiling featuringDrosophila, zebrafish, and mammalian cells, here we characterize the paraoxonase-like APMAP as an ER-localized protein that promotes redox and lipid homeostasis and lipoprotein maturation. APMAP-depleted mammalian cells exhibit defective ER morphology, elevated ER and oxidative stress, lipid droplet accumulation, and perturbed ApoB-lipoprotein homeostasis. Critically, APMAP loss is rescued with chemical antioxidant NAC. Organismal APMAP depletion inDrosophilaperturbs fat and lipoprotein homeostasis, and zebrafish display increased vascular ApoB-containing lipoproteins, particles that are atherogenic in mammals. Lipidomics reveals altered polyunsaturated phospholipids and increased ceramides upon APMAP loss, which perturbs ApoB-lipoprotein maturation. These ApoB-associated defects are rescued by inhibiting ceramide synthesis. Collectively, we propose APMAP is an ER-localized antioxidant that promotes lipid and lipoprotein homeostasis.Key findings summaryAPMAP localizes primarily to the ER network in human cells andDrosophilafat body tissue, and is a type II integral membrane proteinLoss of APMAP orDrosophilaAPMAP (dAPMAP) causes ER membrane expansion, elevates CHOP-associated ER stress, promotes LD accumulation, and alters ApoB-lipoprotein secretionAPMAP-depleted cells and dAPMAP-depletedDrosophilafat tissue exhibit defective redox homeostasis; phenotypes associated with APMAP loss are rescued by antioxidant NACZebrafish-based LipoGlo reporter reveals that loss ofapmapin zebrafish causes increased vascular ApoB-containing lipoproteinsLipidomic profiling indicates that APMAP loss reduces PUFA-phospholipids and elevates intracellular ceramides, which perturbs ApoB maturation
Emergent collective alignment gives competitive advantage to longer cells during range expansion
Authors: van den Berg Nathan , Thijssen Kristian , Nguyen Thu Trang , Sarlet Adrien , Cordero Mireia , Vázquez Alba García , Mitarai Namiko , Doostmohammadi Amin , Jauffred Liselotte 
Published: January 2024

ABSTRACTBacteria’s competition for nutrients and territory drives biofilm evolution (1–4). The factors determining the outcome of competition among diverse bacterial species have a broad impact on a wide range of pathological (5), environmental (6), and microbiome interactions (7). While motility-related traits (8–11) and specific molecular mechanisms (12, 13) have been identified as potential winning attributes in bacteria, a shared and universally conserved feature determining competition remains elusive. Here, we demonstrate that a simple morphological feature of individual bacteria, cell aspect ratio, provides a winning trait for the population. Using growth-based range expansion experiments (14), we show that relatively longer bacteria robustly conquer the expanding front, even when initially in minority. Using an agent-based model of dividing bacteria, we reveal that the takeover mechanism is their emergent collective alignment: groups of locally aligned bacteria form “nematic arms” bridging the central region of the colony to the expanding front. Once at the front, bacteria align parallel to it and block the access of shorter bacteria to nutrients and space. We confirm this observation with single-cell experiments and further generalise our findings by introducing a generic continuum model of alignment-dominated competition, explaining both experimental and cell-based model observations. Moreover, we extend our predictions to spherical range expansions (15) and confirm the competitive advantage of being longer, even though the effect is less pronounced than in surface-attached colonies. Our results uncover a simple, yet hitherto overlooked, mechanical mechanism determining the outcome of bacterial competition, which is potentially ubiquitous among various bacteria. With the current advances in genetic engineering, varying aspect ratios can work as a simple tunable mechanism for the on-demand setting of the outcome of bacterial competitions with widespread implications for biofilm control.
An accessible digital imaging workflow for multiplexed quantitative analysis of adult eye phenotypes inDrosophila melanogaster
Authors: Pipkin Heidi J. J. , Lindsay Hunter L. , Smiley Adam T. , Jurmu Jack D. , Arsham Andrew M 
Published: January 2024

AbstractThe compound eye ofDrosophila melanogasterhas long been a model for studying genetics, development, neurodegeneration, and heterochromatin. Imaging and morphometry of adultDrosophilaand other insects is hampered by the low throughput, narrow focal plane, and small image sensors typical of stereomicroscope cameras. When data collection is distributed among many individuals or extended time periods, these limitations are compounded by inter-operator variability in lighting, sample positioning, focus, and post-acquisition processing. To address these limitations we developed a method for multiplexed quantitative analysis of adultDrosophila melanogasterphenotypes. Efficient data collection and analysis of up to 60 adult flies in a single image with standardized conditions eliminates inter-operator variability and enables precise quantitative comparison of morphology. Semi-automated data analysis using ImageJ and R reduces image manipulations, facilitates reproducibility, and supports emerging automated segmentation methods, as well as a wide range of graphical and statistical tools. These methods also serve as a low-cost hands-on introduction to imaging, data visualization, and statistical analysis for students and trainees.
Human vascular organoids with a mosaicAKT1mutation recapitulate Proteus syndrome
Authors: He Siyu , Zhu Yuefei , Chauhan Shradha , Tavakol Daniel Naveed , Lee Jong Ha , Berris Rayna Batya-Leia , Xu Cong , Lee Jounghyun H. , Lee Caleb , Cai Sarah , McElroy Shannon , Vunjak-Novakovic Gordana , Tomer Raju , Azizi Elham , Xu Bin , Lao Yeh-Hsing , Leong Kam W. 
Published: January 2024

AbstractVascular malformation, a key clinical phenotype of Proteus syndrome, lacks effective models for pathophysiological study and drug development due to limited patient sample access. To bridge this gap, we built a human vascular organoid model replicating Proteus syndrome’s vasculature. Using CRISPR/Cas9 genome editing and gene overexpression, we created induced pluripotent stem cells (iPSCs) embodying the Proteus syndrome-specific AKTE17Kpoint mutation for organoid generation. Our findings revealed that AKT overactivation in these organoids resulted in smaller sizes yet increased vascular connectivity, although with less stable connections. This could be due to the significant vasculogenesis induced by AKT overactivation. This phenomenon likely stems from boosted vasculogenesis triggered by AKT overactivation, leading to increased vascular sprouting. Additionally, a notable increase in dysfunctional PDGFRβ + mural cells, impaired in matrix secretion, was observed in these AKT-overactivated organoids. The application of AKT inhibitors (ARQ092, AZD5363, or GDC0068) reversed the vascular malformations; the inhibitors’ effectiveness was directly linked to reduced connectivity in the organoids. In summary, our study introduces an innovative in vitro model combining organoid technology and gene editing to explore vascular pathophysiology in Proteus syndrome. This model not only simulates Proteus syndrome vasculature but also holds potential for mimicking vasculatures of other genetically driven diseases. It represents an advance in drug development for rare diseases, historically plagued by slow progress.
Centromere landscapes resolved from hundreds of human genomes
Authors: Gao Shenghan , Zhang Yimeng , Bush Stephen J. , Wang Bo , Yang Xiaofei , Ye Kai 
Published: January 2024

AbstractHigh-fidelity (HiFi) sequencing has facilitated the assembly and analysis of the most repetitive region of the genome, the centromere. Nevertheless, our current understanding of human centromeres draws from a relatively small number of telomere-to-telomere assemblies, and so has not yet captured its full diversity. In this study, we investigated the genomic diversity of human centromere higher order repeats (HORs) using both HiFi reads and haplotype-resolved assemblies from hundreds of samples drawn from ongoing pangenome-sequencing projects and reprocessed using a novel HOR annotation pipeline, HiCAT-human. We use this wealth of data to provide a global survey of the centromeric HOR landscape, in particular finding that 23 HORs exhibited significant copy number variability between populations. We detected three centromere genotypes with imbalance population frequencies on each of chromosome 5, 8 and 17. An inter-assembly comparison of HOR loci further revealed that while HOR array structures are diverse, they nevertheless tend to form a number of specific landscapes, each exhibiting different levels of HOR subunit expansion and possibly reflecting a cyclical evolutionary transition from homogeneous to nested structures and back.
GRK2-mediated AKT activation controls cell cycle progression and G2-checkpoint in a p53-dependent manner
Authors: Rivas Verónica , González-Muñoz Teresa , Albitre Ángela , Lafarga Vanesa , Mayor Federico , Penela Petronila 
Published: January 2024

ABSTRACTCell cycle checkpoints, activated by stressful events, halt the cell cycle progression, and prevent the transmission of damaged DNA. These checkpoints prompt cell repair but also trigger cell death if damage persists. Decision-making between these responses is multifactorial and context-dependent, with the tumor suppressor p53 playing a central role. In many tumor cells, p53 alterations lead to G1/S checkpoint loss, rendering cell viability dependent on the strength of the G2 checkpoint through mechanisms not fully characterized. Cells with a strong pro-survival drive can evade cell death despite substantial DNA lesions. Deciphering the integration of survival pathways with p53-dependent and -independent mechanisms governing the G2/M transition is crucial for understanding G2 arrest functionality and predicting tumor cell response to chemotherapy. The serine/threonine kinase GRK2 emerges as a signaling node in cell cycle modulation. In cycling cells, but not in G2 checkpoint-arrested cells, GRK2 protein levels decline during G2/M transition through a process triggered by CDK2-dependent phosphorylation of GRK2 at the S670 residue and Mdm2 ubiquitination. We report now that this downmodulation in G2 prevents the unscheduled activation of the PI3K/AKT pathway, allowing cells to progress into mitosis. Conversely, higher GRK2 levels lead to tyrosine phosphorylation by the kinase c-Abl, promoting the direct association of GRK2 with the p85 regulatory subunit of PI3K and AKT activation in a GRK2 catalytic-independent manner. Hyperactivation of AKT is conditioned by p53’s scaffolding function, triggering FOXO3a phosphorylation, impaired cyclin B1 accumulation, and CDK1 activation, causing a G2/M transition delay. Upon G2 checkpoint activation, GRK2 potentiates early arrest independently of p53 through AKT activation. However, its ability to overcome the G2 checkpoint in viable conditions depends on p53. Our results suggest that integrating the GRK2/PI3K/AKT axis with non-canonical functions of p53 might confer a survival advantage to tumor cells.
Large-scale characterization of drug mechanism of action using proteome-wide thermal shift assays
Authors: Van Vranken Jonathan G. , Li Jiaming , Mintseris Julian , Wei Ting-Yu , Sniezek Catherine M. , Gadzuk-Shea Meagan , Gygi Steven P. , Schweppe Devin K. 
Published: January 2024

AbstractIn response to an ever-increasing demand of new small molecules therapeutics, numerous chemical and genetic tools have been developed to interrogate compound mechanism of action. Owing to its ability to approximate compound-dependent changes in thermal stability, the proteome-wide thermal shift assay has emerged as a powerful tool in this arsenal. The most recent iterations have drastically improved the overall efficiency of these assays, providing an opportunity to screen compounds at a previously unprecedented rate. Taking advantage of this advance, we quantified more than one million thermal stability measurements in response to multiple classes of therapeutic and tool compounds (96 compounds in living cells and 70 compounds in lysates). When interrogating the dataset as a whole, approximately 80% of compounds (with quantifiable targets) caused a significant change in the thermal stability of an annotated target. There was also a wealth of evidence portending off-target engagement despite the extensive use of the compounds in the laboratory and/or clinic. Finally, the combined application of cell-and lysate-based assays, aided in the classification of primary (direct ligand binding) and secondary (indirect) changes in thermal stability. Overall, this study highlights the value of these assays in the drug development process by affording an unbiased and reliable assessment of compound mechanism of action.
Growing a cystic fibrosis-relevant polymicrobial biofilm to probe community phenotypes
Authors: Poirier Sarah , Jean-Pierre Fabrice 
Published: January 2024

ABSTRACTMostin vitromodels lack the capacity to fully probe bacterial phenotypes emerging from the complex interactions observed in real-life environments. This is particularly true in the context of hard-to-treat chronic and polymicrobial biofilm-based infections detected in the airways of persons with cystic fibrosis (pwCF), a multiorgan genetic disease. While multiple microbiome studies have defined the microbial compositions detected in the airway of pwCF, noin vitromodels thus far had fully integrated critical cystic fibrosis (CF)-relevant lung features. Therefore, a significant knowledge-gap in our capacity to investigate the mechanisms driving the pathogenesis of mixed species CF lung infections remained. To tackle this challenge, we have built a four-species microbial community model includingPseudomonas aeruginosa, Staphylococcus aureus, Streptococcus sanguinis, andPrevotella melaninogenicagrown in CF-like conditions. Through the utilization of this system, clinically relevant phenotypes such as antimicrobial (Abx) recalcitrance of several pathogens were observed and explored at the molecular level. The usefulness of thisin vitromodel resides in its standardized workflow that can facilitate the study of interspecies interactions in the context of chronic CF lung infections.SUMMARYIn this protocol, we describe a cystic fibrosis (CF)-lung relevant four-species polymicrobial biofilm model that can be used to explore the impact of bacterial interspecies interactions.
Characterization of antimalarial activity of artemisinin-based hybrid drugs
Authors: Quadros Helenita Costa , Herrmann Lars , Manaranche Jeanne , Paloque Lucie , Borges-Silva Mariana C. , Dziwornu Godwin Akpeko , D’Alessandro Sarah , Chibale Kelly , Basilico Nicoletta , Benoit-Vical Françoise , Tsogoeva Svetlana B. , Moreira Diogo Rodrigo M. 
Published: January 2024

ABSTRACTIn response to the spread of artemisinin (ART) resistance, ART-based hybrid drugs were developed and their activity profile was characterized against drug-sensitive and drug-resistantPlasmodium falciparumparasites. Two hybrids were found to display parasite growth reduction, stage-specificity, speed of activity, additivity of activity in drug combinations, and stability in hepatic microsomes of similar levels to those displayed by dihydroartemisinin (DHA). Conversely, the rate of chemical homolysis of the peroxide bonds is slower in the hybrids than in DHA. From a mechanistic perspective, heme plays a central role in the chemical homolysis of peroxide and in inhibiting heme detoxification and disrupting parasite heme redox homeostasis. The hybrid exhibiting slow homolysis of peroxide bonds was more potent in reducing the viability of ART-resistant parasites in a ring-stage survival assay than the hybrid exhibiting fast homolysis. However, both hybrids showed some limited activity against ART-induced quiescent parasites in the quiescent-stage survival assay. Our findings are consistent with previous results showing that slow homolysis of peroxide-containing drugs may retain activity against proliferating ART-resistant parasites. However, our data suggest that this property does not overcome the limited activity of peroxides in killing non-proliferating parasites in a quiescent state.GRAPHICAL ABSTRACTHepatic and cell-host-mediated metabolism are responsible for short plasma half-lives of antimalarial artemisinins (ARTs), illustrated here by dihydroartemisinin (DHA). ART-based hybrid drugs that overcome rapid degradation can facilitate activity against ART-resistant parasites, as illustrated by hybrid1.
Production of OSU G5P[7] Porcine Rotavirus Expressing a Fluorescent Reporter by Reverse Genetics
Authors: Snyder Anthony J. , Agbemabiese Chantal A. , Patton John T. 
Published: January 2024

ABSTRACTRotaviruses are a significant cause of severe, potentially life-threatening gastroenteritis in the young of many economically important animals. Although vaccines against porcine rotavirus exist, both live oral and inactivated, their effectiveness in preventing gastroenteritis is less than ideal. Thus, a need remains for the development of new generations of porcine rotavirus vaccines. The Ohio State University (OSU) rotavirus strain represents aRotavirus Aspecies with a G5P[7] genotype, the genotype most frequently associated with rotavirus disease in piglets. Using complete genome sequences that were determined by Nanopore sequencing, we developed a robust reverse genetics system enabling recovery of recombinant (r)OSU rotavirus. Although rOSU grew to high titers (∼107plaque-forming units/ml), its growth kinetics were modestly decreased in comparison to laboratory-adapted OSU virus. The reverse genetics system was used to generate rOSU rotavirus that served as an expression vector for foreign protein. Specifically, by engineering a fused NSP3-2A-UnaG open reading frame into the segment 7 RNA, we produced genetically stable rOSU that expressed the fluorescent UnaG protein as a functional separate product. Together, these findings raise the possibility of producing improved live oral porcine rotavirus vaccines through reverse genetics-based modification or combination porcine rotavirus vaccines that can express neutralizing antigens of other porcine enteric diseases.