bluuebunny/crossref_metadata_2025_split huggingface dataset:
The oncogenic CCDC6-RET fusion product is a dual ATP and ADP-dependent kinase that functions viacis-phosphorylation
Authors: Martín-Hurtado Ana , Contreras Julia , Sánchez-Wandelmer Jana , Zarzuela Eduardo , Muñoz Inés G. , Muñoz Javier , Isasa Marta , Plaza-Menacho Iván 
Published: January 2025

SummaryGene fusions products involving protein kinases are known drivers in human cancers and actionable targets for personalized therapy, yet the structural and molecular determinants that control their function are largely unexplored. Here we show that a CCDC6-RET fusion product, a driver and therapeutic target in lung and thyroid cancers, is a highly active dimeric kinase in solution. Time-resolved mass spectrometry analysis together with a robust biochemical and biophysical characterization reveal that CCDC6-RET functions as a dual ATP- and ADP-dependent kinase able to bind both nucleotides and uses them as phosphoryl donors. We also identify a crosstalk between the c-terminal and the activation segments, uncovering a mutually exclusive dependency by the former on activation loop phospho-sites controlling both the processing and the catalytic activity of the fusion protein. Furthermore, we generated a 3D-assembly of a CCDC6-RET homodimer combining electron microscopy (EM) single particle, small-angle X-ray scattering (SAXS) and in silico structural analyses. Our structural model together with cross-linking mass spectrometry data demonstrated that CCDC6-RET forms a face-to-facetrans-inhibited dimer in the apo state characterized by intermolecular-crosslinked activation segments. Upon nucleotide binding and catalytic domains reorientation, fast activation loop phosphorylation is driven by a mechanismin cis. Our work uncover for the first time the molecular and structural determinants that controls CCDC6-RET function and provides a solid framework to study the structure and function of other RET fusion products.
Creation and Validation of a Proteome-Wide Yeast Library for Protein Detection and Analysis
Authors: Baruch Din , Tsirkas Ioannis , Sass Ehud , Dubreuil Benjamin , Asraf Yeynit , Aharoni Amir , Schuldiner Maya , Klein Ofir 
Published: January 2025

AbstractA significant challenge in cell biology is to uncover the function of uncharacterized proteins. Surprisingly a quarter of the proteome is still poorly understood even in the most well studied model organisms. Systematic methodologies, including the use of tagged protein collections, have emerged as a powerful approach to address this gap. Despite the availability of proteome- wide collections featuring various fused proteins, the impact of tag size on protein function highlighted the need for using minimally disruptive tags for functional genomic studies. To rise to this challenge, we have created a proteome-wide collection of yeast strains in which proteins are N-terminally tagged with the Hemagglutinin (HA) epitope. The library leverages the compact size of the HA tag to minimize drawbacks associated with larger tags while enabling efficient functional analysis. We showcase the potential uses of our library for systematically evaluating protein size, abundance and localization using anin vivolabeling approach. Our characterization underscores the potential utility of a proteome-wide HA-tagged library in revealing novel aspects of cell biology, providing an additional powerful tool for functional genomics.
Preparation of stagedCaenorhabditis elegansembryos using size filtration
Authors: Jhaveri Nikita S. , Mastronardo Maya K. , Collins J.B. , Andersen Erik C. 
Published: January 2025

AbstractThe free-living nematodeCaenorhabditis eleganshas been routinely used to study gene functions, gene interactions, and conserved signaling pathways. Most experiments require that the animals are synchronized to be at the same specific developmental stage. Bleach synchronization is traditionally used to obtain a population of staged embryos, but the method can have harmful effects on the embryos. The physical separation of differently sized animals is preferred but often difficult to perform because some developmental stages are the same sizes as others. Microfluidic device filters have been used as alternatives, but they are expensive and require customization to scale up the preparation of staged animals. Here, we present a protocol for the synchronization of embryos using mesh filters. Using filtration, we obtained a higher yield of embryos per plate than using standard bleach synchronization protocol and at a scale beyond microfluidic devices. Importantly, filtration does not affect downstream larval development assays as much as bleach synchronization does. In conclusion, we have exploited the differences in the sizes ofC. elegansdevelopmental stages to isolate embryo cultures suitable for use in high-throughput assays.
Functional Characterization of a Novel Long Non-Coding RNA inLeishmania braziliensisIdentified Through Computational Screening for Conserved RNA Structures
Authors: Espada Caroline R. , Anthon Christian , Magalhães Rubens D.M. , Quilles Junior José Carlos , Teles Natalia M.M. , Pais Fabiano , Orsine Lissur A. , de Almeida Letícia , Defina Tânia P. A. , Dowle Adam , Gorodkin Jan , Walrad Pegine B. , Cruz Angela K. 
Published: January 2025

AbstractLeishmaniaparasites alternate between hosts, facing environmental changes that demand rapid gene expression adaptation. Lacking canonical RNA polymerase II promoters, transcription in these eukaryotes is polycistronic, with gene regulation occurring post-transcriptionally. Although non-coding RNAs (ncRNAs) have been identified inLeishmaniatranscriptomes, their functions remain unclear. Recognizing RNA structure’s importance, we performed a genome-wide alignment ofL. braziliensisand related species, identifying conserved RNA structures, 38 of which overlap with known ncRNAs. One such ncRNA,lncRNA45, was functionally characterized. Using a knockout cell line, we demonstrated thatlncRNA45is crucial for parasite fitness. Reintroducing the wild-typelncRNA45restored fitness, while a version with a single nucleotide substitution in the structured region did not. This mutation also altered RNA-protein interactions. These findings suggest thatlncRNA45’s regulatory role and protein interactions rely on its secondary structure. This study highlights the significance of structured lncRNAs inLeishmaniabiology and their potential as therapeutic targets. Further research into these ncRNAs could uncover new parasite regulation mechanisms and inspire novel treatment strategies.
Evaluation of the Hypertension Surveillance System at Pilot Hypertension Prevention and Control Health Facilities in Addis Ababa, Ethiopia, 2022
Authors: Goshim Nigus 
Published: January 2025

AbstractIntroductionHypertension is a major cause of premature death worldwide. Evaluating a surveillance system promotes the best use of data collection resources and ensures that systems operate effectively. It allows us to determine whether a specific system is useful for a particular public health initiative and is achieving the goals of the public health program and the data collection objectives. Ethiopian hypertension control initiatives were recently launched (2019) and implemented in limited health facilities. Therefore, this evaluation is aimed at the hypertension surveillance system of Addis Ababa, Ethiopia.MethodA descriptive cross-sectional study was conducted from December 21 to 31, 2022. Eight health care facilities were evaluated based on updated CDC guidelines for evaluating public health surveillance systems, and one key informant from each facility was interviewed using semi structured questionnaire. The data were analyzed descriptively. The data were analyzed with Excel 2021 and summarized by the frequency mean, percentage and rate.ResultsThe system detected cases (8/8) and monitored the control rate, attendance rate and missed visits. All interviewed staff (8/8) stated that the case definition, diagnosis and treatment algorism of hypertension were easy. Regarding acceptability, all staff believed that stakeholders were engaged, and 7/8 of the facilities recorded and reported data in a timely manner, but there was no acknowledgment given to the staff (0/8). The stability and data quality were 95% and 72.9, respectively. The representativeness was 24% to 36% based on prevalence, and regarding sex, it ranged from 88% to 108%. predictive positive value (31.4%) and zero flexibility.ConclusionThe system was useful for detecting morbidities and retaining patients on treatment. It has good simplicity, acceptability and stability but requires improvement in its flexibility, representativeness, data quality and positive predictive value. The system is better able to scale up to other health facilities.
Optimizing mouse metatranscriptome profiling by selective removal of redundant nucleic acid sequences
Authors: Roos Morgan , Bunga Samuel , Tan Asako , Maissy Erica , Skola Dylan , Richter Alexander , Whittaker Daniel S. , Desplats Paula , Zarrinpar Amir , Conrad Rick , Kuersten Scott 
Published: January 2025

AbstractMetatranscriptome (MetaT) sequencing is a critical tool for profiling the dynamic metabolic functions of microbiomes. In addition to taxonomic information, MetaT also provides real-time gene expression data of both host and microbial populations, thus permitting authentic quantification of the functional (enzymatic) output of the microbiome and its host. The main challenge to effective and accurate MetaT analysis is the removal of highly abundant rRNA transcripts from these complex mixtures of microbes, which can number in the thousands of individual species. Regardless of methodology for rRNA depletion, the design of rRNA removal probes based solely upon taxonomic content of the microbiome typically requires very large numbers of individual probes, making this approach complex to commercially manufacture, costly, and frequently technically infeasible. In previous work [1], we designed a set of depletion probes for human stool samples using a design strategy based solely on sequence abundance, completely agnostic of the microbiomal species present. Here, we show that the human-based probes are less effective when used with mouse cecal samples. However, adapting additional rRNA depletion probes specifically to cecal content provides both greater efficiency and consistency for MetaT analysis of mouse samples.ImportanceSequencing total RNA from microbiome samples is seriously impaired by the overwhelming proportion of rRNA to mRNA content. As much as 99% of sequencing reads can be assigned to the rRNA content, thus removal of these abundant transcripts is critical to MetaT analysis. The use of Ribo Zero Plus rRNA depletion probes designed for human gut microbiomes proved to be less effective and more inconsistent across mouse cecal donor samples, a common experimental system for microbiome studies. In the present work, we have extended and refined a taxonomically-neutral probe design method for mouse cecal content. The additional probes were carefully chosen to limit the number needed for effective depletion to reduce both the cost and risk of introducing bias to MetaT analysis. Our results demonstrate this method as efficient and consistent for rRNA removal in mouse cecal samples, thus providing a significant increase in the number of mRNA-rich sequencing reads for MetaT analysis.
EEG Dynamical Features during Variable-Intensity Cycling Exercise in Parkinson’s Disease
Authors: Alizadeh Zahra , Arasteh Emad , Mirian Maryam S. , Sacheli Matthew A. , Murray Danielle , Appel-Cresswell Silke , McKeown Martin J. 
Published: January 2025

1.Abstract1.1.BackgroundExercise is increasingly recognized as a beneficial intervention for Parkinson’s disease (PD), yet the optimal type and intensity of exercise remain unclear. This study investigated the relationship between exercise intensity and neural responses in PD patients, using electroencephalography (EEG) to explore potential neural markers that could be ultimately used to guide exercise intensity.1.2.MethodEEG data were collected from 14 PD patients (5 females) and 8 healthy controls (HC) performing stationary pedaling exercises at 60 RPM with resistance adjusted to target heart rates of 30%, 40%, 50%, 60%, and 70% of maximum heart rate. Subjects pedaled for 3 minutes at each intensity level in a counterbalanced order. Canonical Time-series Characteristics (Catch-22) features and Multi-set Canonical Correlation Analysis (MCCA) were utilized to identify common profiles of EEG features at increasing exercise intensity across subjects.1.3.ResultsWe identified a statistically significant MCCA component demonstrating a monotonic relationship with pedaling intensity. The dominant feature in this component was Periodicity Wang (PW), related to the autocorrelation of the EEG. Analysis revealed a consistent trend across features: six features increased with intensity, indicating heightened rhythmic engagement and sustained neural activation, while three features decreased, suggesting reduced variability and enhanced predictability in neural responses. Notably, PD patients exhibited more rigid, consistent response patterns compared to healthy controls (HC), who showed greater flexibility and variability in their neural adaptation across intensities.1.4.ConclusionThis study highlights the feasibility of using EEG-derived features to track exercise intensity in PD patients, identifying specific neural markers correlating with varying intensity levels. PD subjects demonstrate less inter-subject variability in motor responses to increasing intensity. Our results suggest that EEG biomarkers can be used to assess differing brain involvement with the same exercise of increasing intensity, potentially useful for guiding targeted therapeutic strategies and maximizing the neurological benefits of exercise in PD.
Impaired glymphatic clearance independently contributes to poor outcomes in Parkinson’s disease
Authors: Zarkali Angeliki , Thomas George , Paterson Ross , Hannaway Naomi , Dobreva Ivelina , Heslegrave Amanda J , Veleva Elena , Zetterberg Henrik , Weil Rimona S 
Published: January 2025

AbstractBackgroundImpaired glymphatic clearance may contribute to pathological accumulations in Parkinson’s (PD), but how it interacts with other processes causing dementia and poor outcomes remains unclear.ObjectivesClarify how glymphatic clearance impacts cognition in PD and its interaction with established imaging markers.MethodsWe used diffusion tensor image analysis along the perivascular space (DTI-ALPS) as an indirect marker of glymphatic clearance in 98 PD patients (31 PD-poor outcomes: dementia, mild cognitive impairment, frailty or death within 3-year follow-up; 67 PD-good outcomes) and 28 controls. We assessed DTI-ALPS relationship to cognition, white matter (fibre cross-section), cortical thickness, iron accumulation (quantitative susceptibility mapping (QSM)), and plasma markers (phosphorylated tau-181 (p-tau181 and neurofilament light (NFL)) cross-sectionally and longitudinally.ResultsDTI-ALPS was lower in PD-poor outcomes compared to PD-good outcomes and controls (p=0.005) with further longitudinal reductions only in PD-poor outcomes (group*time interaction: β=-0.013, p=0.021). Lower DTI-ALPS was associated with lower fibre cross-section in P, at baseline and longitudinally but with different spatial distribution from white matter changes relating to PD cognition. There was no correlation between baseline DTI-ALPS and plasma ptau-181 (p=0.642), NFL (p=0.448) or baseline cortical thickness. Lower DTI-ALPS was associated with accelerated cortical thinning within left precentral gyrus and changes in brain iron distribution.ConclusionsPD patients who develop poor outcomes show impaired glymphatic clearance at baseline that worsened longitudinally. DTI-ALPS correlated with white matter integrity and brain iron accumulation. However, both showed different spatial distribution than that seen in PD dementia; suggesting impaired glymphatic clearance contributes to cognitive decline in a distinct manner.
Repeated tDCS at clinically-relevant field intensity can boost concurrent motor learning in rats
Authors: Farahani Forouzan , Vöröslakos Mihály , Birnbaum Andrew M. , FallahRad Mohamad , Williams Preston T.J.A. , Martin John H. , Parra Lucas C. 
Published: January 2025

AbstractElectric fields used in clinical trials with transcranial direct current stimulation (tDCS) are small, with magnitudes that have yet to demonstrate measurable effects in preclinical animal models. We hypothesized that weak stimulation will nevertheless produce sizable effects, provided that it is applied concurrently with behavioral training, and repeated over multiple sessions. We tested this here in a rodent model of dexterous motor-skill learning. We developed a preparation that allows concurrent stimulation during the performance of a pellet-reaching task in freely behaving rats. The task was automated to minimize experimenter bias. We measured field magnitudes intracranially to calibrate the stimulation current. In this study, only male rats were used. Animals were trained for 20 min with concurrent epicranial tDCS over 10 daily sessions. Behavior was recorded with high-speed video to quantify reaching dynamics. We also measured motor-evoked potentials (MEPs) bilaterally with epidural microstimulation. The new electrode montage enabled stable stimulation over 10 sessions with a field intensity of 2V/m at the motor cortex. The number of successful reaches improved across days of training, and the rate of learning was higher in the anodal group as compared to sham-control animals (F(1)=7.12, p=0.008, N=24). MEPs were not systematically affected by tDCS. Posthoc analysis suggests that tDCS modulated motor learning only for right-pawed animals, improving success of reaching, but limiting stereotypy in these animals. Repeated and concurrent anodal tDCS can boost motor-skill learning at clinically-relevant field intensities. In this animal model the effect interacted with paw preference and was not associated with corticospinal excitability.Significance StatementThe effects of tDCS have been explored in numerous human clinical trials, but the mechanisms of action of weak electric fields remain unclear. In vitro studies show that constant electric fields at 2.5 V/m can enhance the efficacy of synapses undergoing plasticity. This study demonstrates in a rodent model that tDCS of only 2 Vm when applied concurrently to behavioral training can improve motor skill learning, and reduce stereotypy of reaching behavior. These effects accumulated over 10 days of training. Motor evoked potentials (MEP), which are often used to demonstrate plastic effects in humans on a time scale of hours, were not measurably affected by tDCS on this longer time scale.
RB1sequence variants in retinoblastoma. Analysis ofRB1variants in a database for correlation with pRB protein domains and clinical presentation
Authors: Diana Parma , Nicohol Tovar-Martelo , Florencia Giliberto , Irene Szijan 
Published: January 2025

ABSTRACTRetinoblastoma (RB) is the most common pediatric ocular tumor that occurs due to biallelic inactivation of theRB1tumor suppressor gene. RB may be unilateral or bilateral and is hereditary in 50% of cases. Inactivation of theRB1gene may occur by gross rearrangements (20%) or by small-length changes (80%): single nucleotide substitutions (SNVs) and insertions/deletions (INDELs). We analyzed the SNVs and INDELs of germinal origin, annotated in the databasehttp://rb1-lovd.d-lohmann.de, in order to find the frequency of different variants, their correlation with the protein pRB functional domains and with the clinical presentation. The number of mutation variants analyzed was 2103, 34% of them were nonsense, 34% indels, 22% splice-site and 10% missense. All these variants mainly gave rise to bilateral RB (88%), their frequency and distribution in relation to pRB domains varied between bilateral (Bi) and unilateral hereditary (Ug) RB. Nonsense variants occurred more frequently in Bi vs Ug, whereas missense variants were more frequent in Ug vs Bi. Indels and Splice-site variants were not significantly different between Bi and Ug. The most frequent pRB location of mutation variants was in the Pocket domain (the binding site of the E2F transcription factor), 58% for nonsense, 64% missense, 50% splice site and 45% INDELs. The slice-site of the consensus sequence most mutated was the first nucleotide of the donor, which is the driver of the splicing process.ConclusionsThe highest percentage of variants in RB corresponded to nonsense substitutions and indels, mainly affecting the Pocket domain, which is the major functional site for pRB regulatory process These results indicate the predominance of the most pathogenic variants in retinoblastoma.
Longitudinal wastewater-based surveillance of vancomycin-resistant Enterococci in tertiary-care hospitals
Authors: Au Emily , Acosta Nicole , Waddell Barbara J. , Lee Jangwoo , Du Kristine , Weyant R. Benson , Bautista Maria , McCalder Janine , Van Doorn Jennifer , Low Kashtin , Stefani September , Visser Gail , Clark Rhonda , Pitout Johann , Kim Joseph , Missaghi Bayan , Larios Oscar , Kanji Jamil , Vayalumkal Joseph , Leal Jenine , Westlund Paul , Quinn Robert R. , James Matthew , Lee Bonita , Pang Xiao-Li , Dalton Bruce , Frankowski Kevin , O’Grady Christine , Conly John M. , Hubert Casey R.J. , Parkins Michael D. 
Published: January 2025

AbstractSummaryVancomycin-resistantEnterococcus(VRE) is an important cause of healthcare-associated infections. We adapted wastewater-based surveillance as a tool to longitudinally monitor VRE in hospitals through the detection of vancomycin resistance genesvanAandvanB.MethodsWastewater from four tertiary-care hospitals (three adult and one pediatric, totaling >2300 inpatient beds) and all three municipal wastewater treatment plants (WWTP) in Calgary, Canada (∼1.4 million) was sampled weekly (March to September 2022) and every other week (September 2022 to March 2023). Wastewater pellets were collected, DNA extracted, andvanAandvanBquantified by qPCR.vanAandvanBgene copies were assessed as raw (copies/mL) and normalized with three different fecal biomarkers – total bacterial 16S-rRNA,BacteroidesHF183 16S-rRNA, and human 18S-rRNA. Raw and normalizedvanAandvanBabundance from each site was compared with clinically identified infections, vancomycin prescribing and hemodialysis services.FindingsvanAabundance was up to1085-fold higher p<0.0001, Mann-Whitney) andvanBup to 32-fold higher (p<0.01, Mann-Whitney) in adult hospitals compared to an aggregate municipal signal and exhibited significantly greater variation. Strong correlations between each method of fecal normalization and raw-measuredvanAandvanBwere observed, and no analysis method proved superior (Spearman’s r=0.50–0.96, p<0.0001).vanAabundance was strongly correlated with hemodialysis provision (Spearman’s r=0.8357, p<0.0001) but not vancomycin prescribing.InterpretationWastewater-based surveillance is a comprehensive, real-time tool capable of longitudinal hospital surveillance with the potential to transform the ability of infection control and antimicrobial stewardship programs to dynamically track, understand, and mitigate nosocomial antimicrobial-resistant pathogens.