bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genetic regulation of nascent RNA maturation revealed by direct RNA nanopore sequencing
Authors: Choquet Karine , Chaumont Louis-Philippe , Bache Simon , Baxter-Koenigs Autum R. , Churchman L. Stirling 
Published: August 2024

AbstractQuantitative trait loci analyses have revealed an important role for genetic variants in regulating alternative splicing (AS) and alternative cleavage and polyadenylation (APA) in humans. Yet, these studies are generally performed with mature mRNA, so they report on the outcome rather than the processes of RNA maturation and thus may overlook how variants directly modulate pre-mRNA processing. The order in which the many introns of a human gene are removed can substantially influence AS, while nascent RNA polyadenylation can affect RNA stability and decay. However, how splicing order and poly(A) tail length are regulated by genetic variation has never been explored. Here, we used direct RNA nanopore sequencing to investigate allele-specific pre-mRNA maturation in 12 human lymphoblastoid cell lines. We found frequent splicing order differences between alleles and uncovered significant single nucleotide polymorphism (SNP)-splicing order associations in 17 genes. This included SNPs located in or near splice sites as well as more distal intronic and exonic SNPs. Moreover, several genes showed allele-specific poly(A) tail lengths, many of which also had a skewed allelic abundance ratio.HLAclass I transcripts, which encode proteins that play an essential role in antigen presentation, showed the most allele-specific splicing orders, which frequently co-occurred with allele-specific AS, APA or poly(A) tail length differences. Together, our results expose new layers of genetic regulation of pre-mRNA maturation and highlight the power of long-read RNA sequencing for allele-specific analyses.
A simple MiMIC based approach for tagging endogenous genes to visualise live transcriptionin vivo
Authors: Beadle Lauren Forbes , Sutcliffe Catherine , Ashe Hilary L. 
Published: August 2024

AbstractLive imaging of transcription in theDrosophilaembryo using the MS2 or PP7 systems is transforming our understanding of transcriptional regulation. However, insertion of MS2/PP7 stem loops into endogenous genes requires laborious CRISPR genome editing. Here we exploit the previously described Minos-mediated integration cassette (MiMIC) transposon system inDrosophilato establish a method for simply and rapidly inserting MS2/PP7 cassettes into any of the thousands of genes carrying a MiMIC insertion. In addition to generating a variety of stem loop donor fly stocks, we have made new stocks expressing the complementary coat proteins fused to different fluorescent proteins. We show the utility of this MiMIC-based approach by MS2/PP7 tagging and live imaging transcription of endogenous genes and the long non-coding RNA,roX1, in the embryo. We also present live transcription data from larval brains, the wing disc and ovary, thereby extending the tissues that can be studied using the MS2/PP7 system. Overall, this first high throughput method for tagging mRNAs inDrosophilawill facilitate the study of transcription dynamics of thousands of endogenous genes in a range ofDrosophilatissues.
Training Data Diversity Enhances the Basecalling of Novel RNA Modification-Induced Nanopore Sequencing Readouts
Authors: Wang Ziyuan , Liu Ziyang , Fang Yinshan , Zhang Hao Helen , Sun Xiaoxiao , Hao Ning , Que Jianwen , Ding Hongxu 
Published: August 2024

ABSTRACTAccurately basecalling sequence backbones in the presence of nucleotide modifications remains a substantial challenge in nanopore sequencing bioinformatics. It has been extensively demonstrated that state-of-the-art basecallers are less compatible with modification-induced sequencing signals. A precise basecalling, on the other hand, serves as the prerequisite for virtually all the downstream analyses. Here, we report that basecallers exposed to diverse training modifications gain the generalizability to analyze novel modifications. With synthesized oligos as the model system, we precisely basecall various out-of-sample RNA modifications. From the representation learning perspective, we attribute this generalizability to basecaller representation space expanded by diverse training modifications. Taken together, we conclude increasing the training data diversity as a novel paradigm for building modification-tolerant nanopore sequencing basecallers.
A Comparison of Skeletal Muscle Diffusion Tensor Imaging Tractography Seeding Methods
Authors: Damon Bruce M. , Guzman Roberto Pineda , Lockard Carly A. , Zhou Xingyu 
Published: August 2024

AbstractThe internal arrangement of a muscle’s fibers with respect to its mechanical line of action (muscle architecture) is a major determinant of muscle function. Muscle architecture can be quantified using diffusion tensor magnetic resonance imaging-based tractography, which propagates streamlines from a set of seed points by integrating vectors that represent the direction of greatest water diffusion (and by inference, the local fiber orientation). Previous work has demonstrated that tractography outcomes are sensitive to the method for defining seed points, but this sensitivity has not been fully examined. To do so, we developed a realistic simulated muscle architecture and implemented four novel methods for tract seeding: seeding along the muscle-aponeurosis boundary with an updated procedure for rounding seed points prior to lookup in the muscle boundary mask and diffusion tensor matrix (APO-3); voxel-based seeding throughout the muscle volume at a user-specified spatial frequency (VXL-1); voxel-based seeding throughout the muscle volume at a variable spatial frequency (VXL-2), and seeding near external and internal muscle boundaries (VXL-3). We then implemented these methods in an example human dataset. The updated aponeurosis seeding procedures allow more accurate and robust tract propagation from seed points. The voxel-based seeding methods had quantification outcomes that closely matched the updated aponeurosis seeding method. Further, the voxel-based methods can accelerate the overall workflow and may be beneficial in high throughput analysis of multi-muscle datasets. Continued evaluation of these methods in a wider range of muscle architectures is warranted.
Distinct functional domains of Dystroglycan regulate inhibitory synapse formation and maintenance in cerebellar Purkinje cells
Authors: Jahncke Jennifer N. , Schnell Eric , Wright Kevin M. 
Published: August 2024

AbstractDystroglycan is a cell adhesion molecule that localizes to synapses throughout the nervous system. While Dystroglycan is required to maintain inhibitory synapses from cerebellar molecular layer interneurons (MLIs) onto Purkinje cells (PCs) whether initial synaptogenesis during development is dependent on Dystroglycan has not been examined. We show that conditional deletion ofDystroglycanfrom Purkinje cells prior to synaptogenesis results in impaired MLI:PC synapse formation and function due to reduced presynaptic inputs and abnormal postsynaptic GABAAreceptor clustering. Using genetic manipulations that disrupt glycosylation of Dystroglycan or truncate its cytoplasmic domain, we show that Dystroglycan’s role in synapse function requires both extracellular and intracellular interactions, whereas synapse formation requires only extracellular interactions. Together, these findings provide molecular insight into the mechanism of inhibitory synapse formation and maintenance in cerebellar cortex.
Brain activity discriminates acoustic simulations of the same environment
Authors: Matten Viola G , Stirnberg Rüdiger , van de Par Steven , Ewert Stephan D , Flanagin Virginia L 
Published: August 2024

AbstractIn complex acoustic environments, sound localization involves the integration of numerous interrelated auditory and cognitive cues, making it challenging to understand their relationship to brain activity. Here, we use virtual acoustics to probe the brain’s response to auditory distance cues in a realistic environment. We developed a system to record the actual MRI environment, simulated the same room with different degrees of accuracy, then presented sounds at one of two locations in the room. We implemented a novel auditory fMRI sequence to record brain activity. Despite only minor differences in acoustics between the auralizations, it was possible to decode all three rooms from brain activity. A systematic analysis revealed that the direct-to-reverberant energy ratio (DRR) drove brain activity differences between auralizations, centered on the posterior auditory cortex (AC). The results provide strong evidence that the posterior AC processes DRR for spatial auditory perception.Impact statementA novel fMRI sequence and recording technique are combined with virtual acoustics and multi-variate analyses to decode room simulations from brain activity during distance perception and identify the auditory factors that drive the pattern of activity in the brain.
EXPERIMENTAL DESIGN ISSUES ASSOCIATED WITH CLASSIFICATIONS OF HYPERSPECTRAL IMAGING DATA
Authors: Nansen Christian , Lee Hyoseok , Mesgaran Mohsen B. 
Published: August 2024

AbstractHyperspectral imaging has emerged as a pivotal tool to classify plant materials (seeds, leaves, and whole plants), pharmaceutical products, food items, and many other objects. This communication addresses two issues, which appear to be over-looked or ignored in >99% of hyperspectral imaging studies: 1) the “small N, large P” problem, when number of spectral bands (explanatory variables, “P”) surpasses number of observations, (“N”) leading to potential model over-fitting, and 2) absence of independent validation data in performance assessments of classification models. Based on simulations of randomly generated data, we illustrate risks associated with these issues. We explore and discuss consequences of over-fitting and risks of misleadingly high accuracy that can result from having a large number of variables relative to observations. We highlight connections of these issues with radiometric repeatability (levels of stochastic noise). A method is proposed wherein a theoretical dataset is generated to mirror the structure of an actual dataset, with the classification of this theoretical dataset serving as a reference. By shedding light on important and common experimental design issues, we aim to enhance methodological rigor and transparency in classifications of hyperspectral imaging data and foster improved and effective applications across various science domains.
Sld3CBD-Cdc45 structural insights into Cdc45 recruitment for CMG complex formation
Authors: Li Hao , Ishizaki Izumi , Kato Koji , Sun XiaoMei , Muramatsu Sachiko , Itou Hiroshi , Ose Toyoyuki , Araki Hiroyuki , Yao Min 
Published: August 2024

AbstractDNA replication requires Cdc45 and GINS recruitment into the MCM duplex hexamer by initiation factors to form an active helicase, the Cdc45–MCM–GINS (CMG) complex, at the replication origins. The initiation factor Sld3 is a central regulator of Cdc45 and GINS recruitment worked with Sld7 together. However, the mechanism through which Sld3 regulates CMG complex formation remains unclear. Here, we present the structure of the Sld3 Cdc45-binding-domain in complex with Cdc45 (Sld3CBD–Cdc45), showing detailed interactions and conformational changes required for binding to each other. The mutant analysis indicated that the binding between Sld3CBD and Cdc45 could be broken easily. We also revealed that Sld3CBD, GINS, and MCM bind to different sites on Cdc45 in the Sld3CDB–CMG model, indicating that after recruitment of Cdc45, Sld7–Sld3 could remain in Cdc45–MCM until CMG formation. The consistency between the particle size of Sld7–Sld3–Cdc45 and the distance between Sld3CBDs in the Cdc45–MCM dimer indicated the binding manner of the Cdc45–Sld3–[Sld7]2–Sld3–Cdc45 off/on MCM duplex hexamer. A DNA-binding assay of Sld3 and its complexes with single-stranded ARS1 fragments revealed a relationship between the dissociation of Sld7–Sld3 from CMG and the unwound single-stranded DNA. These findings help to further our understanding of the molecular basis of regulation of CMG complex formation by Sld3.
A microendovascular system can record precise neural signals from cortical and deep vessels with minimal invasiveness
Authors: Iwata Takamitsu , Nakamura Hajime , Uemura Takafumi , Araki Teppei , Matsumura Takaki , Abe Takaaki , Nezu Toshikazu , Takagaki Masatoshi , Ozaki Tomohiko , Miura Shinpei , Fukuma Ryohei , John Sam E. , Grayden David B. , Kishima Haruhiko , Sekitani Tsuyoshi , Yanagisawa Takufumi 
Published: August 2024

AbstractMinimally invasive intravascular electroencephalography (ivEEG) signals are a promising tool for developing clinically feasible brain–computer interfaces (BCIs) that restore communication and motor functions in paralyzed patients. However, current ivEEG techniques can only record signals from the superior sagittal sinus (SSS), making it challenging to record motor responses related to hand and mouth movements from brain regions distant from the SSS, despite their critical role in BCIs. Here, using micro intravascular electrodes, ivEEGs recorded from the cortical or deep veins of eight pigs could measure cortical activities with greater signal power and better spatial resolution than those recording in the SSS, thus allowing mapping of the sensorimotor and visual functional areas. Additionally, electrical stimulation in the cortical vein between the micro intravascular electrodes induced muscle contractions contralateral to the stimulated area in five anesthetized pigs. These results demonstrate that ivEEG using micro intravascular electrodes is a promising tool for developing BCIs.
RNA functional control by hydrolysis reversible acylation
Authors: Liu Kaizheng , Kietrys Anna M. 
Published: August 2024

AbstractWe present the hydrolysis reversible acylation of RNA 2′-OH for functional control. Compared with previously reported acylating reagents, our new moleculeEST1Aeliminated the need of toxic organic molecules in deacylation. Instead, the acylation was removed by biocompatible intracellular stimuli such as esterase and histidine. After validating the reversibility using oligonucleotides, we applied our acylating reagent on functional RNAs, including broccoli aptamer, eGFP mRNA, and eGFP siRNA, and gained activity control on them. Further inhibitory and enhancive assays in HepG2 and U87MG cells confirmed that both carboxylesterase and cholinesterase contributed to deacylationin cellulo. By taking advantage of differences in esterase expression among cell lines, our new strategy can benefit the development of cell-selective gene therapeutics.
Analysis and control of untemplated DNA polymerase activity for guided synthesis of kilobase-scale DNA sequences
Authors: Castle Simeon D. , Woolfson Adrian , Linshiz Gregory , Riley Blake T. , Samuel Ifor D.W. , Holliger Philipp , Oldfield Lauren , Hessel Andrew , Gorochowski Thomas E. 
Published: August 2024

AbstractDNA polymerases are complex molecular machines able to replicate genetic material using a template-driven process. While the copying function of these enzymes is well established, their ability to perform untemplated DNA synthesis is less well characterized. Here, we explore the ability of DNA polymerases to synthesize DNA fragments in the absence of template. We use long-read nanopore sequencing and real-time PCR to observe the synthesis of pools of DNA products derived from a diverse set of natural and engineered DNA polymerases across varying temperatures and buffer compositions. We detail the features of the DNA fragments generated, enrichment of select sequence motifs, and demonstrate that the sequence composition of the synthesized DNA may be altered by modifying environmental conditions. This work provides an extensive data set to better discern the process of untemplated DNA polymerase activity and may support its potential repurposing as a technology for the guided synthesis of DNA sequences on the kilobase-scale and beyond.
Roles for the canonical polarity machinery in thede novoestablishment of polarity in budding yeast spores
Authors: Cooperman Benjamin , McMurray Michael 
Published: August 2024

ABSTRACTThe yeastSaccharomyces cerevisiaebuds at sites pre-determined by cortical landmarks deposited during prior budding. During mating between haploid cells in the lab, external pheromone cues override the cortical landmarks to drive polarization and cell fusion. By contrast, in haploid gametes (called spores) produced by meiosis, a pre-determined polarity site drives initial polarized morphogenesis independent of mating partner location. Spore membranes are madede novoso existing cortical landmarks were unknown, as were the mechanisms by which the spore polarity site is made and how it works. We find that the landmark canonically required for distal budding, Bud8, stably marks the spore polarity site along with Bud5, a GEF for the GTPase Rsr1 that canonically links cortical landmarks to the conserved Cdc42 polarity machinery. Cdc42 and other GTPase regulators arrive at the site during its biogenesis, after spore membrane closure but apparently at the site where membrane synthesis began, and then these factors leave, pointing to the presence of discrete phases of maturation. Filamentous actin may be required for initial establishment of the site, but thereafter Bud8 accumulates independent of actin filaments. These results suggest a distinct polarization mechanism that may provide insights into gamete polarization in other organisms.SIGNIFICANCE STATEMENTDormant budding yeast spores possess a single, stable cortical site that marks the location where polarized growth occurs upon dormancy exit. It was not known how the site forms or which molecules comprise it.Using fluorescently tagged proteins in living cells undergoing sporulation, the authors found proteins canonically involved in polarization of non-spore cells arriving at the polarity site in a choreographed manner and required for site function.These findings point to a distinct polarity mechanism from non-spore cells and raise new questions about polarity protein interactions with membranes that may be applicable to gametogenesis in other organisms.
Changes in hypothalamic subunits volume and their association with metabolic parameters and gastrointestinal appetite-regulating hormones following bariatric surgery
Authors: Lachance Amélie , Daoust Justine , Pelletier Mélissa , Caron Alexandre , Carpentier André C. , Biertho Laurent , Maranzano Josefina , Tchernof André , Dadar Mahsa , Michaud Andréanne 
Published: August 2024

AbstractBackgroundSome nuclei of the hypothalamus are known for their important roles in maintaining energy homeostasis and regulating food intake. Moreover, obesity has been associated with hypothalamic inflammation and morphological alterations, as indicated by increased volume. However, the reversibility of these changes after bariatric surgery-induced weight loss remains underexplored.ObjectiveThe aim of this study was to characterize volume changes in hypothalamic subunits up to two years following bariatric surgery and to determine whether these differences were associated with changes in metabolic parameters and levels of gastrointestinal appetite-regulating hormone levels.MethodsParticipants with severe obesity undergoing bariatric surgery were recruited. They completed high-resolution T1-weighted brain magnetic resonance imaging (MRI) before bariatric surgery and at 4, 12 and 24 months post-surgery. Blood samples collected during the fasting and postprandial states were analyzed for glucagon-like peptide 1 (GLP-1), peptide YY (PYY), and ghrelin concentrations. The hypothalamus was segmented into 5 subunits per hemisphere using a publicly available automated tool. Linear mixed-effects models were employed to examine volume changes between visits and their associations with variables of interest.ResultsA total of 73 participants (mean age 44.5 ± 9.1 years, mean BMI 43.5 ± 4.1 kg/m2) were included at baseline. Significant volume reductions were observed in the whole left hypothalamus 24 months post-surgery. More specifically, decreases were noted in both the left anterior-superior and left posterior subunits at 12 and 24 months post-surgery (all p<0.05, after FDR correction). These reductions were significantly associated with the percentage of total weight loss (both subunits p<0.001), improvements in systolic blood pressure (both subunits p<0.05), and an increase in postprandial PYY (both subunits p<0.05).ConclusionThese results suggest that some hypothalamic morphological alterations observed in the context of obesity could potentially be reversed with bariatric surgery induced-weight loss.
Inverse correlation between NLRP3 and TMEM176B as a possible tool for colorectal cancer progression
Authors: Cambui Raylane Adrielle Gonçalves , Roa Mariela Estefany Vera , Leal Vinicius Nunes Cordeiro , Yamada Suemy Melim , de Oliveira Leonardo Antônio Teixeira , Nadaf Izabel Nazira , Lelis Cleiton Ribeiro , da Cunha Pedroso Rafaela Cássia , do Nascimento Costa Rafaela Cardoso , Amorim Raquel Maria Neves , do Amaral Jefferson Rodrigues , Lemes Polyana Silva , Rizzo Leonardo Amorim , do Espírito Santo Gilmar Ferreira , Elias Rosa Maria , Cogliati Bruno , Hill Marcelo , Pontillo Alessandra 
Published: August 2024

AbstractPurposeTMEM176B has been recently identified as a novel player in anti-cancer immune responses by negatively modulating the NLRP3 inflammasome activation in colorectal cancer (CRC). Yet, the TMEM176B/NLRP3 axis in CRC needs to be deeply investigated.MethodsTMEM176B and NLRP3 expression were evaluated in CRC patients by immunohistochemistry and RT-qPCR assays. The prognostic relevance of TMEM176B and NLRP3 was determined by in silico analysis. The NLRP3 inflammasome activation in the CRC microenvironment was assessed in a peripheral blood mononuclear cells (PBMC) and CRC cell line (HCT-116) spheroids co-culture assay.ResultsReduced NLRP3 and increased TMEM176B expression correlates to CRC stages and poor survival. The in vitro assay showed HCT-116 cells activated NLRP3 inflammasome in PBMC.ConclusionThese findings evidence the inverse correlation between NLRP3 and TMEM176B in CRC progression, suggesting them as a predictive tool.