bluuebunny/crossref_metadata_2025_split huggingface dataset:
Degradation and release of dissolved environmental RNAs from zebrafish cells
Authors: Xu Zhongneng 
Published: July 2023

ABSTRACTEnvironmental RNAs in water are gradually being applied in aquatic ecological surveys, water pollution monitoring, etc., but the current methods to detect environmental RNAs in water can mainly measure the RNAs in the filters that are used for filtering water samples, neglecting dissolved environmental RNAs in water. The sources and degradation profiles of dissolved environmental RNAs in water remain unknown. The present study was conducted to measure the permeability of extracted RNAs from zebrafish cells through filters, the degradation of extracted RNAs from zebrafish cells in tubes, and the release rate and degradation of dissolved environmental RNAs from living zebrafish cells and dying zebrafish cells, aiming to provide dynamic information from dissolved environmental RNAs in water. The results showed that there were no significant differences between the levels of extracted RNAs from zebrafish cells before filtration with 0.45 µm filters and those in the filtrates. The extracted RNAs from zebrafish cells degraded in water in the tubes, and after 2 months, more than 15% of RNAs in the groups of RNAs in water were still detected. The half-life of all the RNAs in the tubes was approximately 20∼43 days. During the 6-day experiment of the release and degradation of dissolved RNAs from living cells, an average of 4.1×10-4∼ 1.7×10-3pg dissolved RNAs (7.6×105∼ 3.2×106RNA bases) were secreted per cell per day into the liquid environment. During the 6-day experiment of the release and degradation of dissolved RNAs from dying cells, approximately 4.2 pg of dissolved RNAs released by a dying zebrafish cell in water could be detected. The dissolved environmental RNAs in water from zebrafish cells degraded faster in the presence of zebrafish cells: under the conditions without zebrafish cells, the average survival rate of the dissolved environmental RNAs in water per day was 98.4%/day; under the conditions with living zebrafish cells, the average survival rate per day was 49.7%/day; and under the conditions with dying zebrafish cells, the average survival rate per day was 34.9%/day. The estimated levels of dissolved environmental RNAs in water in fish tanks were too low to be detected by the current techniques. Although the methods in the present study need to be improved, this study may provide information to develop new ways to measure the dynamics of dissolved environmental RNAs in water and quantitatively analyze RNAs released into liquid environments of living and dying cells.
γδ T cell-mediated cytotoxicity against patient-derived healthy and cancer cervical organoids
Authors: Dong Junxue , Holthaus David , Peters Christian , Koster Stefanie , Ehsani Marzieh , Quevedo-Olmos Alvaro , Berger Hilmar , Zarobkiewicz Michal , Mangler Mandy , Gurumurthy Rajendra Kumar , Hedemann Nina , Chumduri Cindrilla , Kabelitz Dieter , Meyer Thomas F. 
Published: July 2023

AbstractCervical cancer is a leading cause of death among women globally, primarily driven by high-risk papillomaviruses. However, the effectiveness of chemotherapy is limited, underscoring the potential of personalized immunotherapies. Patient-derived organoids, which possess cellular heterogeneity, proper epithelial architecture and functionality, and long-term propagation capabilities offer a promising platform for developing viable strategies.In addition to αβ T cells and natural killer (NK) cells, γδ T cells represent a cell population with significant therapeutic potential against both hematologic and solid tumours.To evaluate the efficacy of γδ T cells in cervical cancer treatment, we generated patient-derived healthy and cancer ectocervical organoids. Furthermore, we examined transformed healthy organoids, expressing HPV16 oncogenes E6 and E7. We analysed the effector function ofin vitroexpanded γδ T cells upon co-culture with organoids. Our findings demonstrated that healthy cervical organoids were less susceptible to γδ T cell-mediated cytotoxicity compared to HPV-transformed organoids and cancerous organoids.To identify the underlying pathways involved in this observed cytotoxicity, we performed bulk-RNA sequencing on the organoid lines, revealing differences in DNA-damage and cell cycle checkpoint pathways, as well as transcription of potential γδ T cell ligands. We validated these results using immunoblotting and flow cytometry. We also demonstrated the involvement of BTN3A1 and BTN2A1, crucial molecules for γδ T cell activation, as well as differential expression of PDL1/CD274 in cancer, E6/E7+ and healthy organoids. Interestingly, we observed a significant reduction in cytotoxicity upon blocking MSH2, a protein involved in DNA mismatch-repair.In summary, we established a co-culture system of γδ T cells with cervical cancer organoids, providing a novelin vitromodel to optimize innovative patient-specific immunotherapies for cervical cancer.
PMS1as a target for splice modulation to prevent somatic CAG repeat expansion in Huntington’s disease
Authors: McLean Zachariah L. , Gao Dadi , Correia Kevin , Roy Jennie C. L. , Shibata Shota , Farnum Iris N. , Valdepenas-Mellor Zoe , Rapuru Manasa , Morini Elisabetta , Ruliera Jayla , Gillis Tammy , Lucente Diane , Kleinstiver Benjamin P. , Lee Jong-Min , MacDonald Marcy E. , Wheeler Vanessa C. , Pinto Ricardo Mouro , Gusella James F. 
Published: July 2023

AbstractHuntington’s disease (HD) is a dominantly inherited neurodegenerative disorder whose motor, cognitive, and behavioral manifestations are caused by an expanded, somatically unstable CAG repeat in the first exon ofHTTthat lengthens a polyglutamine tract in huntingtin. Genome-wide association studies (GWAS) have revealed DNA repair genes that influence the age-at-onset of HD and implicate somatic CAG repeat expansion as the primary driver of disease timing. To prevent the consequent neuronal damage, small molecule splice modulators (e.g., branaplam) that targetHTTto reduce the levels of huntingtin are being investigated as potential HD therapeutics. We found that the effectiveness of the splice modulators can be influenced by genetic variants, both atHTTand other genes where they promote pseudoexon inclusion. Surprisingly, in a novel hTERT-immortalized retinal pigment epithelial cell (RPE1) model for assessing CAG repeat instability, these drugs also reduced the rate ofHTTCAG expansion. We determined that the splice modulators also affect the expression of the mismatch repair genePMS1, a known modifier of HD age-at-onset. Genome editing at specificHTTandPMS1sequences using CRISPR-Cas9 nuclease confirmed that branaplam suppresses CAG expansion by promoting the inclusion of a pseudoexon inPMS1, making splice modulation ofPMS1a potential strategy for delaying HD onset. Comparison with another splice modulator, risdiplam, suggests that other genes affected by these splice modulators also influence CAG instability and might provide additional therapeutic targets.
Polyamine transport inhibition and cisplatin synergistically enhance tumor control through oxidative stress in murine head and neck cancer models
Authors: Yassin-Kassab Abdulkader , Wang Nathaniel , Foley Jackson , Stewart Tracy Murray , Burns Mark R. , Casero Robert A. , Harbison R. Alex , Duvvuri Umamaheswar 
Published: July 2023

AbstractBackgroundSurgery and/or platinum-based chemoradiation remain standard of care for patients with head and neck squamous cell carcinoma (HNSCC). While these therapies are effective in a subset of patients, a substantial proportion experience recurrence or treatment resistance. As cisplatin mediates cytotoxicity through oxidative stress while polyamines play a role in redox regulation, we posited that combining cisplatin with polyamine transport inhibitor, AMXT-1501, would increase oxidative stress and tumor cell death in HNSCC cells.MethodsCell proliferation was measured in syngeneic mouse HNSCC cell lines treated with cisplatin ± AMXT-1501. Synergy was determined by administering cisplatin and AMXT-1501 at a ratio of 1:10 to cancer cellsin vitro. Cancer cells were transferred onto mouse flanks to test the efficacy of treatmentsin vivo. Reactive oxygen species (ROS) were measured. Cellular apoptosis was measured with flow cytometry using Annexin V/PI staining. High-performance liquid chromatography (HPLC) was used to quantify polyamines in cell lines. Cell viability and ROS were measured in the presence of exogenous cationic amino acids.ResultsThe combination of cisplatin and AMXT-1501 synergizein vitroon HNSCC cell lines.In vivocombination treatment resulted in tumor growth inhibition greater than either treatment individually. The combination treatment increased ROS production and induced apoptotic cell death. HPLC revealed the synergistic mechanism was independent of intracellular polyamine levels. Supplementation of cationic amino acids partially rescued cancer cell viability and reduced ROS.ConclusionAMXT-1501 enhances the cytotoxic effects of cisplatinin vitroandin vivoin aggressive HNSCC cell lines through a polyamine-independent mechanism.
Integrative analysis of DNA replication origins and ORC/MCM binding sites in human cells reveals a lack of overlap
Authors: Tian Mengxue , Wang Zhenjia , Su Zhangli , Shibata Etsuko , Shibata Yoshiyuki , Dutta Anindya , Zang Chongzhi 
Published: July 2023

AbstractBased on experimentally determined average inter-origin distances of ∼100 kb, DNA replication initiates from ∼50,000 origins on human chromosomes in each cell cycle. The origins are believed to be specified by binding of factors like the Origin Recognition Complex (ORC) or CTCF or other features like G-quadruplexes. We have performed an integrative analysis of 113 genome-wide human origin profiles (from five different techniques) and 5 ORC-binding profiles to critically evaluate whether the most reproducible origins are specified by these features. Out of ∼7.5 million union origins identified by all datasets, only 0.27% were reproducibly obtained in at least 20 independent SNS-seq datasets and contained in initiation zones identified by each of three other techniques (20,250 shared origins), suggesting extensive variability in origin usage and identification. 21% of the shared origins overlap with transcriptional promoters, posing a conundrum. Although the shared origins overlap more than union origins with constitutive CTCF binding sites, G-quadruplex sites and activating histone marks, these overlaps are comparable or less than that of known Transcription Start Sites, so that these features could be enriched in origins because of the overlap of origins with epigenetically open, promoter-like sequences. Only 6.4% of the 20,250 shared origins were within 1 kb from any of the ∼13,000 reproducible ORC binding sites in human cancer cells, and only 4.5% were within 1 kb of the ∼11,000 union MCM2-7 binding sites in contrast to the nearly 100% overlap in the two comparisons in the yeast,S. cerevisiae. Thus, in human cancer cell lines, replication origins appear to be specified by highly variable stochastic events dependent on the high epigenetic accessibility around promoters, without extensive overlap between the most reproducible origins and currently known ORC- or MCM-binding sites.
A signature of platelet reactivity in CBC scattergrams reveals genetic predictors of thrombotic disease risk
Authors: Verdier Hippolyte , Thomas Patrick , Batista Joana , Kempster Carly , McKinney Harriet , Gleadall Nicholas , Danesh John , Mumford Andrew , Heemskerk Johan , Ouwehand Willem H. , Downes Kate , Astle William J. , Turro Ernest 
Published: July 2023

AbstractGenetic studies of platelet reactivity (PR) phenotypes may identify novel antiplatelet drug targets. However, discoveries have been limited by small sample sizes (n<5,000) due to the complexity of measuring PR. We trained a model to predict PR from complete blood count (CBC) scattergrams. A GWAS of this phenotype in 29,806 blood donors identified 21 distinct associations implicating 20 genes, of which six have been identified previously. The effect size estimates were significantly correlated with estimates from a study of flow-cytometry measured PR and a study of a phenotype ofin vitrothrombus formation. A genetic score of PR built from the 21 variants was associated with myocardial infarction and pulmonary embolism. Mendelian randomisation analyses showed PR to be causally associated with the risks of coronary artery disease, stroke and venous thromboembolism. Our approach provides a blueprint for employing phenotype imputation to study the determinants of hard-to-measure but biologically important haematological traits.Key pointsPlatelet reactivity can be predicted from scattergrams generated by haematology analysers of a type in widespread clinical use.Genetic analysis ofpredictedplatelet reactivity reveals associations with the risks of thrombotic diseases, including stroke.
Differential expansion and retention patterns of LRR-RLK genes across plant evolution
Authors: Kileeg Zachary , Haldar Aparna , Khan Hasna , Qamar Arooj , Mott G. Adam 
Published: July 2023

AbstractTo maximize overall fitness, plants must accurately respond to a host of growth, developmental, and environmental signals throughout their life. Many of these internal and external signals are perceived by the leucine-rich repeat receptor-like kinases, which play roles in regulating growth, development, and immunity. This largest family of receptor kinases in plants can be divided into subfamilies based on conservation of the kinase domain, which demonstrates that shared evolutionary history often indicates shared molecular function. Here we investigate the evolutionary history of this family across the evolution of 112 plant species. We identify lineage-specific expansions of the malectin-domain containing subfamily LRR subfamily I primarily in the Brassicales and bryophytes. Most other plant lineages instead show a large expansion in LRR subfamily XII, which in Arabidopsis is known to contain key receptors in pathogen perception. This striking asymmetric expansion may reveal a dichotomy in the evolutionary history and adaptation strategies employed by plants. A greater understanding of the evolutionary pressures and adaptation strategies acting on members of this receptor family offers a way to improve functional predictions for orphan receptors and simplify identification of novel stress related receptors.
Transcriptomic Effects of Low-Dose Inorganic Arsenic Exposure on Murine Bone Marrow-Derived Macrophages
Authors: Illingworth Emily J. , Maertens Alexandra , Sillé Fenna C.M. 
Published: July 2023

ABSTRACTBoth tissue-resident macrophages and monocytes recruited from the bone marrow that transform into tissue-resident cells play critical roles in mediating homeostasis as well as in the pathology of inflammatory diseases. Inorganic arsenic (iAs) is the most common drinking water contaminant worldwide and represents a major public health concern. Several diseases that macrophages have implicated involvement in are caused by iAs exposure, including cardiovascular disease, cancer, and increased risk of infectious disease. Therefore, understanding the effects of iAs exposure on macrophages can help us better grasp the full range of arsenic immunotoxicity and better design therapeutic targets for iAs-induced diseases particularly in exposed populations. In this study, we analyzed the transcriptome of low dose iAs-exposed male and female murine bone marrow-derived macrophages (BMDMs) with either M0, M1, or M2 stimulation. We identified differentially expressed genes by iAs in a sex- and stimulation-dependent manner and used bioinformatics tools to predict protein-protein interactions, transcriptional regulatory networks, and associated biological processes. Overall, our data suggest that M1-stimulated, especially female-derived, BMDMs are most susceptible to iAs exposure. Most notably, we observed significant downregulation of major proinflammatory transcription factors, like IRF8, and its downstream targets, as well as genes encoding proteins involved in pattern recognition and antigen presentation, such as TLR7, TLR8, and H2-D1, potentially providing causal insight regarding arsenic’s role in perturbing immune responses to infectious diseases. We also observed significant downregulation of genes involved in processes crucial to coordinating a proinflammatory response including leukocyte migration, differentiation, and cytokine and chemokine production and response. Finally, we discovered that 24 X-linked genes were dysregulated in iAs-exposed female stimulation groups compared to only 3 across the iAs-exposed male stimulation groups. These findings elucidate the potential mechanisms underlying the sex-differential iAs-associated immune-related disease risk.
An allelic series at theEDNRB2locus controls diverse piebalding patterns in the domestic pigeon
Authors: Maclary Emily T. , Wauer Ryan , Phillips Bridget , Brown Audrey , Boer Elena F. , Samani Atoosa M. , Shapiro Michael D. 
Published: July 2023

ABSTRACTVariation in pigment patterns within and among vertebrate species reflects underlying changes in cell migration and function that can impact health, reproductive success, and survival. The domestic pigeon (Columba livia) is an exceptional model for understanding the genetic changes that give rise to diverse pigment patterns, as selective breeding has given rise to hundreds of breeds with extensive variation in plumage color and pattern. Here, we map the genetic architecture of a suite of pigmentation phenotypes known as piebalding. Piebalding is characterized by patches of pigmented and non-pigmented feathers, and these plumage patterns are often breed-specific and stable across generations. Using a combination of quantitative trait locus mapping in F2laboratory crosses and genome-wide association analysis, we identify a locus associated with piebalding across many pigeon breeds. This shared locus harbors a candidate gene,EDNRB2, that is a known regulator of pigment cell migration, proliferation, and survival. We discover multiple distinct haplotypes at theEDNRB2locus in piebald pigeons, which include a mix of protein-coding, noncoding, and structural variants that are associated with depigmentation in specific plumage regions. These results identify a role forEDNRB2in pigment patterning in the domestic pigeon, and highlight how repeated selection at a single locus can generate a diverse array of stable and heritable pigment patterns.AUTHOR SUMMARYBoth wild and domestic birds show striking variation in pigment patterning, and these pigment patterns can play critical roles in mate choice, communication, and camouflage. Despite the importance of pigment patterning for survival and reproductive success, the mechanisms that control pigment patterning remain incompletely understood. In domestic pigeons, artificial selection has given rise to a wide array of pigmentation patterns within a single species, including a suite of phenotypes called “piebalding” that is characterized by regional loss of feather pigment. Here, we took advantage of the wide array of distinct piebalding phenotypes in domestic pigeons to map the genetic basis of region-specific loss of plumage pigment. Rather than focusing on a single piebalding pattern, we sought to broadly understand genetic control of regional pigment loss by examining several breeds with different piebalding patterns. We compared the genomes of piebald and non-piebald pigeons using both genetic crosses and genome-wide association studies, and identified several genetic changes affecting the endothelin receptor geneEDNRB2. Our findings highlight how independent mutations at a single locus can drive diversification of plumage pigment patterning within a species.
DNA methylation insulates genic regions from CTCF loops near nuclear speckles
Authors: Roseman Shelby A. , Siegenfeld Allison P. , Lee Ceejay , Lue Nicholas Z. , Waterbury Amanda L. , Liau Brian B. 
Published: July 2023

AbstractThe insulator protein CTCF is essential for mediating chromatin loops and regulating gene expression. While it is established that DNA methylation hinders CTCF binding, the impacts of this methylation-sensitive CTCF binding on chromatin architecture and transcription are poorly defined. Here, we used a selective DNMT1 inhibitor to investigate the characteristics and functions of ‘reactivated’ CTCF peaks resulting from global DNA demethylation. We found that reactivated peaks preferentially form chromatin loops on gene bodies and interact with highly-looping partner peaks located in regions of active chromatin. Notably, both reactivated CTCF peaks and their highly-looping partners are enriched near nuclear speckles, condensate bodies implicated in transcription and splicing. Utilizing targeted protein degradation, we specifically depleted CTCF and nuclear speckles to elucidate their functional interplay. By degrading CTCF upon DNMT1 inhibition, we revealed that CTCF is important for DNMT1 inhibitor-dependent interactions between chromatin and speckle proteins. Moreover, we found that CTCF promotes the activation of genes near speckles upon DNMT1 inhibition. Conversely, acute depletion of nuclear speckles revealed that they influence transcription but do not maintain CTCF binding or looping. Collectively, our study suggests a model wherein DNA methylation prevents spurious CTCF occupancy and interactions with regulatory elements near nuclear speckles, yet CTCF looping is robust towards the loss of speckles.
Evolution of the enhancer-rich regulatory region of thePOU1F1gene
Authors: Wallis Michael , Xu Qianlan , Krawczyk Michal , Skowronska-Krawczyk Dorota 
Published: July 2023

AbstractPrecise spatio-temporal expression of genes in organogenesis is regulated by the coordinated interplay of DNA elements such as promoter and enhancers present in the regulatory region of a given locus. POU1F1 transcription factor plays a crucial role in the development of somatotrophs, lactotrophs and thyrotrophs in the anterior pituitary gland, and in maintaining high expression of growth hormone, prolactin and TSH. In mouse, expression ofPOU1F1is controlled by a region fenced by two CTCF sites, containing 5 upstream enhancer elements, designated E-A (5’ to 3’). We performed comparative sequence analysis of this regulatory region and discovered that three elements, B, C and E, are present in all vertebrate groups except Agnatha. One very long (>2kb) element (A) is unique to mammals suggesting a specific change in regulation of the gene in this group. Using DNA accessibility assay (ATAC-seq) we showed that conserved elements in anterior pituitary of four non-mammals are open, suggesting functionality as regulatory elements. We showed that, in many non-mammalian vertebrates, an additional upstream exon closely follows element E, leading to alternatively spliced transcripts. Here, element E functions as an alternative promoter, but in mammals this feature is lost, suggesting that conversion of alternative promoter to enhancer could be one evolutionary mechanism for enhancer birth. Our work shows that regulation ofPOU1F1changed markedly during the course of vertebrate evolution, use of a small number of enhancer elements combined with alternative promoters in non-mammalian vertebrates being replaced by use of a unique combination of enhancers in mammals.