bluuebunny/crossref_metadata_2025_split huggingface dataset:
KaryoTap Enables Aneuploidy Detection in Thousands of Single Human Cells
Authors: Mays Joseph C. , Mei Sally , Kogenaru Manjunatha , Quysbertf Helberth M. , Bosco Nazario , Zhao Xin , Bianchi Joy J. , Goldberg Aleah , Kidiyoor Gururaj Rao , Holt Liam J. , Fenyö David , Davoli Teresa 
Published: September 2023

AbstractInvestigating chromosomal instability and aneuploidy within tumors is essential for understanding tumorigenesis and developing diagnostic and therapeutic strategies. Single-cell DNA sequencing technologies have enabled such analyses, revealing aneuploidies specific to individual cells within the same tumor. However, it has been difficult to scale the throughput of these methods to detect rare aneuploidies while maintaining high sensitivity. To overcome this deficit, we developed KaryoTap, a method combining custom targeted DNA sequencing panels for the Tapestri platform with a computational framework to enable detection of chromosome- and chromosome arm-scale aneuploidy (gains or losses) and copy number neutral loss of heterozygosity in all human chromosomes across thousands of single cells simultaneously. KaryoTap allows detecting gains and losses with an average accuracy of 83% for arm events and 91% for chromosome events. Importantly, together with chromosomal copy number, our system allows us to detect barcodes and gRNAs integrated into the cells’ genome, thus enabling pooled CRISPR- or ORF-based functional screens in single cells. As a proof of principle, we performed a small screen to expand the chromosomes that can be targeted by our recently described CRISPR-based KaryoCreate system for engineering aneuploidy in human cells. KaryoTap will prove a powerful and flexible approach for the study of aneuploidy and chromosomal instability in both tumors and normal tissues.
Visualizing interactions of VDAC1 in live cells using a tetracysteine tag
Authors: Pilic Johannes , Oflaz Furkan E. , Gottschalk Benjamin , Erdogan Yusuf C. , Graier Wolfgang F. , Malli Roland 
Published: September 2023

SummaryThe voltage-dependent anion channel 1 (VDAC1) is a crucial gatekeeper in the outer mitochondrial membrane, controlling metabolic and energy homeostasis. The available methodological approaches fell short of accurate visualization of VDAC1 in living cells. To permit precise VDAC1 imaging, we used the tetracysteine (TC)-tag approach and visualized VDAC1 dynamics in living cells. TC-tagged VDAC1 had a cluster-like distribution on mitochondria. The majority of VDAC1-clusters were localized at endoplasmic reticulum (ER)-mitochondria contact sites. Notably, VDAC1 colocalized with BCL-2 Antagonist/Killer (BAK)-clusters upon apoptotic stimulation. Additionally, VDAC1 was found at mitochondrial fission sites, likely promoting mitochondrial fragmentation. These findings highlight the suitability of the TC-tag for live-cell imaging of VDAC1, shedding light on the roles of VDAC1 in multiple cellular processes.
Genetic manipulation of anIxodes scapulariscell line
Authors: Singh Nisha , Rolandelli Agustin , O’Neal Anya J. , Butler L. Rainer , Samaddar Sourabh , Laukaitis-Yousey Hanna J. , Butnaru Matthew , Mohr Stephanie E. , Perrimon Norbert , Pedra Joao H. F. 
Published: September 2023

AbstractAlthough genetic manipulation is one of the hallmarks in model organisms, its applicability to non-model species has remained difficult due to our limited understanding of their fundamental biology. For instance, manipulation of a cell line originated from the blacklegged tickIxodes scapularis,an arthropod that serves as a vector of several human pathogens, has yet to be established. Here, we demonstrate the successful genetic modification of the commonly used tick ISE6 line through ectopic expression and clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) genome editing. We performed ectopic expression using nucleofection and attained CRISPR-Cas9 editing via homology dependent recombination. Targeting the E3 ubiquitin ligase X-linked inhibitor of apoptosis (xiap) and its substratep47led to alteration in molecular signaling within the immune deficiency (IMD) network and increased infection of the rickettsial agentAnaplasma phagocytophiluminI. scapularisISE6 cells. Collectively, our findings complement techniques for genetic engineering of ticksin vivoand aid in circumventing the long-life cycle ofI. scapularis,of which limits efficient and scalable molecular genetic screens.ImportanceGenetic engineering in arachnids has lagged compared to insects, largely because of substantial differences in their biology. This study unveils the implementation of ectopic expression and CRISPR-Cas9 gene editing in a tick cell line. We introduced fluorescently tagged proteins in ISE6 cells and edited its genome via homology dependent recombination. We ablated the expression ofxiapandp47, two signaling molecules present in the immune deficiency (IMD) pathway ofI. scapularis. Impairment of the tick IMD pathway, an analogous network of the tumor necrosis factor receptor in mammals, led to enhanced infection of the rickettsial agentA. phagocytophilum. Altogether, our findings provide a critical technical resource to the scientific community to enable a deeper understanding of biological circuits in the blacklegged tickIxodes scapularis.
No model to rule them all: a systematic comparison of 83 thermal performance curve models across traits and taxonomic groups
Authors: Kontopoulos Dimitrios - Georgios , Sentis Arnaud , Daufresne Martin , Glazman Natalia , Dell Anthony I. , Pawar Samraat 
Published: September 2023

AbstractIn ectotherms, the performance of physiological, ecological and life-history traits universally increases with temperature to a maximum before decreasing again. Identifying the most appropriate thermal performance model for a specific trait type has broad applications, from metabolic modelling at the cellular level to forecasting the effects of climate change on population, ecosystem and disease transmission dynamics. To date, numerous mathematical models have been designed, but a thorough comparison among them is lacking. In particular, we do not know if certain models consistently outperform others and how factors such as sampling resolution and trait or organismal identity influence model performance. To fill this knowledge gap, we compile 2,739 thermal performance datasets from diverse traits and taxa, to which we fit a comprehensive set of 83 existing mathematical models. We detect remarkable variation in model performance that is not primarily driven by sampling resolution, trait type, or taxonomic information. Our results reveal a surprising lack of well-defined scenarios in which certain models are more appropriate than others. To aid researchers in selecting the appropriate set of models for any given dataset or research objective, we derive a classification of the 83 models based on the average similarity of their fits.
Ultrasensitive quantification of HIV-1 cell-to-cell transmission in primary human CD4+T cells measures viral sensitivity to broadly neutralizing antibodies
Authors: Mazurov Dmitriy , Herschhorn Alon 
Published: September 2023

ABSTRACTHIV-1 efficiently replicatesin vivoby direct transmission from infected to uninfected CD4+T cells at confined local sites designated virological synapses (VSs). VSs are formed by cell junctions between HIV-1 envelope glycoproteins (Envs) on an infected cell and CD4 on an uninfected cell. These sites facilitate highly efficient viral transmission and contribute to HIV-1 evasion from neutralizing antibodies, but accurate quantification of the efficiency of cell-cell transmission is still challenging. Here, we developed an ultrasensitive HIV-1 cell- to-cell transmission assay that triggers the expression of thenanoluciferase (nluc)gene in target cells upon transmission and after reverse transcription of the HIV-1 RNA genome. The assay is based on insertion of thenlucgene in an antisense orientation into HIV-1 provirus; Nluc expression is blocked in virus-producing cells because thenlucgene contains an intron that can be efficiently spliced out only when mRNA is transcribed from the opposite (sense) strand. Thus, only sense mRNA that is spliced and subsequently reversed transcribed during transmission to target cells will support Nluc expression. Assay optimization resulted in a very low background, >99% splicing efficiency, high sensitivity and wide dynamic range for detection of cell-cell transmission in T cell lines and primary CD4+T cells. The new reporter vector can detect cell-cell transmission using single-round viral vectors and HIV-1 molecular clones, which provide viral proteins of different HIV-1 strains, and reproducibly measures sensitivity of HIV-1 transmission to antibody neutralizationin vitro. This assay will contribute to understanding fundamental mechanisms of HIV-1 cell-to-cell transmission, allow evaluation of pre-existing or acquired HIV-1 resistance in clinical trials, and can be adapted to study other retroviruses.
Discovering Disease Genes in PPI Networks: A Bridge from Centrality to Communities
Authors: Khan Mehwish Wahid , Shams Malick Rauf Ahmed , Cherifi Hocine 
Published: September 2023

AbstractTargeted therapies have become pivotal in modern clinical oncology, driven by a molecularlevel understanding of cancer’s intricacies, its progression, and innovative research and technology. Personalized and targeted treatments hinge on identifying key genes, hub genes, or biomarkers. Protein-protein interaction (PPI) networks are instrumental in understanding the molecular basis of diseases. While existing literature has identified significant genes based on network centrality, investigations based on community-aware centrality have been notably absent. This omission matters because disease networks frequently display modular structures, necessitating a new perspective. This study bridges the gap between network centrality and community-based investigations. Indeed, in modular networks, node influence can be categorized into two types: local impact within its community, determined by intra-community connections, and global effect on nodes in other communities, established through inter-community links. This concept extends conventional centrality measures to networks with a community structure. Initially, we performed a comparative analysis of seven PPI networks related to cancer and noncancerous conditions. We explore the correlation between classical network centralities and their equivalents at the global (inter-community) and local (intra-community) levels. Notably, we consistently observed a high correlation between network degree and local degree centrality in all PPIs, leading us to select local degree centrality for further investigation. Pronounced modularity characterizes prostate and cervical disease networks. Consequently, we investigate these networks to identify key genes at the local community level and validate them by examining their expression levels. Variations in gene expression between cancerous and non-cancerous tissues bolster our findings. We identify a novel set of genes as potential key players in prostate and cervical cancer. Specifically, in cervical cancer, the top genes at the mesoscopic level include AKT1, CDK2, BRCA1, VEGFA, SRC, PSMD14, MRPL3, TP53, and NUP37. Meanwhile, the top genes identified in prostate cancer are FOS, TP53, UBA52, HLA-B, TSPO, and CD19. Although we focus on cancer data, our methodology’s versatility makes it applicable to other disease networks, opening avenues to identify key genes as potential drug targets.
CCR3 inhibition suppresses inflammation-driven recruitment of peripheral immune cells to the eye
Authors: Lopez Yesenia , Caryotakis Sofia , Raina Sharda , Rege Sanket V. , Harish Reema , Ray Rebecca , Kosti Idit , Teichert Arnaud , Minami S. Sakura , Dhande Onkar S. 
Published: September 2023

AbstractC-C chemokine receptor type 3 (CCR3) has been linked with age-related macular degeneration (AMD) pathologies. Specifically, its function as an immune modulator in AMD remains unclear. To address this question, we investigated the impact of CCR3 inhibition on inflammation, a key driver of AMD pathologies, by assessing inflammatory cytokines and infiltrating immune cells in two models of ocular inflammation.Mice were orally dosed twice a day with AKST4290, a CCR3 small molecular inhibitor, in the sodium iodate (NaIO3) and myelin oligodendrocyte glycoprotein (MOG) models. A combination of autoradiography and analytical chemistry techniques were used to assess drug concentration and distribution. Bead-based multiplexing technology was used to determine cytokine concentrations, and flow cytometry and immunohistochemistry was used to ascertain ocular immune-cell composition.CCR3 expression was detected in the retinal pigment epithelium (RPE)/choroid complex where AKST4290 was found to preferentially accumulate at sustained levels. In the NaIO3model, inhibition of CCR3 with AKST4290 significantly decreased both the concentration of specific chemokines and the number of multiple populations of infiltrating peripheral immune cell. Furthermore, effects of CCR3 inhibition on immune cell infiltration were confirmed in the MOG model.These data demonstrate that CCR3 inhibition strongly modulates local inflammation by impacting both cytokine concentrations and immune cell composition in ocular diseases.Moreover, these findings together with the known role of CCR3 in promoting pathologic angiogenesis implicate a pleiotropic role for CCR3 in AMD.
Machine learning to identify clinically relevantCandidayeast species
Authors: Shankarnarayan Shamanth A. , Charlebois Daniel A. 
Published: September 2023

AbstractBackgroundFungal infections, especially due toCandidaspecies, are on the rise. Multi-drug resistant organism such asCandida aurisare difficult and time consuming to identify accurately.Machine learning is increasingly being used in health care, especially in medical imaging. In this study, we evaluated the effectiveness of six convolutional neural networks (CNNs) to identify four clinically importantCandidaspecies.Materials and MethodsWet-mounted images were captured using bright field live-cell microscopy followed by separating single cells, budding cells, and cell group images which were then subjected to different machine learning algorithms (custom CNN, VGG16, ResNet50, InceptionV3, EfficientNetB0, and EfficientNetB7) to learn and predictCandidaspecies.ResultsAmong the six algorithms tested, the InceptionV3 model performed best in predictingCandidaspecies from microscopy images. All models performed poorly on raw images obtained directly from the microscope. The performance of all models increased when trained on single and budding cell images. The InceptionV3 model identified budding cells ofC. albicans, C. auris,C. glabrata(Nakaseomyces glabrata), andC. haemuloniiin 97.0%, 74.0%, 68.0%, and 66.0% cases, respectively. For single cells ofC. albicans, C. auris,C. glabrata, andC. haemuloniiInceptionV3 identified 97.0%, 73.0%, 69.0%, and 73.0% cases, respectively. The sensitivity and specificity of InceptionV3 were respectively 77.1% and 92.4%.ConclusionThis study provides proof of concept that microscopy images from wet mounted slides can be used to identifyCandidayeast species using machine learning quickly and accurately.
Integrative Genomic Analyses Reveal Putative Cell Type-specific Targets of theDrosophilaEts Transcription Factor Pointed
Authors: Bollepogu Raja Komal Kumar , Yeung Kelvin , Shim Yoon-Kyung , Mardon Graeme 
Published: September 2023

AbstractThe Ets domain transcription factors direct diverse biological processes throughout all metazoans and are implicated in development as well as in tumor initiation, progression and metastasis. TheDrosophilaEts transcription factor Pointed (Pnt) is required for several aspects of eye development and regulates cell cycle progression, specification, and differentiation. Despite its critical role in development, very few targets of Pnt have been reported previously. Here, we used chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) to determine the genome-wide occupancy of Pnt in late larval eye discs. We identified enriched regions that mapped to an average of 6,941 genes, the vast majority of which are novel putative Pnt targets. Integrating ChIP-seq data with two other larval eye single cell genomics datasets (scRNA-seq and snATAC-seq) reveals genes that may be putative cell type-specific genes regulated by Pnt. Finally, our ChIP-seq data predict cell type-specific functional enhancers that were not reported previously. Our study provides a greatly expanded list of putative Pnt targets in the eye and is a resource for future studies that will allow mechanistic insights into complex developmental processes regulated by Pnt.
Mitonuclear effects on sex ratio persist across generations in interpopulation hybrids
Authors: Edmands Suzanne , Denova Jacob R. , Flanagan Ben A. , Jah Murad , Applebaum Scott L. 
Published: September 2023

AbstractEukaryotic energy production requires tight coordination between gene products from both the nuclear and mitochondrial genomes. Because males and females often have different energetic strategies, this mitonuclear coordination can be expected to differentially impact the two sexes. Previous work found evidence for sex-specific mitonuclear effects in the copepodTigriopus californicusby comparing two parental lines and their reciprocal F1 crosses. However, an alternative hypothesis is that the patterns could instead be driven by the parental source of nuclear alleles. Here we test this alternative hypothesis by extending the same cross to F2 hybrids, who receive both maternal and paternal nuclear alleles from F1 hybrids. Results confirm mitonuclear effects on sex ratio, with distorted ratios persisting from the F1 to F2 generations, despite reduced fitness in F2 hybrids. No sex by cross interactions were found for other phenotypic traits measured. Mitochondrial DNA content was shown to be higher in females, the more stress-tolerant sex. Both routine metabolic rate and oxidative DNA damage were found to be lower in F2 hybrids than in parentals. Confirmation of sex-biased mitonuclear effects inT. californicusis notable, given that the species lacks sex chromosomes, which can confound interpretations of sex-specific mitochondrial effects.
Overexpression of Rheb, a positive Tor regulator, reveals principles of rod outer segment size control
Authors: Gilliam Larissa , Willoughby John J , Annan William , White Diana , Jensen Abbie M 
Published: September 2023

AbstractThe outer segment of retinal rod photoreceptors is a highly modified cilium with the typical ciliary microtubular axoneme but additionally there are hundreds of densely packed, stacked, discrete intramembranous discs containing phototransduction machinery. The outer segment is continuously regenerated or renewed throughout the life of the animal through the combined processes of proximal growth and distal shedding. Since the pioneering experiments decades ago that demonstrated the process of rod outer segment renewal very little research has been directed towards understanding mechanistic principles of renewal and homeostasis, and many important questions remain unaddressed. The rod outer segment is a prime example of tight size control that is amenable to experimental and computational analyses. This study uses zebrafish molecular genetics with high resolution imaging, quantitative analyses, and computational modeling to uncover principles that regulate rod outer segment homeostatic length control. By genetically increasing rod outer segment growth, we show that outer segments lengthen and shedding does not accelerate to offset this added growth.
Intrinsic and extrinsic factors regulate FtsZ function inCaulobacter crescentus
Authors: Barrows Jordan M , Anderson Ashley S , Talavera-Figueroa Barbara K , Goley Erin D 
Published: September 2023

AbstractBacterial cell division is crucial for replication and requires careful coordination via a complex set of proteins collectively known as the divisome. The tubulin-like GTPase FtsZ is the master regulator of this process and serves to recruit downstream divisome proteins and regulate their activities. Upon arrival at mid-cell, FtsZ associates with the membrane via anchoring proteins and exhibits treadmilling motion, driven by its GTP binding and hydrolysis activities. Treadmilling is proposed to play a role in Z-ring condensation, as well as in distribution and regulation of peptidoglycan (PG) cell wall remodeling enzymes. FtsZ polymer superstructure and dynamics are central to its function, yet their regulation is incompletely understood. We sought to address these gaps in knowledge by modulating intrinsic and extrinsic regulators of FtsZ and evaluating their effectsin vitroand in cells, alone and in combination. To do this, we leveraged the cell cycle control features ofCaulobacter crescentus.We observed thatCaulobacterFtsZ variants that abrogate GTP hydrolysis impact FtsZ dynamics and Z-ring positioning, with little to no effect on Z-ring structure or constriction. Production of an FtsZ variant lacking its disordered C-terminal linker (ΔCTL) resulted in aberrant Z-ring dynamics and morphology, misregulated PG metabolism, and cell lysis. Combining ΔCTL and GTPase mutations was additive, suggesting they each act independently to control the Z-ring. Modulating levels of FtsA resulted in formation of multiple Z-rings that failed to constrict, suggesting roles in regulating both FtsZ superstructure and the activity of downstream divisome components. Collectively, our results indicate that GTP hydrolysis serves primarily to position the Z-ring at mid-cell, the CTL regulates both Z-ring structure and downstream signaling, and FtsA contributes to all aspects of FtsZ assembly and function. The additive effects of these elements are required to support robust and efficient cell division.
Potential of Ilhéus virus to emerge
Authors: Plante Kenneth S. , Plante Jessica A. , Azar Sasha R. , Shinde Divya P. , Scharton Dionna , Versiani Alice F. , Oliveira da Silva Natalia Ingrid , Strange Taylor , Sacchetto Lívia , Fokam Eric B. , Rossi Shannan L. , Weaver Scott C. , Marques Rafael E. , Nogueira Mauricio L. , Vasilakis Nikolaos 
Published: September 2023

AbstractIlhéus virus (ILHV)(Flaviviridae: Orthoflavivirus) is an arthropod-borne virus (arbovirus) endemic to Central and South America and the Caribbean. First isolated in 1944, most of our knowledge derives from surveillance and seroprevalence studies. These efforts have detected ILHV in a broad range of mosquito and vertebrate species, including humans, but laboratory investigations of pathogenesis and vector competence have been lacking. Here, we develop several immune intact murine models that closely recapitulate human neuroinvasive disease with strain- and age-specific virulence, as well as a uniformly lethal immunocompromised model. Replication kinetics in several vertebrate and invertebrate cell lines demonstrate that ILHV is capable of replicating to high titers in a wide variety of potential host and vector species. Lastly, vector competence studies provide strong evidence for efficient infection of and potential transmission byAedesspecies mosquitoes, despite ILHV’s phylogenetically clustering withCulexvectored flaviviruses, suggesting ILHV is poised for emergence in the neotropics.TeaserMurine models of ILHV mimic human disease, andAedesspecies of mosquitoes are highly susceptible to infection and dissemination.