bluuebunny/crossref_metadata_2025 huggingface dataset:
Introducing ARTiMiS: A low-cost flow imaging microscope for phytoplankton monitoring in engineered and natural environments
Authors: Gincley Benjamin , Khan Farhan , Hartnett Elaine , Fisher Autumn , Pinto Ameet J.
Published: March 2024

Manual microscopy is the gold standard for phytoplankton monitoring in diverse engineered and natural environments. However, it is both labor-intensive and requires specialized training for accuracy and consistency, and therefore difficult to implement on a routine basis without significant time investment. Automation can reduce this burden by simplifying the measurement to a single indicator (e.g., chlorophyll fluorescence) measurable by a probe, or by processing samples on an automated cytometer for more granular information. The cost of commercially available flow imaging cytometers, however, poses a steep financial barrier to adoption. To overcome these labor and cost barriers, we developed ARTiMiS: the Autonomous Real-Time Microbial ‘Scope. The ARTiMiS is a low-cost flow imaging microscopy-based platform with onboard software capable of providing species-level quantitation of phytoplankton communities in real-time. ARTiMiS leverages novel multi-modal imaging and onboard machine learning-based data processing that is currently optimized for a curated and expandable database of industrially relevant microalgae. We demonstrate its operational limits, performance in identification of laboratory-cultivated microalgae, and potential for continuous monitoring of complex microalgal communities in full-scale cultivation systems.

Synopsis

We introduce a platform for low-cost real-time imaging monitoring of phytoplankton and demonstrate its utility in real-time monitoring of laboratory- and full-scale microalgal cultivation systems.
Early-stage lung adenocarcinoma is driven by an injury-associated, plastic cell state dependent on a KRAS-ITGA3-SRC axis
Authors: Moye Aaron L. , Dost Antonella F. M. , Ietswaart Robert , Sengupta Shreoshi , Ya VanNashlee , Aluya Chrystal , Fahey Caroline G. , Louie Sharon M. , Paschini Margherita , Kim Carla F.
Published: March 2024

Glycine 12 mutations in the GTPase KRAS (KRASG12) are a known initiating event for lung adenocarcinoma (LUAD) with broad clinical relevance. KRASG12mutations promote cell-intrinsic rewiring of the lung alveolar type II progenitor (AT2) cells, but to what extent such changes interplay with pathways essential for lung homeostasis and cell fate is unclear. We used single-cell RNA-seq (scRNA-seq) from AT2-mesenchyme organoid co-cultures, mouse models, and stage IA LUAD patient samples to identify conserved regulators of AT2 cell transcriptional dynamics and the impact of KRASG12Dwith temporal resolution. In AT2WTorganoids, a transient injury/plasticity state preceded AT2 self-renewal and AT1 differentiation. Early-stage AT2KRAScells exhibited perturbed gene expression dynamics most noted by retention of the injury/plasticity state. At later time points in tumorigenesis, AT2KRAScells consisted of heterogeneous populations that could be defined by either the injury state or high expression of an AT2 cell signature. The injury state in AT2KRAScells of LUAD in patients, mice, and organoids was distinguishable from AT2WTstates by altered receptor expression, including co-expression of ITGA3 and SRC. The combination of clinically relevant KRASG12Dand SRC inhibitors to target the oncogenic injury cell state impaired AT2KRASorganoid growth. Thus, an injury/plasticity signature characterized as an essential step in lung repair is used during alveolar cell self-renewal and during initiation and progression of LUAD. Early-stage lung cancer may be susceptible to intervention by targeting the oncogenic-specific nature of this cell state.
Evolution Under Thermal Stress AffectsEscherichia coli’s Resistance to Antibiotics
Authors: Bullivant Austin , Lozano-Huntelman Natalie , Tabibian Kevin , Leung Vivien , Armstrong Dylan , Dudley Henry , Savage Van M. , Rodríguez-Verdugo Alejandra , Yeh Pamela J
Published: March 2024

Exposure to both antibiotics and temperature changes can induce similar physiological responses in bacteria. Thus, changes in growth temperature may affect antibiotic resistance. Previous studies have found that evolution under antibiotic stress causes shifts in the optimal growth temperature of bacteria. However, little is known about how evolution under thermal stress affects antibiotic resistance. We examined 100+ heat-evolved strains ofEscherichia colithat evolved under thermal stress. We asked whether evolution under thermal stress affects optimal growth temperature, if there are any correlations between evolving in high temperatures and antibiotic resistance, and if these strains’ antibiotic efficacy changes depending on the local environment’s temperature. We found that: (1) surprisingly, most of the heat-evolved strains displayed a decrease in optimal growth temperature and overall growth relative to the ancestor strain, (2) there were complex patterns of changes in antibiotic resistance when comparing the heat-evolved strains to the ancestor strain, and (3) there were few significant correlations among changes in antibiotic resistance, optimal growth temperature, and overall growth.

Importance

Escherichia coli, a bacteria species often found within the intestinal tract of warm-blooded organisms, can be harmful to humans. Like all species of bacteria,E. colican evolve, particularly in the presence of stressful conditions such as extreme temperatures or antibiotic treatments. Recent evidence suggests that when encountering one source of stress, an organism’s ability to deal with a different source of stress is also affected. With global climate change and the continued evolution of antibiotic-resistant bacteria, the need to further investigate how temperature and antibiotics interact is clear. The significance of our research is in identifying possible correlations between temperature and antibiotic stress, broadening our understanding of how stressors affect organisms, and allowing for insights into possible future evolutionary pathways.
PNAbind: Structure-based prediction of protein-nucleic acid binding using graph neural networks
Authors: Sagendorf Jared M. , Mitra Raktim , Huang Jiawei , Chen Xiaojiang S. , Rohs Remo
Published: March 2024

The recognition and binding of nucleic acids (NAs) by proteins depends upon complementary chemical, electrostatic and geometric properties of the protein-NA binding interface. Structural models of protein-NA complexes provide insights into these properties but are scarce relative to models of unbound proteins. We present a deep learning approach for predicting protein-NA binding given the apo structure of a protein (PNAbind). Our method utilizes graph neural networks to encode spatial distributions of physicochemical and geometric properties of the protein molecular surface that are predictive of NA binding. Using global physicochemical encodings, our models predict the overall binding function of a protein and can discriminate between specificity for DNA or RNA binding. We show that such predictions made on protein structures modeled with AlphaFold2 can be used to gain mechanistic understanding of chemical and structural features that determine NA recognition. Using local encodings, our models predict the location of NA binding sites at the level of individual binding residues. Binding site predictions were validated against benchmark datasets, achieving AUROC scores in the range of 0.92-0.95. We applied our models to the HIV-1 restriction factor APOBEC3G and show that our predictions are consistent with experimental RNA binding data.
Cambrian origin of the arachnid brain reveals early divergence of Chelicerata
Authors: Strausfeld Nicholas J. , Hirth Frank
Published: March 2024

Fossils from the lower Cambrian provide crucial insights into the diversification of arthropod lineages: Mandibulata, exemplified by centipedes, insects, and crustaceans; and Chelicerata, represented by sea spiders, horseshoe crabs, and arachnids, the last including spiders, scorpions, and ticks1. Two mid-Cambrian genera claimed as stem chelicerates areMollisoniaandSanctacaris,defined by a carapaced prosoma equipped with clustered limbs, followed by a segmented trunk opisthosoma equipped with appendages for swimming and respiration2–4. Up to the present, the phyletic status of Mollisoniidae and Sanctacariidae has been that of a basal chelicerate2, stemward of Leanchoiliidae, whose neuromorphology resembles that of extant Merostomata (horseshoe crabs)5. Here, we identify preserved traces of neuronal tissues inMollisonia symmetricathat crucially depart from a merostome organization. Instead, a radiating organization of metameric neuropils occupying most of its prosoma is situated behind a pair of oval unsegmented neuropils directly connected to paired chelicerae extending from the front of the prosoma. The disposition and connections of these neuropils is the neuroanatomical signature that denotes a complete reversal of the three genetically distinct domains defining arthropod brains6. Thus, inMollisoniathe deutocerebrum is the most rostral domain with the proso-and protocerebral domains folded backwards such that tracts from the principal eyes extend caudally to reach their prosocerebral destination. These defining neuromorphological features illuminate a marine origin of Arachnida from which evolved the planet’s most successful arthropodan predators.
Open Iris - An Open Source Framework for Video-Based Eye-Tracking Research and Development
Authors: Sadeghi Roksana , Ressmeyer Ryan , Yates Jacob , Otero-Millan Jorge
Published: March 2024

Eye-tracking is an essential tool in many fields, yet existing solutions are often limited for customized applications due to cost or lack of flexibility. We present OpenIris, an adaptable and user-friendly open-source framework for video-based eye-tracking. OpenIris is developed in C# with modular design that allows further extension and customization through plugins for different hardware systems, tracking, and calibration pipelines. It can be remotely controlled via a network interface from other devices or programs. Eye movements can be recorded online from camera stream or offline post-processing recorded videos. Example plugins have been developed to track eye motion in 3-D, including torsion. Currently implemented binocular pupil tracking pipelines can achieve frame rates of more than 500Hz. With the OpenIris framework, we aim to fill a gap in the research tools available for high-precision and high-speed eye-tracking, especially in environments that require custom solutions that are not currently well-served by commercial eye-trackers.

CCS CONCEPTS

Applied computing→ Life and medical sciences.
Geny: A Genotyping Tool for Allelic Decomposition of Killer Cell Immunoglobulin-Like Receptor Genes
Authors: Zhou Qinghui , Ghezelji Mazyar , Hari Ananth , Ford Michael K.B. , Holley Connor ,   COVNET Consortium, Mirabello Lisa , Chanock Stephen , Sahinalp S. Cenk , Numanagić Ibrahim
Published: March 2024

Accurate genotyping of Killer cell Immunoglobulin-like Receptor (KIR) genes plays a pivotal role in enhancing our understanding of innate immune responses, disease correlations, and the advancement of personalized medicine. However, due to the high variability of the KIR region and high level of sequence similarity among different KIR genes, the currently available genotyping methods are unable to accurately infer copy numbers, genotypes and haplotypes of individual KIR genes from next-generation sequencing data. Here we introduce Geny, a new computational tool for precise genotyping of KIR genes. Geny utilizes available KIR haplotype databases and proposes a novel combination of expectation-maximization filtering schemes and integer linear programming-based combinatorial optimization models to resolve ambiguous reads, provide accurate copy number estimation and estimate the haplotype of each copy for the genes within the KIR region. We evaluated Geny on a large set of simulated short-read datasets covering the known validated KIR region assemblies and a set of Illumina short-read samples sequenced from 25 validated samples from the Human Pangenome Reference Consortium collection and showed that it outperforms the existing genotyping tools in terms of accuracy, precision and recall. We envision Geny becoming a valuable resource for understanding immune system response and consequently advancing the field of patient-centric medicine.
Expression of fibroblast activation protein-α in human deep venous thrombus
Authors: Oguri Nobuyuki , Gi Toshihiro , Nakamura Eriko , Furukoji Eiji , Goto Hiroki , Maekawa Kazunari , Tsuji Atsushi B. , Nishii Ryuichi , Aman Murasaki , Moriguchi-Goto Sayaka , Sakae Tatefumi , Azuma Minako , Yamashita Atsushi
Published: March 2024

Background

Fibroblast activation protein-α (FAP), a type-II transmembrane serine protease, is expressed during wound healing, in cancer-associated fibroblasts, and in chronic fibrosing diseases. However, its expression in deep vein thrombus (DVT) remains unclear. Therefore, in this study, we investigated FAP expression and localization in DVT.

Methods

First, we pathologically accessed aspirated thrombi of patients with DVT (n=14), classifying thrombotic areas as follows; fresh, cellular lysis, endothelialization, fibroblastic reaction. Endothelialization and fibroblastic reaction were defined as organizing reactions. We immunohistochemically examined FAP-expressed areas and expressed cells. Second, we analyzed FAP expression in cultured dermal fibroblasts.

Results

All the aspirated thrombi showed a mixture of at least three of the four thrombotic areas. Specifically, 83 % of aspirated thrombi showed fresh and organizing reactions within each thrombus. Immunohistochemical expression of FAP was restricted in organizing area. Further, FAP expression in the thrombi was mainly found in vimentin-positive or α-smooth muscle actin-positive fibroblasts in double immunofluorescence. Some CD163-positive macrophages also showed FAP expression. FAP mRNA and protein levels significantly increased in cultured fibroblasts with low- proliferative activity under 0.1% fetal bovine serum (FBS) than that in fibroblasts under 10% FBS. Fibroblasts cultured in 10% FBS showed a significant decrease in FAP mRNA levels following supplementation with hemin, but not with thrombin.

Conclusions

The heterogeneous composition of the venous thrombi suggests the existence of a multistep thrombus formation process in human DVT. Further, fibroblasts or myofibroblasts may express FAP in organizing process in DVT. FAP expression may increase in fibroblasts with low-proliferative activity.
Arctic greening drives changes in the diet and gut microbiome of a resident herbivore with consequences for fitness
Authors: Kamenova Stefaniya , Albon Steve D. , Loe Leif Egil , Irvine R. Justin , Langvatn Rolf , Gusarova Galina , de Muinck Eric Jacques , Trosvik Pål
Published: March 2024

Rapid climate warming is “greening” the tundra due to higher plant productivity, particularly of deciduous shrubs and grasses, potentially changing the nutritional base for herbivores. However, the consequences for herbivores are unclear. Although the gut microbiome is an essential factor, mediating the capacity to adapt to dietary change, few studies have investigated the effect of annual changes in the diet-gut microbiome nexus on fitness traits. In a model system, the Svalbard reindeer, a species experiencing the greatest rate of climate warming on Earth, we investigate how climate-induced changes in diet and the gut microbiome impact reindeer body mass. Using high-resolution DNA metabarcoding, we quantified diet and microbiome composition in rumen samples from 97 animals culled in October from 1998 to 2004 in four different valleys. Overall diet diversity and grass consumption were significantly higher following warmer summers, while intake of the dwarf shrubSalix polarisincreased with the maximum normalised difference vegetation index (NDVI). The proportion ofSalixhad a significant positive effect on autumn body mass, which in turn increases ovulation rates and overwinter survival. There was a significant positive effect ofSalixon the microbiome diversity, while between-year variation in Bacteroidota, the most common bacteria phylum, was also significantly, positively related to maximum NDVI. However, a structural equation model revealed that the direct “path” effect of the proportion ofSalixin the diet on reindeer body mass was stronger than the indirect path effect, mediated through the gut microbiome. Our results suggest a positive impact of climate-driven Arctic greening on Svalbard reindeer fitness, operating through a change in diet composition, partially mediated by the changes in the gut microbiome. These findings advance our understanding of the mechanistic underpinnings of Arctic warming on herbivore populations and contribute to the debate about the importance of the diet−gut microbiome nexus in facilitating species resilience in the face of climate change.
Automated model discovery for human cardiac tissue: Discovering the best model and parameters
Authors: Martonová Denisa , Peirlinck Mathias , Linka Kevin , Holzapfel Gerhard A. , Leyendecker Sigrid , Kuhl Ellen
Published: March 2024

For more than half a century, scientists have developed mathematical models to understand the behavior of the human heart. Today, we have dozens of heart tissue models to choose from, but selecting the best model is limited to expert professionals, prone to user bias, and vulnerable to human error. Here we take the human out of the loop and automate the process of model discovery. Towards this goal, we establish a novel incompressible orthotropic constitutive neural network to simultaneously discover both, model and parameters, that best explain human cardiac tissue. Notably, our network features 32 individual terms, 8 isotropic and 24 anisotropic, and fully autonomously selects the best model, out of more than 4 billion possible combinations of terms. We demonstrate that we can successfully train the network with triaxial shear and biaxial extension tests and systematically sparsify the parameter vector withL1-regularization. Strikingly, we robustly discover a four-term model that features a quadratic term in the second invariantI2, and exponential quadratic terms in the fourth and eighth invariantsI4f,I4n, andI8fs. Importantly, our discovered model is interpretable by design and has parameters with well-defined physical units. We show that it outperforms popular existing myocardium models and generalizes well, from homogeneous laboratory tests to heterogeneous whole heart simulations. This is made possible by a new universal material subroutine that directly takes the discovered network weights as input. Automating the process of model discovery has the potential to democratize cardiac modeling, broaden participation in scientific discovery, and accelerate the development of innovative treatments for cardiovascular disease.

Our source code, data, and examples are available athttps://github.com/LivingMatterLab/CANN.
ROUA Database: 300 Human Genomes from the border of Ukraine and Romania
Authors: Shchubelka Khrystyna , Wolfsberger Walter W. , Oleksyk Olga T. , Hasynets Yaroslava , Patskun Silviya , Vakerych Mykhailo , Kish Roman , Mirutenko Violeta , Mirutenko Vladislav , Cotoraci Coralia Adina , Pop Calin , Neagu Olimpia , Baltă Cornel , Herman Hildegard , Mare Paula , Dumitra Simona , Papiu Horatiu , Hermenean Anca , Oleksyk Taras K.
Published: March 2024

We present a multi-layered data source (ROUA Database) providing the results of Whole Genome Sequencing of two human populations in the Carpathian Mountains region, specifically Ukraine’s Transcarpathia and Romania’s Satu Mare and Baia Mare provinces, areas previously underexplored in population genomics. The database contains the raw and annotated files of the whole genome sequences from 300 individuals from these regions, including annotations of common and unique genetic variants following a sampling protocol designed to capture the genetic diversity of Ukrainians and Romanians, including minority groups like Wallachians and Roma. The data is hosted on a dedicated web resource. We provide information on how to access to results of primary and secondary analysis of the data, including comparative analysis with previously published populations from Ukraine, and populations from International Genome Sample Resource and Human Genome Diversity Project. The free research access to this database is contributing to growing understanding of human genetic diversity in Central Europe. This effort emphasizes the potential for reuse of the generated data, advocating for open access to support future research in genomics, bioinformatics, and personalized medicine.
The contribution of de novo coding mutations to meningomyelocele
Authors: Ha Yoo-Jin , Tang Isaac , Nisal Ashna , Jhamb Ishani , Wallace Cassidy , Schroeder Sarah , Lee Chanjae , Vong Keng loi , Meave Naomi , Jiwani Fiza , Barrows Chelsea , Lee Sangmoon , Jiang Nan , Patel Arzoo , Blanco Francisco A. , Yu Seyoung , Jeong Hui Su , Plutzer Isaac , Major Michael B. , Benoit Béatrice , Poüs Christian , Heffner Caleb , Kibar Zoha , Bot Gyang Markus , Northrup Hope , Au Kit Sing , Strain Madison , Ashley-Koch Allison , Finnell Richard H. , Le Joan T. , Meltzer Hal , Araujo Camila , Machado Helio R. , Stevenson Roger E. , Yurrita Anna , Mumtaz Sara , Mutchinick Osvaldo M. , Medina-Bereciartu José Ramón , Hildebrandt Friedhelm , Melikishvili Gia , Marwan Rony , Capra Valeria , Noureldeen Mahmoud M. , Salem Aida M.S. , Issa Mahmoud Y. , Zaki Maha S. , Lee Ji Eun , Alkelai Anna , Shuldiner Alan R. , Kingsmore Stephen F. , Murray Stephen A. , Gee Heon Yung , Miller W. Todd , Tolias Kimberley F. , Wallingford John B. ,   Spina Bifida Sequencing Consortium, Kim Sangwoo , Gleeson Joseph G.
Published: March 2024

Meningomyelocele (MM) is considered a genetically complex disease resulting from failure of neural tube closure (NTD). Patients display neuromotor disability and frequent hydrocephalus requiring ventricular shunting. A few proposed genes contribute to disease susceptibility, but most risk remains unexplained1. We postulated that de novo mutations (DNMs) under purifying selection contribute to MM risk2. Here we recruited a cohort of 851 MM trios requiring shunting at birth, compared with 732 control trios, and found that de novo likely gene disrupting or damaging missense mutations occur in approximately 22.3% of subjects, 28% of which are estimated to contribute to disease risk. The 187 genes with damaging DNMs collectively define networks including actin cytoskeleton and microtubule-based processes, axon guidance, and histone modification. Gene validation demonstrates partial or complete loss of function, impaired signaling and defective neural tube closure inXenopusembryos. Our results suggest DNMs make key contributions to MM risk, and highlight critical pathways required for neural tube closure in human embryogenesis.
Glutamine addiction is targetable via altering splicing of nutrient sensors and epitranscriptome regulators
Authors: Bojko Jodie , Kollareddy Madhu , Szemes Marianna , Bellamy Jacob , Poon Evon , Moukachar Ahmad , Legge Danny , Vincent Emma E , Jones Nicholas , Malik Sally , Greenhough Alex , Paterson Alex , Park Ji Hyun , Gallacher Kelli , Chesler Louis , Malik Karim
Published: March 2024

About 50% of poor prognosis neuroblastoma arises due to MYCN over-expression. We previously demonstrated that MYCN and PRMT5 proteins interact and PRMT5 knockdown led to apoptosis of MYCN amplified (MNA) neuroblastoma. Here we evaluate PRMT5 inhibitors GSK3203591/GSK3326593 as targeted therapeutics for MNA neuroblastoma and show MYCN-dependent growth inhibition and apoptosis. RNAseq revealed dysregulated MYCN transcriptional programmes and altered mRNA splicing, converging on key regulatory pathways such as DNA damage response, epitranscriptomics and cellular metabolism. Metabolic tracing showed glutamine metabolism was impeded following GSK3203591 treatment, which disrupted the MLX/Mondo nutrient sensors via intron retention of MLX mRNA. Glutaminase (GLS) protein was decreased by GSK3203591 despite unchanged transcript levels, suggesting post-transcriptional regulation. We demonstrate the RNA methyltransferase METTL3 and cognate reader YTHDF3 proteins are lowered following splicing alterations; accordingly, we observed hypomethylation of GLS mRNA and decreased GLS following YTHDF3 knockdown. In vivo efficacy of GSK3326593 was confirmed by increased survival ofTh-MYCNmice together with splicing events and protein decreases consistent with in vitro data. Our study supports the spliceosome as a key vulnerability of MNA neuroblastoma and rationalises PRMT5 inhibition as a targeted therapy.

GRAPHICAL ABSTRACT
Machine Learning Classification of 53BP1 Foci
Authors: Benítez-Jones María Xóchitl , Keegan Sarah , Jamshahi Sebastian , Fenyö David
Published: March 2024

Background

53BP1 foci are reflective of DNA double-strand break formation and have been used as radiation markers. Manual focus counting, while prone to bias and time constraints, remains the most accurate mode of detecting 53BP1 foci. Several studies have pursued automated focus detection to replace manual methods. Deep learning, spatial 3D images, and segmentation techniques are main components of the highest performing automated methods. While these approaches have achieved promising results regarding accurate focus detection and cell classification, they are not compatible with time-sensitive large-scale applications due to their demand for long run times, advanced microscopy, and computational resources. Further, segmentation of overlapping foci in 2D images has the potential to represent focus morphologies inaccurately.

Results

To overcome these limitations, we developed a novel method to classify 2D fluorescence microscopy images of 53BP1 foci. Our approach consisted of three key features: (1) general 53BP1 focus classes, (2) varied parameter space composed of properties from individual foci and their respective Fourier transform, and (3) widely-available machine learning classifiers. We identified four main focus classes, which consisted of blurred foci and three levels of overlapping foci. Our parameter space for the training focus library, composed of foci formed by fluorescently-tagged BP1-2, showed a wide correlation range between variables which was validated using a publicly-available library of immunostained 53BP1 foci. Random forest achieved one of the highest and most stable performances for binary and multiclass problems, followed by a support vector machine and k-nearest neighbors. Specific metrics impacted the classification of blurred and low overlap foci for both train and test sets.

Conclusions

Our method classified 53BP1 foci across separate fluorescent markers, resolutions, and damage-inducing methods, using off-the-shelf machine learning classifiers, a diverse parameter space, and well-defined focus classes.
Karyotype Evolution in Response to Chemoradiotherapy and Upon Recurrence of Esophageal Adenocarcinomas
Authors: van der Sluis K. , van Sandick J. W. , Koemans W. J. , van den Bosch T. , Broeks A. , Peters D. , Seignette I. M. , Rausch C. R. , van Dijk E. , Snaebjornsson P. , van den Berg J. G. , van Grieken N. C. T. , Ylstra B. , Carvalho B. , Miedema D. M. , Kodach L. L.
Published: March 2024

The genome of esophageal adenocarcinoma (EAC) is highly unstable and might evolve over time. Here, we track karyotype evolution in EACs in response to treatment and upon recurrence through multi-region and longitudinal analysis. To this end, we introduce L-PAC, a bio-informatics technique that allows inference of absolute copy number aberrations (CNA) of low-purity samples by leveraging information of high-purity samples from the same cancer. Quantitative analysis of matched absolute CNAs reveals that the amount of karyotype evolution induced by chemoradiotherapy (CRT) is predictive for early recurrence and depends on the initial level of karyotype intra-tumor heterogeneity. We observe that CNAs acquired in response to CRT are partially reversed back to the initial state upon recurrence. CRT hence alters the fitness landscape to which tumors can adjust by adapting their karyotype. Together, our results indicate that karyotype plasticity contributes to therapy resistance of EACs.
Biotic and abiotic drivers of ecosystem functioning differ between a temperate and a tropical region
Authors: van Dijk Laura J. A. , Miraldo Andreia , Raharinjanahary Dimby , Rajoelison Eric Tsiriniaina , Fisher Brian L. , Goodsell Robert M. , Iwaszkiewicz-Eggebrecht Elzbieta , Åhlén David , Högvall Johanna , Lundberg Erika , Rova Emma , Łukasik Piotr , Ronquist Fredrik , Roslin Tomas , Tack Ayco J. M.
Published: March 2024

Any single ecosystem will provide many ecosystem functions. Whether these functions tend to increase in concert or trade off against each other is a question of much current interest. Equally topical are the drivers behind ecosystem function rates. Yet, we lack large-scale systematic studies that investigate how abiotic factors can directly or indirectly — via effects on biodiversity — drive ecosystem functioning. In this study, we assessed the impact of climate, landscape and biotic community on ecosystem functioning and multifunctioning in the temperate and tropical zone, and investigated potential trade-offs among ecosystem functions in both zones. To achieve this, we measured a diverse set of insect-related ecosystem functions — including herbivory, seed dispersal, predation, decomposition and pollination — at 50 sites across Madagascar and 171 sites across Sweden, and characterized the insect community at each site using Malaise traps. We used structural equations models to infer causality of the effects of climate, landscape, and biodiversity on ecosystem functioning. For the temperate zone, we found that abiotic factors were more important than biotic factors in driving ecosystem functioning, while in the tropical zone, effects of biotic drivers were most pronounced. In terms of trade-offs among functions, in the temperate zone, only seed dispersal and predation were positively correlated, while all other functions were uncorrelated. By contrast, in the tropical zone, most ecosystem functions increased in concert, highlighting that tropical ecosystems can simultaneously provide a diverse set of functions. These correlated functions in Madagascar could for the most part be explained by similar responses to local climate, landscape, and biota. Our study suggests that the functioning of temperate and tropical ecosystems differs fundamentally in patterns and drivers. Without a better understanding of these differences, it will be impossible to correctly predict shifts in ecosystem functioning in response to environmental disturbances. To identify global patterns and drivers of ecosystem functioning, we will next need replicate sampling across biomes – as here achieved for two regions, thus paving the road and setting the baseline expectations.
Super Pangenome of Grapevines Empowers Improvement of the Oldest Domesticated Fruit
Authors: Guo Li , Wang Xiangfeng , Ayhan Dilay Hazal , Rhaman Mohammad Saidur , Yan Ming , Jiang Jianfu , Wang Dongyue , Zheng Wei , Mei Junjie , Ji Wei , Jiao Jian , Chen Shaoying , Sun Jie , Yi Shu , Meng Dian , Wang Jing , Bhuiyan Mohammad Nasim , Qin Guochen , Guo Linling , Yang Qingxian , Zhang Xuenan , Sun Haisheng , Liu Chonghuai , Ye Wenxiu
Published: March 2024

Grapevine (Vitis) is the oldest domesticated fruit crop with great cultural and economic importance. Here, we assemble and annotate haplotype-resolved genomes of 72 Vitis accessions including 25 wild and 47 cultivated grapevines, and a haplotype-resolved complete genome of V. vinifera. Coalescent phylogenomics of 142 haplotype genomes disentangles the mysterious hybridization history of grapevines, revealing enormous genetic diversity among species. Pangenome analysis together with phenotyping data reveals that European cultivars, more susceptible to the most destructive disease downy mildew (DM), had a smaller repertoire of disease resistance genes of NLR family. Through extensive structural variation (SV) characterization, phenotyping, transcriptome profiling of 113 Vitis accessions, and SV-eQTL analysis, we have identified over 79 SVs and their relevant genes significantly associated with DM resistance, exemplified by a lysine histidine transporter, VvLHT8. This haplotype-resolved complete genome and pangenome of Vitis genus will accelerate grapevine breeding and enrich our understanding of the evolution and biology of grapevines.
MAssively-Parallel Flow cytometry Xplorer (MAPFX): A Toolbox for Analysing Data from the Massively-Parallel Cytometry Experiments
Authors: Liao Hsiao-Chi , Speed Terence P. , McCarthy Davis J. , Salim Agus
Published: March 2024

Massively-Parallel Cytometry (MPC) experiments allow cost-effective quantification of more than 200 surface proteins at single-cell resolution. The Infinity Flow (Inflow) analysis protocol was developed to measure highly informative protein ‘backbone’ markers on all cells in all wells distributed across three 96-well plates, along with well-specific exploratory protein ‘infinity’ markers. Backbone markers can be used to impute the infinity markers on cells in all other wells using machine learning methods. This protocol offers unprecedented opportunities for more comprehensive classification of cell types. However, some aspects of the protocol can be improved, including methods for background correction and removal of unwanted variation. Here, we proposeMAPFXas an end-to-end toolbox that carefully pre-processes the raw data from MPC experiments, and further imputes the ‘missing’ infinity markers in the wells without those measurements. Our pipeline starts by performing background correction on raw intensities to remove the noise from electronic baseline restoration and fluorescence compensation by adapting a normal-exponential convolution model. Unwanted technical variation, from sources such as well effects, is then removed using a log-normal model with plate, column, and row factors, after which infinity markers are imputed using the informative backbone markers as predictors. The completed dataset can then be used for clustering and other statistical analyses. Unique features of our approach include performing background correction prior to imputation and removing unwanted variation from the data at the cell-level, while explicitly accounting for the potential association between biology and unwanted factors. We benchmark our pipeline against alternative pipelines and demonstrate that our approach is better at preserving biological signals, removing unwanted variation, and imputing unmeasured infinity markers.
Estimation of the Achilles tendon twist in vivo by individual triceps surae muscle stimulation
Authors: Laura Lecompte , Marion Crouzier , Stéphane Baudry , Benedicte Vanwanseele
Published: March 2024

The Achilles tendon (AT) is comprised of three distinct subtendons, each arising from the one of the three heads of the triceps surae muscles: gastrocnemius medialis (GM), gastrocnemius lateralis (GL) and soleus (SOL). These subtendons exhibit a twisted structure, classified as low (Type I), medium (Type II), and high (Type III) twist, based on cadaveric studies. Nevertheless, the in-vivo investigation of AT twist is notably scarce, resulting in a limited understanding of its functional significance. The aim of this study was to give insights into the complex 3D AT structure in vivo. 30 healthy participants underwent individual stimulation of each of the triceps surae muscles at rest with the foot attached to the pedal of an isokinetic dynamometer. Ultrasound images were captured to concomitantly examine the displacement of the superficial, middle and deep AT layers. SOL stimulation resulted in the highest AT displacement followed by GM and GL stimulation. Independent of the muscle stimulated, non-uniformity within the AT was observed with the deep layer exhibiting more displacement compared to the middle and superficial layers, hence important inter-individual differences in AT displacement were noticeable. By leveraging these individual displacement patterns during targeted stimulations in conjunction with cadaveric twist classifications providing insights into the area of each specific subtendon, our classification identified 19 subjects with a ’low’ and 11 subjects with a ’high’ AT twist. More research is needed to understand the complexity of the AT twisted structure in vivo to further understand its effect on AT properties and behaviour.
Transfer learning of multicellular organization via single-cell and spatial transcriptomics
Authors: Tan Yecheng , Wang Ai , Wang Zezhou , Lin Wei , Yan Yan , Nie Qing , Shi Jifan
Published: March 2024

Spatial tissues exhibit complex gene expression and multicellular patterns that are difficult to dissect. Single-cell RNA sequencing (scRNA-seq) provides full coverages of genes, but lacking spatial information, whereas spatial transcriptomics (ST) measures spatial locations of individual or group of cells, with more restrictions on gene information. To integrate scRNA-seq and ST data, we introduce a transfer learning method to decipher spatial organization of cells named iSORT. iSORT trains a neural network that maps gene expressions to spatial locations using scRNA-seq data along with ST slices as references. iSORT can find spatial patterns at single-cell scale, identify key genes that drive the patterning, and infer pseudo-growth trajectories using a concept of SpaRNA velocity. Benchmarking on simulation data and comparing with multiple existing tools show iSORT’s robustness and accuracy in reconstructing spatial organization. Using our own new human artery datasets, iSORT shows its capability of dissecting atherosclerosis. Applications to a range of biological systems, such as mouse embryo, mouse brain,Drosophilaembryo, and human developmental heart, demonstrate that iSORT can utilize both scRNA-seq and ST datasets to uncover multilayer spatial information of single cells.
Expansion microscopy for super-resolution imaging of collagen-abundant tissues
Authors: Chuang Ya-Han , Wu Yueh-Feng , Lin Ya-Hui , Zhou Yu-Xian , Hsu Shao-Chun , Lin Sung-Jan , Chu Li-An
Published: March 2024

Expansion microscopy (ExM) is popular for three-dimensional ultrastructural imaging of cultured cells and tissue slices at nanoscale resolution with conventional microscopes via physical expansion of biological tissues. However, the application of this technology to collagen-abundant thick tissues is challenging. We demonstrate a new method, collagen expansion microscopy (ColExM), optimized for expanding tissues containing more than 70% collagen. ColExM succeeded in 4.5-fold linear expansion with minimal structural distortion of corneal and skin tissues. It was also compatible with immunostaining, allowing super-resolution visualization of three-dimensional neural structures innervating hair follicles and corneas. With ColExM, we succeeded in identifying individual mitochondria and previously unrecognized dendritic spine-like structures of corneal nerves. ColExM also enabled fine mapping of structural rearrangement of tight junctions and actin cytoskeletons. Therefore, this method can facilitate the exploration of three-dimensional nanoscale structures in collagen-rich tissues.
Breaking the Burst: Unveiling Mechanisms Behind Fragmented Network Bursts in Patient-derived Neurons
Authors: Doorn Nina , Voogd Eva J.H.F. , Levers Marloes R. , van Putten Michel J.A.M. , Frega Monica
Published: March 2024

Fragmented network bursts (NBs) are observed as a phenotypic driver in many patient-derived neuronal networks on multi-electrode arrays (MEAs), but the pathophysiological mechanisms underlying this phenomenon are unknown. Here, we used our previously developed biophysically detailedin silicomodel to investigate these mechanisms. Fragmentation of NBs in our model simulations occurred only when the level of short-term synaptic depression (STD) was enhanced, suggesting that STD is a key player. Experimental validation with Dynasore, an STD enhancer, induced fragmented NBs in healthy neuronal networksin vitro. Additionally, we showed that strong asynchronous neurotransmitter release, NMDA currents, or short-term facilitation (STF) can support the emergence of multiple fragments in NBs by producing excitation that persists after high-frequency firing stops. Our results provide important insights into disease mechanisms and potential pharmaceutical targets for neurological disorders modeled using hiPSC-derived neurons.
Exploring the potential ofCeltis aetnensis compoundsthrough Molecular Docking for Hepatocellular carcinoma
Authors: Kavitha K.N. , Revathi K. , Tamiraikani Tamilamban
Published: March 2024

Hepatocellular carcinoma (HCC) accounts for 1% of the liver malignances; HCC progresses in hepatocytes primarily as a result of inflammation, oxidative stress, and primary liver disease. Thanks to the in-silico methods that help in the identification of the targets and the drugs/potential drugs that can inhibit these proteins that cater to the growth of the cancer cells. Plant actives have been long considered as potential sources of anticancer drugs. In this regard, we studied the actives fromCeltis tournefortii Lam (Celtis aetnensis) usingGCMS analysis. The protein structures of three receptors—NF-B P50 homodimer, FGF receptor 4, and vascular endothelial growth factor receptor 2 (VEGFR2)—were then subjected to docking studies using phytocompounds fromCeltis aetnensis. Three receptor proteins were used in the docking analysis against 7 ligands. In this study, the PDB was used to obtain the structures of 3 cancer-related receptor proteins. The proteins were produced by eliminating water molecules, ligands using PyMol, and were then exposed to docking experiments. Docking studies revealed that the compounds with a binding energy ranging from -1.83 kcal/mol to -6.12 kcal/mol. The docking results revealed that eicosatetraenoic acid has a binding energy of-6.12 kcal/mol against the FGFR4 receptor. Sulfadoxine’s binding energy to the VEGFR2 receptor protein was-4.9 kcal/mol. Promethazine sulfoxide docks with an energy of -5.36 kcal/mol against NFKB P50. By attaching to the protein, these substances demonstrated good inhibitory activity. The results are supportive that these compounds may be used to treat HCC.
Light-directed evolution of dynamic, multi-state, and computational protein functionalities
Authors: Gligorovski Vojislav , Labagnara Marco , Rahi Sahand Jamal
Published: March 2024

Directed evolution is a powerful method in biological engineering. Current approaches draw on time-invariant selection mechanisms, ideal for evolving steady-state properties such as enzymatic activity or fluorescence intensity. A fundamental problem remains how to continuously evolve dynamic, multi-state, or computational functionalities, e.g., on-off kinetics, state-specific activity, stimulus-responsiveness, or switching and logic capabilities. These require selection pressure on all of the states of a protein of interest (POI) and the transitions between them. We realized that optogenetics and cell cycle oscillations could be leveraged for a novel directed evolution paradigm (‘optovolution’) that is germane for this need: We designed a signaling cascade in budding yeast where optogenetic input switches the POI between off (0) and on (1) states. In turn, the POI controls a Cdk1 cyclin, which in the re-engineered cell cycle system is essential for one cell cycle stage but poisonous for another. Thus, the cyclin must oscillate (1-0-1-0…) for cell proliferation. In this system, evolution can act efficiently on the dynamics, transient states, and input-output relations of the POI in every cell cycle. Further, controlling the pacemaker, light, directs and tunes selection pressures. Optovolution is in vivo, continuous, self-selecting, and efficient. We first evolved two optogenetic systems, which relay 0/1 input to 0/1 output: We obtained 25 new variants of the widely used LOV transcription factor El222 that were stronger, less leaky, or green-and red-responsive. The latter was conjectured to be impossible for LOV domains but is needed for multiplexing and lowering phototoxicity. Evolving the PhyB-Pif3 optogenetic system, we discovered that loss of YOR1 makes supplementing the expensive and unstable chromophore phycocyanobilin (PCB) unnecessary. Finally, we demonstrate the generality of the method by evolving a destabilized rtTA transcription factor, which performs an AND operation between transcriptional and doxycycline input. Optovolution makes coveted, difficult-to-change protein functionalities continuously evolvable.
Next Generation Neuropeptide Y Receptor Small Molecule Agonists Inhibit Mosquito Biting Behavior
Authors: Zeledon Emely V. , Baxt Leigh A. , Khan Tanweer A. , Michino Mayako , Miller Michael , Huggins David J. , Jiang Caroline S. , Vosshall Leslie B. , Duvall Laura B.
Published: March 2024

FemaleAedes aegyptimosquitoes can spread disease-causing pathogens when they bite humans to obtain blood nutrients required for egg production. Following a complete blood meal, host-seeking is suppressed until eggs are laid. Neuropeptide Y-like Receptor 7 (NPYLR7) plays a role in endogenous host-seeking suppression and previous work identified small molecule NPYLR7 agonists that suppress host-seeking and blood feeding when fed to mosquitoes at high micromolar doses. Using structure activity relationship analysis and structure-guided design we synthesized 128 compounds with similarity to known NPYLR7 agonists. Althoughin vitropotency (EC50) was not strictly predictive ofin vivoeffect, we identified 3 compounds that suppressed blood feeding from a live host when fed to mosquitoes at a 1 μM dose, a 100-fold improvement over the original reference compound. Exogenous activation of NPYLR7 represents an innovative vector control strategy to block mosquito biting behavior and prevent mosquito/human host interactions that lead to pathogen transmission.
Quantitative profiling of human translation initiation reveals regulatory elements that potently affect endogenous and therapeutically modified mRNAs
Authors: Lewis Cole J.T. , Xie Li , Bhandarkar Shivani , Jin Danni , Abdallah Kyrillos S. , Draycott Austin S. , Chen Yixuan , Thoreen Carson C. , Gilbert Wendy V.
Published: March 2024

mRNA therapeutics offer a potentially universal strategy for the efficient development and delivery of therapeutic proteins. Current mRNA vaccines include chemically modified nucleotides to reduce cellular immunogenicity. Here, we develop an efficient, high-throughput method to measure human translation initiation on therapeutically modified as well as endogenous RNAs. Using systems-level biochemistry, we quantify ribosome recruitment to tens of thousands of human 5′ untranslated regions and identify sequences that mediate 250-fold effects. We observe widespread effects of coding sequences on translation initiation and identify small regulatory elements of 3-6 nucleotides that are sufficient to potently affect translational output. Incorporation of N1-methylpseudouridine (m1Ψ) selectively enhances translation by specific 5′ UTRs that we demonstrate surpass those of current mRNA vaccines. Our approach is broadly applicable to dissect mechanisms of human translation initiation and engineer more potent therapeutic mRNAs.

Highlights

Measurement of >30,000 human 5′ UTRs reveals a 250-fold range of translation output

Systematic mutagenesis demonstrates the causality of short (3-6nt) regulatory elements

N1-methylpseudouridine alters translation initiation in a sequence-specific manner

Optimal modified 5′ UTRs outperform those in the current class of mRNA vaccines