bluuebunny/crossref_metadata_2025 huggingface dataset:
Echolocating bats prefer a high risk-high gain foraging strategy to increase prey profitability
Authors: Stidsholt Laura , Hubancheva Antoniya , Greif Stefan , Goerlitz Holger R. , Johnson Mark , Yovel Yossi , Madsen Peter T.
Published: October 2022

Most bats catch nocturnal prey during active flight guided by echolocation but some species depart from this ancestral behaviour to capture ground prey using passive listening. Here, we explore the costs and benefits of these hunting transitions by combining high-resolution biologging data and DNA metabarcoding to quantify the relative contributions of aerial and ground prey to the total food intake of wild greater mouse-eared bats. We show that these bats use both foraging strategies with similar average nightly captures of 25 small, aerial insects and 30 large, ground-dwelling insects per bat, but with higher capture success in air (78 % in air vs 30 % on ground). However, owing to the 3 to 20 times heavier ground prey, 85 % of the estimated nightly food acquisition comes from ground prey despite the 2.5 times higher failure rates. Further, we find that most bats use the same foraging strategy on a given night suggesting that bats adapt their hunting behaviour to weather and ground conditions. We conclude that prey switching matched to environmental dynamics plays a key role in covering the energy intake even in specialised predators.
Technical report on best practices for hybrid and long readde novoassembly of bacterial genomes utilizing Illumina and Oxford Nanopore Technologies reads
Authors: Hackl Simon T. , Harbig Theresa A. , Nieselt Kay
Published: October 2022

The emergence of commercial long read sequencing technologies in the 2010s and the concomitant development of new bioinformatics tools bears the potential ofde novogenome assemblies of unprecedented contiguity and quality. However, until today these novel technologies suffer from high rates of sequencing errors. These may be overcome by using long and short reads in combination, in so called hybrid approaches, or by increasing the through-put and thereby the coverage of sequencing runs. In particular the latter will thereby increase the cost of the assembly inevitably. Herein, to-date long read and hybrid assemblers were tested on real whole genome sequencing Illumina and Oxford Nanopore Technologies read data sets and sub samples of these in order to elaborate a best practice forde novoassembly. The findings suggest that although long reads alone can be used to reconstruct complete and contiguous genomes, in particular the single-nucleotide and indel error rate remains high compared to hybrid approaches and that this can impact downstream applications such as variation discovery and gene prediction negatively.
The role of hydrodynamic flow in the self-assembly of dragline spider silk proteins
Authors: Herrera-Rodríguez Ana M. , Dasanna Anil Kumar , Daday Csaba , Cruz-Chú Eduardo R. , Aponte-Santamaría Camilo , Schwarz Ulrich S. , Gräter Frauke
Published: October 2022

Hydrodynamic flow in the spider duct induces conformational changes in dragline spider silk proteins (spidroins) and drives their assembly, but the underlying physical mechanisms are still elusive. Here we address this challenging multiscale problem with a complementary strategy of atomistic and coarse-grained Molecular Dynamics (MD) simulations with uniform flow. The conformational changes at the molecular level were analyzed for single tethered spider silk peptides. Uniform flow leads to coiled-to-stretch transitions and pushes alanine residues intoβ-sheet and Poly-Proline II (PPII) conformations. Coarse-grained simulations of the assembly process of multiple semi-flexible block copolymers using multi-particle collision dynamics reveal that the spidroins aggregate faster but into low-order assemblies when they are less extended. At medium-to-large peptide extensions (50%-80%), assembly slows down and becomes reversible with frequent association and dissociation events, while spidroin alignment increases and alanine repeats form ordered regions. Our work highlights the role of flow in guiding silk self-assembly into tough fibers by enhancing alignment and kinetic reversibility, a mechanism likely relevant for other proteins whose function depends on hydrodynamic flow.
Overproduction of Native and Click-able Colanic Acid Slime from EngineeredEscherichia coli
Authors: Sadler Joanna C. , Brewster Richard C. , González Alba F. , Nirkko Jessica S. , Varzandeh Simon , Wallace Stephen
Published: October 2022

The fundamental biology and application of bacterial exopolysaccharides is gaining increasing attention. However, current synthetic biology efforts to produce the major component ofEscherichia sp. slime, colanic acid, and functional derivatives thereof have been limited. Herein, we report the overproduction of colanic acid (up to 1.32 g/L) from D-glucose in an engineered strain ofE. coliJM109. Furthermore, we report that chemically-synthesized L-fucose analogues containing an azide motif can be metabolically incorporated into the slime layer via a heterologous fucose salvage pathway fromBacteroides sp. and used in a click reaction to attach an organic cargo to the cell surface. This molecular engineered bio-polymer possesses enormous potential as a new tool for use in chemical, biological and materials research.
Rugged fitness landscapes minimize promiscuity in the evolution of transcriptional repressors
Authors: Meger Anthony T. , Spence Matthew A. , Sandhu Mahakaran , Jackson Colin J. , Raman Srivatsan
Published: October 2022

How a protein’s function influences the shape of its fitness landscape, smooth or rugged, is a fundamental question in evolutionary biochemistry. Smooth landscapes arise when incremental mutational steps lead to a progressive change in function, as commonly seen in enzymes and binding proteins. On the other hand, rugged landscapes are poorly understood because of the inherent unpredictability of how sequence changes affect function. Here, we experimentally characterize the entire sequence phylogeny, comprising 1158 extant and ancestral sequences, of the DNA-binding domain (DBD) of the LacI/GalR transcriptional repressor family. Our analysis revealed an extremely rugged landscape with rapid switching of specificity even between adjacent nodes. Further, the ruggedness arises due to the necessity of the repressor to simultaneously evolve specificity for asymmetric operators and disfavors potentially adverse regulatory crosstalk. Our study provides fundamental insight into evolutionary, molecular, and biophysical rules of genetic regulation through the lens of fitness landscapes.
SETD2 safeguards the genome against isochromosome formation
Authors: Mason Frank M. , Kounlavong Emily S. , Tebeje Anteneh T. , Dahiya Rashmi , Guess Tiffany , Vlach Logan , Norris Stephen R. , Lovejoy Courtney A. , Dere Ruhee , Ohi Ryoma , Ly Peter , Walker Cheryl L. , Rathmell W. Kimryn
Published: October 2022

Factors governing the faithful replication of chromosomes are essential for cellular and genomic integrity. While a variety of mechanisms to manage breaks and promote repair of DNA are widely recognized, epigenetic landmarks that preserve telomere-to-telomere replication fidelity and prevent genome instability are not well-understood. SETD2 is the histone methyltransferase responsible for trimethylation on histone H3 lysine 36 and is newly recognized as a tumor suppressor that acts to maintain genome stability. Importantly, SETD2 is frequently lost in cancers that exhibit extensive intratumoral heterogeneity. Here, we demonstrate that loss ofSETD2and H3K36me3 promotes chromosome segregation errors and DNA bridging during mitosis, and that these bridges are driven by the formation of dicentric chromosomes. Cytogenetic analyses revealed that these chromosomes were comprised of mirror-imaged isochromosomes and isodicentric chromosomes that contain two active centromeres. These data demonstrate that the SETD2 histone methyltransferase is essential to prevent a palindromic replication intermediate, whose loss precipitates the formation of a mutable chromatin structure known to initiate a cascade of genomic instability in cancer.
Gating interactions steer loop conformational changes in the active site of the L1 metallo-β-lactamase
Authors: Zhao Zhuoran , Shen Xiayu , Chen Shuang , Gu Jing , Wang Huan , Mojica Maria F. , Samanta Moumita , Bhowmik Debsindhu , Vila Alejandro J. , Bonomo Robert A. , Haider Shozeb
Published: October 2022

β-lactam antibiotics are the most important and widely used antibacterial agents across the world. However, the widespread dissemination of β-lactamases among pathogenic bacteria limits the efficacy of β-lactam antibiotics. This has created a major public health crisis. The use of β-lactamase inhibitors has proven useful in restoring the activity of β-lactam antibiotics, yet, effective clinically approved inhibitors against class B metallo-β-lactamases (MBLs) are not available. L1, a class B3 enzyme expressed byStenotrophomonas maltophilia, is a significant contributor to the β-lactam resistance displayed by this opportunistic pathogen. Structurally, L1 is a tetramer with two elongated loops, α3-β7 and β12-α5, present around the active site of each monomer. Residues in these two loops influence substrate/inhibitor binding. To study how the conformational changes of the elongated loops affect the active site in each monomer, enhanced sampling molecular dynamics (MD) simulations were performed, Markov State Models (MSM) were built, and convolutional variational autoencoder (CVAE)-based deep learning was applied. The key identified residues (D150a, H151, P225, Y227, R236) were mutated and the activity of the generated L1 variants was evaluated in cell-based experiments. The results demonstrate that there are extremely significant gating interactions between α3-β7 and β12-α5 loops. Taken together, the gating interactions with the conformational changes of the key residues play an important role in the structural remodeling of the active site. These observations offer insights into the potential for novel drug development exploiting these gating interactions.
Characterization and utility of novel monoclonal antibodies to cholera toxin B subunit
Authors: Garcia Noel Verjan , Celis Ian Carlosalberto Santisteban , Dent Matthew , Matoba Nobuyuki
Published: October 2022

Cholera toxin B subunit (CTB) is a potent immunomodulator exploitable in mucosal vaccine and immunotherapeutic development. To aid in the characterization of pleiotropic biological functions of CTB and its variants, we generated a panel of anti-CTB monoclonal antibodies (mAbs). By ELISA and surface plasmon resonance, two mAbs, 7A12B3 and 9F9C7, were analyzed for their binding affinities to cholera holotoxin (CTX), CTB, and EPICERTIN: a recombinant CTB variant possessing mucosal healing activity. Both 7A12B3 and 9F9C7 bound efficiently to CTX, CTB, and EPICERTIN with equilibrium dissociation constants at low to sub-nanomolar concentrations but bound weakly, if at all, toEscherichia coliheat-labile enterotoxin B subunit. In a cyclic adenosine monophosphate (cAMP) assay using Caco2 human colon epithelial cells, the 7A12B3 mAb was found to be a potent inhibitor of CTX, whereas 9F9C7 had relatively weak inhibitory activity. Meanwhile, the 9F9C7 mAb effectively detected CTB and EPICERTIN bound to the surface of Caco2 cells and mouse spleen leukocytes by flow cytometry. Using 9F9C7 in immunohistochemistry, we confirmed the preferential localization of EPICERTIN in colon crypts following oral administration of the protein in mice. Collectively, these mAbs provide valuable tools to investigate the biological functions and preclinical development of CTB variants.
Time in allopatry does not predict the outcome of secondary contact in lowland Panamanian birds
Authors: Mclaughlin Jessica F. , Miller Matthew J.
Published: October 2022

Geographic speciation models assume that time in allopatry should result in greater reproductive isolation between populations. Here we test the prediction that greater time in allopatry results in greater reproductive isolation using comparative ultraconserved element (UCE) data from ten bird lineages in secondary contact in Panama, measuring both genome-wide divergence and the geographic extent of hybridization. The best-fit models for the proportion of fixed Z-linked and autosomal loci to our data includes a combination of both time (as measured by mtDNA divergence) and hand-wing index, emphasizing that the role of time is tempered by dispersal capability. Furthermore, time does not predict the extent of genome-wide introgression as measured by the median width of diagnostic loci clines or the degree of variation in cline centers or widths. These metrics of the outcome of secondary contact were best predicted by ecological and genomic factors, including diet, hand-wing index, and genome-wideFSTrespectively, that are understood to serve as proxies for dispersal, the variability of population size, and overall genomic divergence. We find a primary role for ecological factors instead of isolation time in determining secondary contact outcomes for a lineage, highlighting how ecology shapes the development of reproductive isolation.
Dissecting the effects of GTPase and kinase domain mutations on LRRK2 endosomal localization and activity
Authors: Rinaldi Capria , Waters Christopher S. , Kumbier Karl , Rao Lee , Nichols R. Jeremy , Jacobson Matthew P. , Wu Lani F. , Altschuler Steven J.
Published: October 2022

Parkinson’s disease-causing LRRK2 mutations lead to varying degrees of Rab GTPase hyperphosphorylation. Puzzlingly, LRRK2 GTPase-inactivating mutations—which do not affect intrinsic kinase activity—lead to higher levels of cellular Rab phosphorylation than kinase-activating mutations. Here, we investigated whether mutation-dependent differences in LRRK2 cellular localization could explain this discrepancy. We discovered that blocking endosomal maturation leads to the rapid formation of mutant LRRK2+endosomes on which LRRK2 phosphorylates substrate Rabs. LRRK2+endosomes are maintained through positive feedback, which mutually reinforces membrane localization of LRRK2 and phosphorylated Rab substrates. Furthermore, across a panel of mutants, cells expressing GTPase-inactivating mutants formed strikingly more LRRK2+endosomes than cells expressing kinase-activating mutants, resulting in higher total cellular levels of phosphorylated Rabs. Our study suggests that an increased probability of LRRK2 GTPase-inactivating mutants to be retained on intracellular membranes over the kinase-activating mutants leads to higher substrate phosphorylation.
Lymph node expansion predicts magnitude of vaccine immune response
Authors: Najibi Alexander J. , Lane Ryan S. , Sobral Miguel C. , Freedman Benjamin R. , Estupinan Joel Gutierrez , Elosegui-Artola Alberto , Tringides Christina M. , Dellacherie Maxence O. , Williams Katherine , Müller Sören , Turley Shannon J. , Mooney David J.
Published: October 2022

Lymph nodes (LNs) dynamically expand in response to immunization, but the relationship between LN expansion and the accompanying adaptive immune response is unclear. Here, we first characterized the LN response across time and length scales to vaccines of distinct strengths. High-frequency ultrasound revealed that a bolus ‘weak’ vaccine induced a short-lived, 2-fold volume expansion, while a biomaterial-based ‘strong’ vaccine elicited an ∼7-fold LN expansion, which was maintained several weeks after vaccination. This latter expansion was associated with altered matrix and mechanical properties of the LN microarchitecture. Strong vaccination resulted in massive immune and stromal cell engagement, dependent on antigen presence in the vaccine, and conventional dendritic cells and inflammatory monocytes upregulated genes involved in antigen presentation and LN enlargement. The degree of LN expansion following therapeutic cancer vaccination strongly correlated with vaccine efficacy, even 100 days post-vaccination, and direct manipulation of LN expansion demonstrated a causative role in immunization outcomes.
A contamination focused approach for optimizing the single-cell RNA-seq experiment
Authors: Arceneaux Deronisha , Chen Zhengyi , Simmons Alan J. , Heiser Cody N. , Southard-Smith Austin N. , Brenan Michael J. , Yang Yilin , Chen Bob , Xu Yanwen , Choi Eunyoung , Campbell Joshua D. , Liu Qi , Lau Ken S.
Published: October 2022

Achieving high data quality in single-cell RNA-seq (scRNA-seq) experiments has always been a significant challenge stemming from minute signal that can be detected in individual cells. Droplet-based scRNA-seq additionally suffers from ambient contamination, comprising nucleic acid materials released by dead cells into the loading buffer and co-encapsulated with real cells, which further washes out real biological signals. Here, we developed quantitative, ambient contamination-based metrics and an associated software package that can both evaluate current datasets and guide new experimental optimizations. We performed a series of experimental optimizations using the inDrops platform to address the mechanical and microfluidic cell encapsulation aspect of an scRNA-seq experiment, with a focus on minimizing ambient contamination. We report improvements that can be achieved via cell fixation, microfluidic loading, microfluidic dilution, and nuclei versus cell preparation; many of these parameters are inaccessible on commercial platforms. We provide insights into previously obscured factors that can affect scRNA-seq data quality and suggest mitigation strategies that can guide future experiments.
Metabolic modeling ofHermetia illucenslarvae resource allocation for high-value fatty acid production
Authors: Grausa Kristina , Siddiqui Shahida A , Lameyer Norbert , Wiesotzki Karin , Smetana Sergiy , Pentjuss Agris
Published: October 2022

All plant and animal kingdom organisms use highly connected biochemical networks to facilitate sustaining, proliferation and growth functions. While biochemical network details are well known, the understanding of intense regulation principles is still limited.

We chose to investigateHermetia illucensfly at the larval stage as it is crucial for successful resource accumulation and allocation for the consequential organism’s developmental stages. We combined the iterative wet lab experiments and innovative metabolic modeling design approaches, to simulate and explain theH. illucenslarval stage resource allocation processes and biotechnology potential. We performed time-based growth and high-value chemical compound accumulation wet lab chemical analysis experiments in larvae and Gainesville diet composition. To predict diet-based alterations on fatty acid allocation potential, we built and validated the firstH. illucensmedium-size stoichiometric metabolic model.

Using optimization methods like Flux balance and Flux variability analysis on the novel insect metabolic model, it predicted that doubled essential amino acid consumption increased the growth rate by 32%, but pure glucose consumption had no positive impact on growth. In the case of doubled pure valine consumption, the model predicted a 2% higher growth rate. In this study, we describe a new framework to research the impact of dietary alterations on the metabolism of multi-cellular organisms at different developmental stages for improved, sustainable and directed high-value chemicals.

Significance Statement

Metabolic modeling serves as a platform for researchers to investigate and study in depth the possible states of the system based on the existing knowledgebase (e.g. metabolic reactions, substrates, products and their stoichiometry). These models can be applied for different industrial applications, to simulate resource allocation potential and growth conditions. Moreover, these models predict the required diet for living organisms and insects to improve survival and growth rates and accumulate higher-value products, like fatty acids.
Transcription factors use a unique combination of cofactors to potentiate different promoter-dependent steps in transcription
Authors: Bell Charles C , Talarmain Laure , Scolamiero Laura , Lam Enid YN , Ang Ching-Seng , Gilan Omer , Dawson Mark A
Published: October 2022

Transcription factors use DNA binding domains to recognise specific sequences and transactivation domains to recruit the cofactor proteins necessary for transcription. However, how specific cofactors contribute to transactivation at different genes remains unclear. Here, we couple Gal4-transactivation assays with comparative CRISPR-Cas9 screens to identify the cofactors required by nine different transcription factors and nine different core promoters in human cells. We classify cofactors as ubiquitous or specific, discover novel transcriptional co-dependencies and demonstrate that submodules within large co-activator complexes, such as the tail 2 and kinase modules of Mediator, facilitate transcriptional elongation. Rather than displaying discrete mechanisms of action, we discover that each TF requires a unique combination of cofactors, which influence its ability to potentiate distinct steps in the transcriptional process. Our findings help reconcile models of cofactor-promoter compatibility by demonstrating that transcription at different classes of promoters is constrained by either initiation or pause release. These differences dictate cofactor compatibility and the dynamic range of gene expression. Overall, our screens provide insight into TF-cofactor relationships and their ability to potentiate different steps in transcription at different classes of promoters.
Gradual cerebral hypoperfusion in a knock-in mouse model of Alzheimer’s disease triggers cortical network dysfunctions
Authors: Singh Surjeet , Lacoursiere Sean G. , Mehla Jogender , Nazari Mojtaba , Sutherland Robert J. , McDonald Robert J. , Mohajerani Majid H.
Published: October 2022

Alzheimer’s disease (AD) is characterized neuropathologically by amyloid-β (Aβ) plaques and neurofibrillary tangles. Vascular pathology caused by chronic cerebral hypoperfusion (HP) is hypothesised to exacerbate AD pathology and has emerged as an increasing cause of age-related cognitive impairment. In this study we examined the effects of gradual cerebral HP on cognitive dysfunction, Aβ pathology, microgliosis, and cortical network dynamics in C57BL/6J mice and a single App knock-in mouse model of AD (AppNL-G-F). We performed unilateral common carotid artery gradual occlusion (UCAgO) in two-month-old mice using an ameroid constrictor. At 4 months of age, animals were tested in a behavioral battery consisting of tests of spatial learning and memory (Morris water task), recognition memory (novel object recognition task), and motor coordination (balance beam). Following behavioural testing,in vivomesoscale wide-field voltage imaging was done to assess cortical functional connectivity and sensory-evoked cortical activity, and brains were harvested for pathology characterization using immunohistochemistry. We found that UCAgO reduced cerebral blood flow (CBF) in the occluded hemisphere (OH), however, subtle behavioural deficits were observed due to HP. A dissociative effect of HP was observed in resting-state functional connectivity analysis, where HP led to hyper-connectivity in C57 mice and hypo-connectivity in App mice. Interestingly, sensory stimulation of limbs contralateral to OH revealed hyper-cortical activations in the non-occluded hemisphere of C57 HP mice, however, hypo-cortical activations were observed in App HP mice. Furthermore, we found that the UCAgO increased cortical and hippocampal microgliosis in both hemispheres of C57 and App mice, a bilateral increase in Aβ deposition was only observed in App mice. These results suggest that gradual cerebral HP leads to cortical network alterations in AD, which is partly mediated via activation of microglia.
A Robust Bayesian Approach to Bulk Gene Expression Deconvolution with Noisy Reference Signatures
Authors: Ghaffari Saba , Bouchonville Kelly J. , Saleh Ehsan , Schmidt Remington E. , Offer Steven M. , Sinha Saurabh
Published: October 2022

Background

Differential gene expression in bulk transcriptomics data can reflect change of transcript abundance within a cell type and/or change in the proportion of cell types within the sample. Expression deconvolution methods can help differentiate these scenarios and enable more accurate inference of gene regulation by estimating the contributions of individual cell types to bulk transcriptomic profiles. However, the accuracy of these methods is sensitive to technical and biological differences between bulk profiles and the cell type-signatures required by them as references.

Results

We present BEDwARS, a Bayesian deconvolution method specifically designed to address differences between reference signatures and the unknown true signatures underlying bulk transcriptomic profiles. Through extensive benchmarking utilizing eight different datasets derived from pancreas and brain, we demonstrate that BEDwARS outperforms leading in-class methods for estimating cell type proportions and signatures. Furthermore, we systematically show that BEDwARS is more robust to noisy reference signatures than all compared methods. Finally, we apply BEDwARS to newly generated RNA-seq and scRNA-seq data on over 100 induced pluripotent stem cell-derived neural organoids to study mechanisms underlying a rare pediatric condition (DihydropyridineDehydrogenase deficiency), identifying the possible involvement of ciliopathy and impaired translational control in the etiology of the disorder.

Conclusion

We propose a new approach to bulk gene expression deconvolution which estimates the cell type proportions and cell type signatures simultaneously and is robust to commonly seen mismatches between reference and true cell type signatures. Application of our method lead to novel findings about mechanisms of a rare pediatric condition.
Development of Highly Efficient CRISPR-Mediated Gene Editing in the RotiferBrachionus manjavacas
Authors: Feng Haiyang , Bavister Gemma , Gribble Kristin E. , Welch David B. Mark
Published: October 2022

Rotifers have been studied in the laboratory and field for over 100 years and are an emerging modern model system for investigation of the molecular mechanisms of genome evolution, development, DNA repair, aging, life history strategy, and desiccation tolerance, and have a long been used in studies of microevolution, ecological dynamics, and ecotoxicology. However, a lack of gene editing tools and transgenic strains has limited the ability to link genotype to phenotype and dissect molecular mechanisms. To facilitate genetic manipulation and the creation of reporter lines, we developed a protocol for highly efficient, transgenerational, CRISPR-mediated gene editing in the monogonont rotiferBrachionus manjavacasby microinjection of Cas9 protein and synthetic single guide RNA into the vitellaria of young amictic (asexual) females. To demonstrate the efficacy of the method, we created knockout mutants of the developmental genevasaand the DNA mismatch repair genemlh3. More than half of mothers survived injection and produced offspring. Genotyping these offspring and successive generations revealed that most carried at least one CRISPR-induced mutation, with many apparently mutated at both alleles or mosaic. In addition, we achieved precise CRISPR-mediated knockin of a stop codon cassette in themlh3locus, with half of injected mothers producing 33% or more F2 offspring with an insertion of the cassette. These results demonstrate the efficacy of the CRISPR/Cas9 system in rotifers to provide insight into the function of specific genes and further advance rotifers as a model system for biological discovery.
Directed functional brain connectivity is altered in sub-threshold amyloid-β accumulators
Authors: Mijalkov Mite , Veréb Dániel , Canal-Garcia Anna , Volpe Giovanni , Pereira Joana B. ,   Alzheimer’s Disease Neuroimaging Initiative
Published: October 2022

Several studies have shown that amyloid-β (Aβ) deposition below the clinically relevant cut-off levels is associated with subtle changes in cognitive function and increases the risk for developing future Alzheimer’s disease (AD). Although functional MRI is sensitive to early alterations occurring during AD, sub-threshold changes in Aβ levels have not been linked to functional connectivity measures. This study aimed to apply directed functional connectivity to identify early changes in network function in cognitively unimpaired participants who, at baseline, exhibit Aβ accumulation below the clinically relevant threshold. To this end, we analyzed baseline functional MRI data from 113 cognitively unimpaired participants of the Alzheimer’s Disease Neuroimaging Initiative cohort who underwent at least one18F-florbetapir-PET after the baseline scan. Using the longitudinal PET data, we classified these participants as Aβ negative (Aβ−) non-accumulators (n=46) and Aβ-accumulators (n=31). We also included 36 individuals who were amyloid-positive (Aβ+) at baseline and continued to accumulate Aβ (Aβ+ accumulators). For each participant, we calculated whole-brain directed functional connectivity networks using our own anti-symmetric correlation method and evaluated their global and nodal properties using measures of network segregation (clustering coefficient) and integration (global efficiency). When compared to Aβ-non-accumulators, the Aβ-accumulators showed lower global clustering coefficient. Moreover, the Aβ+ accumulator group exhibited reduced global efficiency and clustering coefficient, which at the nodal level mainly affected the superior frontal gyrus, anterior cingulate cortex and caudate nucleus. In Aβ-accumulators, global measures were associated with lower baseline regional PET uptake values, as well as higher scores on the Modified Preclinical Alzheimer Cognitive Composite. Our findings indicate that directed connectivity network properties are sensitive to subtle changes occurring in individuals who have not yet reached the threshold for Aβ positivity, which makes them a potentially viable marker to detect negative downstream effects of very early Aβ pathology.

Significance Statement

Directed functional connectivity measures are sensitive to very early changes in amyloid-β deposition.

Amyloid-β burden is associated with decreases in global and nodal directed measures of network integration and segregation.

Measures of directed network integration and segregation are associated with subclinical cognitive changes in Aβ-negative accumulators.
Generalizability and Clinical Implications of Electrocardiogram Denoising with Cardio-NAFNet
Authors: Lim Chanho , Chung Yunsung , Hamm Jihun , Ding Zhengming , Mekhael Mario , Noujaim Charbel , Assaf Ala , Younes Hadi , Chouman Nour , Makan Noor , Donnellan Eoin , Marrouche Nassir
Published: October 2022

The rise of mobile electrocardiogram (ECG) devices came with the rise of frequent large magnitudes of noise in their recordings. Several artificial intelligence (AI) models have had great success in denoising, but the model’s generalizability and the enhancement in clinical interpretability are still questionable. We propose Cardio-NAFNet, a novel AI-based approach to ECG denoising by employing a modified version of Non-Linear Activation Free Network (NAFNET). We conducted three experiments for quantitative and qualitative evaluation of denoising, clinical implications and generalizability. In the first experiment, Cardio-NAFNet achieved 53.74dB average signal to noise ratio across varying magnitude of noise in beat-to-beat denoising, which is a significant improvement over the current state of the art model in ECG denoising. In the second experiment, we tested the enhancement in clinical interpretation of the ECG signals by utilizing a pretrained ECG classifier using 8 second long noise-free ECG signals. When the classifier was tested using noisy ECG signals and their denoised counterparts, Cardio-NAFNet’s denoised signals provided 26% boost in classification results. Lastly, we provide an external validation dataset composed of single-lead mobile ECG signals along with signal quality evaluation from physician experts. Our paper suggests a settling method to capture and reconstruct critical features of ECG signals not only in terms of quantitative evaluation, but also through generalizable qualitative evaluation.
Development and Validation of Sex-Specific Hip Fracture Prediction Models using Electronic Health Records
Authors: Li Gloria Hoi-Yee , Cheung Ching-Lung , Tan Kathryn Choon-Beng , Kung Annie Wai-Chee , Kwok Timothy Chi-Yui , Lau Wallis Cheuk-Yin , Wong Janus Siu-Him , Hsu Warrington W.Q. , Fang Christian , Wong Ian Chi-Kei
Published: October 2022

Background

Hip fracture is associated with immobility, morbidity, mortality, and high medical cost. Due to limited availability of dual-energy X-ray absorptiometry (DXA), hip fracture prediction models without using bone mineral density (BMD) data are essential. We aimed to develop and validate 10-year sex-specific hip fracture prediction models using electronic health records (EHR) without BMD.

Methods

In this population-based study, the derivation cohort comprised 161,051 public healthcare service users (91,926 female; 69,125 male) in Hong Kong aged≥60. Sex-stratified derivation cohort was randomly split to 80% training and 20% internal testing datasets. An external validation cohort comprised 3,046 community-dwelling participants. With 395 potential predictors (age, diagnosis and drug prescription records from EHR), 10-year sex-specific hip fracture prediction models were developed using stepwise selection by logistic regression (LR) and four machine learning (ML) algorithms (gradient boosting machine, random forest, eXtreme gradient boosting, and single-layer neural networks) in the training cohort. Model performance was evaluated in both internal and external validation cohorts.

Findings

In female, the LR model had the highest AUC (0.815) and adequate calibration in internal validation. Reclassification metrics showed ML algorithms could not further improve the performance of the LR model. Similar performance was attained by the LR model in external validation, with high AUC (0.841) comparable to other ML algorithms. In internal validation for male, LR model had high AUC (0.818) and it outperformed all ML models as indicated by reclassification metrics, with adequate calibration. In external validation, the LR model had high AUC (0.898) comparable to ML algorithms. Reclassification metrics demonstrated that LR model had the best discrimination performance.

Interpretation

Even without using BMD data, the 10-year hip fracture prediction models developed by conventional LR had better discrimination performance than the models developed by ML algorithms. Upon further validation in independent cohorts, the LR models could be integrated into the routine clinical workflow, aiding the identification of people at high risk for DXA scan.

Funding

This study was funded by the Health and Medical Research Fund, Food and Health Bureau, Hong Kong SAR Government (reference: 17181381).
One-Step PCR Amplicon Library Construction (OSPALC, version 1)
Authors: Ni Jiahao , Pan Jiao , Wang Yaohai , Chen Tianhao , Feng Xinshi , Li Yichen , Lin Tongtong , Lynch Michael , Long Hongan , Li Weiyi
Published: October 2022

High-throughput sequencing of amplicons has been widely used to precisely and efficiently identify species compositions and analyze community structures, greatly promoting biological studies involving large amounts of complex samples, especially those involving environmental and pathogen-monitoring ones. However, commercial library preparation kits for amplicon sequencing, which generally require multiple steps, including adapter ligation and indexing, are expensive and time-consuming, especially for applications at a large scale. Here, to overcome this technical hurdle, we present a one-step PCR amplicon library construction (OSPALC) protocol for amplicon library preparations in the lab along with a QIIME2-based amplicon analysis pipeline. High-quality reads have been generated by this approach to reliably identify species compositions of mock bacterial communities and environmental samples. With this protocol, the amplicon library construction is completed through one regular PCR with long primers, and the total cost per DNA/cDNA sample decreases to just 1/15 of the typical cost via service providers. Empirically tested primers to construct OSPALC libraries for 16S rDNA V4 regions are demonstrated as a case study. Criteria to design primers targeting at any regions of prokaryotic and eukaryotic genomes are also suggested. In principle, OSPALC can be conveniently used in amplicon library constructions of any target gene of both prokaryotes and eukaryotes at the DNA or RNA levels, and will facilitate research in numerous fields.
A pan-cancer PDX histology image repository with genomic and pathological annotations for deep learning analysis
Authors: White Brian S , Woo Xing Yi , Koc Soner , Sheridan Todd , Neuhauser Steven B , Wang Shidan , Evrard Yvonne A , Landua John David , Mashl R Jay , Davies Sherri R , Fang Bingliang , Raso Maria Gabriela , Evans Kurt W , Bailey Matthew H , Chen Yeqing , Xiao Min , Rubinstein Jill , pour Ali Foroughi , Dobrolecki Lacey Elizabeth , Fujita Maihi , Fujimoto Junya , Xiao Guanghua , Fields Ryan C , Mudd Jacqueline L , Xu Xiaowei , Hollingshead Melinda G , Jiwani Shahanawaz ,   PDXNet consortium, Davis-Dusenbery Brandi , Wallace Tiffany A , Moscow Jeffrey A , Doroshow James H , Mitsiades Nicholas , Kaochar Salma , Pan Chong-xian , Chen Moon S , Carvajal-Carmona Luis G , Welm Alana L , Welm Bryan E , Govindan Ramaswamy , Li Shunqiang , Davies Michael A , Roth Jack A , Meric-Bernstam Funda , Xie Yang , Herlyn Meenhard , Ding Li , Lewis Michael T , Bult Carol J , Dean Dennis A , Chuang Jeffrey H
Published: October 2022

Patient-derived xenografts (PDXs) model human intra-tumoral heterogeneity in the context of the intact tissue of immunocompromised mice. Histological imaging via hematoxylin and eosin (H&E) staining is performed on PDX samples for routine assessment and, in principle, captures the complex interplay between tumor and stromal cells. Deep learning (DL)-based analysis of largehumanH&E image repositories has extracted inter-cellular and morphological signals correlated with disease phenotype and therapeutic response. Here, we present an extensive, pan-cancer repository of nearly 1,000PDXand paired human progenitor H&E images. These images, curated from the PDXNet consortium, are associated with genomic and transcriptomic data, clinical metadata, pathological assessment of cell composition, and, in several cases, detailed pathological annotation of tumor, stroma, and necrotic regions. We demonstrate that DL can be applied to these images to classify tumor regions and to predict xenograft-transplant lymphoproliferative disorder, the unintended outgrowth of human lymphocytes at the transplantation site. This repository enables PDX-specific, investigations of cancer biology through histopathological analysis and contributes important model system data that expand on existing human histology repositories. We expect the PDXNet Image Repository to be valuable for controlled digital pathology analysis, both for the evaluation of technical issues such as stain normalization and for development of novel computational methods based on spatial behaviors within cancer tissues.
ASO targeting temperature-controlledRBM3poison exon splicing prevents neurodegeneration in vivo
Authors: Preußner Marco , Smith Heather L , Zhang Min , Hughes Daniel , Emmerichs Ann-Kathrin , Scalzitti Silvia , Peretti Diego , Swinden Dean , Neumann Alexander , Haltenhof Tom , Mallucci Giovanna R , Heyd Florian
Published: October 2022

Neurodegenerative diseases are increasingly prevalent in the aging population, yet currently no disease-modifying treatments are available. Increasing the expression of the cold-shock protein, RBM3, through therapeutic hypothermia is remarkably neuroprotective, but cooling poses a health risk itself, strongly limiting its clinical application. Selective upregulation of RBM3 at normothermia thus holds immense therapeutic potential. Here we identify a poison exon within the RBM3 gene that is solely responsible for cold-induced RBM3 expression. Genetic removal or ASO-mediated manipulation of this exon yields high RBM3 levels independent of cooling. Notably, a single administration of ASO to exclude the poison exon, using FDA-approved chemistry, results in long-lasting increase of RBM3 expression in mouse brains. In prion-diseased mice, this treatment leads to remarkable neuroprotection, with prevention of neuronal loss and spongiosis despite high levels of prion protein. RBM3-inducing ASOs could thus broadly deliver protection in humans in conditions ranging from acute brain injury to Alzheimer’s disease.

One sentence summary

Inducing cold shock protein RBM3 by modulating its alternative splicing at normothermia is neuroprotective in vivo
Examining the early distribution of the artemisinin-resistantPlasmodium falciparumkelch13 R561H mutation in Rwanda
Authors: Kirby Rebecca , Giesbrecht David , Karema Corine , Watson Oliver , Lewis Savannah , Munyaneza Tharcisse , de Dieu Butera Jean , Juliano Jonathan J , Bailey Jeffrey , Mazarati Jean Baptiste
Published: October 2022

Artemisinin resistance mutations inPlasmodium falciparum kelch13(Pfk13) have begun to emerge in Africa.Pfk13-R561H was the first reported African mutation found in Rwanda in 2014, but limited sampling left questions about its early distribution and origin. We detected 476 parasitemias among 1873 residual blood spots from a 2014-15 Rwanda Demographic Health Survey. We sequenced 351 samples revealing 341/351 were wild type (97.03% weighted) and 4 samples (1.34% weighted) harbored R561H which were significantly spatially clustered. Our study better defines the early distribution of R561H in Rwanda and suggests that the origin may have involved higher-transmission regions.
In Silico Analysis of NovelVHLGermline Mutations in Iranian RCH Patients
Authors: Naseripour Masood , Bagherzadeh Kowsar , Khakpoor Golnaz , Sedaghat Ahad , Mirshahi Reza , Kasraee Hengameh , Kiaee Reza Afshar , Azimi Fatemeh
Published: October 2022

Von Hippel-Lindau (VHL) syndrome is an autosomal dominant inherited multisystem neoplasia disorder caused by theVHLtumor suppressor gene, coding for VHL protein (pVHL), variants. Various types ofVHLvariants present different clinical phenotypes that later lead to events resulting in benign or malignant lesions including Retinal Capillary Hemangioblastoma (RCH). We reported on 3 novel mutation sites observed in 3 families (5 RCH patients), including c.511A>C, c.514C>T, and c.511A>T in exon 3 of theVHLgene. According to the ACMG classifications, c.514C>T and c.511A>T variations are likely pathogenic, and c.511A>C is a variant of uncertain significance (VUS) and in accordance with autosomal dominant inheritance. The location and impact of the incidence mutations on pVHL were computed using in silico analysis. The obtained structural information and computational analysis showed that the studied mutations induce conformational changes that limit the flexibility of the pVHL interaction interface with elonginB/C, elongin C/B, and cullin2, which is necessary for HIF1 alpha binding. The recently added gene variants and their related clinical phenotypes will improve the VHL diagnosis accuracy and the patients’ population carryingVHLgene mutations. These pioneering results could be a model for future functional studies.
AI-Enabled Classification of Head and Neck Tumors using Convolutional Neural Networks with Dynamic Optical Contrast Imaging
Authors: Odion Ren , Mukdad Laith , Alhiyari Yazeed , Tam Kenric , Shori Ramesh , Vo-Dinh Tuan , John Maie A. St.
Published: October 2022

Background

Recent advances in artificial intelligence (AI) in the field of imaging have resulted in new opportunities for automated tumor detection and margin assessment. In particular, AI deep learning techniques such as the Convolutional Neural Network (CNN) have greatly advanced the field of computer vision. Here we introduce the application of a CNN model for use with Dynamic Optical Contrast Imaging (DOCI), an imaging technique developed by our group that creates a unique molecular signature on tissue targets by obtaining the autofluorescence decay of several spectral bands in the UV-Vis range.

Methods

21 patients undergoing surgical resection for tonsillar squamous cell carcinoma (SCC) were identified on a prospective basis. DOCI images were analyzed and compared to the pathology results as ground truth. A CNN model was used to segment sections of DOCI images and provide a percentage chance of tumor presence, allowing for automated tumor margin delineation without a-priori knowledge of the tumor tissue composition.

Results

CNN outputs yielded a 99.98% confidence in classifying non-tumor tissue and 76.02% confidence in classifying tumor tissue.

Conclusions

Our results indicate that a CNN-based classification model for DOCI allows for real-time analysis of tissue, providing improved sensitivity and accuracy of determining true margins and thus enabling the head and neck cancer surgeon to save healthy tissue and improve patient outcomes.
The function and decline of the female reproductive tract at single-cell resolution
Authors: Winkler Ivana , Tolkachov Alexander , Lammers Fritjof , Lacour Perrine , Schneider Nina , Koch Marie-Luise , Panten Jasper , Grünschläger Florian , Daugelaite Klaudija , Poth Tanja , Haas Simon , Odom Duncan T. , Goncalves Angela
Published: October 2022

The female reproductive tract (FRT) undergoes extensive remodeling during each reproductive cycle, regulated by systemic changes in sex hormones. Whether this recurrent remodeling influences a specific organ’s aging trajectory is unknown. To address this, we systematically characterized at single-cell resolution the morphological and transcriptional changes that occur in ovary, oviduct, uterus, cervix, and vagina at each phase of the mouse estrus cycle, during decidualization, and into aging. Transcriptional and cell-to-cell communication networks in estrus cycle and aging are enriched for ECM reorganization and inflammation, two essential components of FRT remodeling. We directly link the organ-specific level of these two processes over reproductive lifespan with the gradual, age-related development of fibrosis and chronic inflammation. Our data represent a comprehensive atlas of the FRT lifespan, revealing pathological consequences of incomplete resolution of recurrent inflammation and tissue repair.
Transcription factor HSFA7b controls ethylene signaling and meristem maintenance at the shoot apical meristem during thermomemory
Authors: John Sheeba , Apelt Federico , Kumar Amit , Bents Dominik , Annunziata Maria Grazia , Fichtner Franziska , Mueller-Roeber Bernd , Olas Justyna J.
Published: October 2022

The shoot apical meristem (SAM) is responsible for overall shoot growth by generating all above-ground structures. Recent research identified that the SAM displays an autonomous heat stress (HS) memory of a previous non-lethal HS event. Considering the importance of the SAM for plant growth it is essential to unlock how its thermomemory is mechanistically controlled. Here, we report thatHEAT SHOCK TRANSCRIPTION FACTOR A7b(HSFA7b) plays a crucial role in this process in Arabidopsis. We found that HSFA7b directly regulates ethylene response at the SAM by binding to promoters of key regulators of ethylene signaling includingETHYLENE-INSENSITIVE 3to establish thermotolerance. Moreover, HSFA7b controls maintenance of the SAM stem cell pool during thermomemory by regulating the expression of the master regulatorWUSCHELthrough direct transcriptional activation of theSPLAYEDchromatin remodelling factor.