bluuebunny/crossref_metadata_2025 huggingface dataset:
Bioinformatics Analysis of next generation sequencing data for Risk Prediction in Patients with Type 1 diabetes mellitus
Authors: Vastrad Basavaraj , Vastrad Chanabasayya
Published: March 2022

Type 1 diabetes mellitus (T1DM) comprise the most common forms of autoimmune disease. The aim of this investigation was to apply a bioinformatics approach to reveal related pathways or genes involved in the development of T1DM. The next generation sequencing (NGS) dataset GSE182870 was downloaded from the gene expression omnibus (GEO) database. Differentially expressed gene (DEG) analysis was performed using DESeq2. The g:Profiler was utilized to analyze the functional enrichment, gene ontology (GO) and REACTOME pathway of the differentially expressed genes. Protein-protein interaction (PPI) network, modules, miRNA- hub gene regulatory network, and TF-hub gene regulatory network were conducted via comprehensive target prediction and network analyses. Finally, hub genes were validated by using receiver operating characteristic curve analysis. A total of 860 DEGs were screened out from NGS dataset, among which 477 genes were up regulated and 383 genes were down regulated. GO enrichment analysis indicated that up regulated genes were mainly involved in cellular metabolic process, intracellular anatomical structure and catalytic activity, and the down regulated genes were significantly enriched in cellular nitrogen compound biosynthetic process, protein containing complex and heterocyclic compound binding. REACTOME pathway enrichment analysis showed that the up regulated genes were mainly enriched in metabolism of carbohydrates and the down regulated genes were significantly enriched in metabolism of RNA. A PPI network, modules, miRNA-hub gene regulatory network and TF - hub gene regulatory network were constructed, and hub genes (MAPK14, RHOC, MAD2L1, TAF1, TRAF2, HSP90AA1, TP53, HSP90AB1, UBA52 and RACK1) were identified. This study provides further insights into the underlying pathogenesis of T1DM.
Characterization of Quantum Dots with Hyperspectral Fluorescence Microscopy for Multiplexed Optical Imaging of Biomolecules
Authors: Zhang Shuyan , Xin Cheng Joseph Yong , Chua Jian Jun , Olivo Malini
Published: March 2022

The optical properties of quantum dots were extensively characterized using a hyperspectral fluorescence microscopy system. The system provides a single excitation wavelength in the ultraviolet and 311 emission wavelength channels in the visible. This allows detection of multiple fluorophores (e.g. different quantum dots) with a high spectral resolution in one go which is not achievable with a conventional fluorescence microscope where different filter sets have to be used. A spectral library was established based on the spectral profiles of six types of quantum dots. Notably, a slight spectral shift was observed for all samples while the sample was drying. Subsequently, two quantum dot mixture samples were studied. Using the spectral unmixing approach, the relative proportions of each quantum dot within a homogeneous mixture and the spatial distribution of each quantum dot within a heterogeneous mixture were calculated. The calculated values match well with the theoretical predictions. Hence, the analysis method presented here can be used for simultaneous imaging of multiple fluorophores using hyperspectral imaging technology. The results provide valuable information for the realization of real-time multi-channel in vivo fluorescent imaging of biomolecules.
POU2AF2/C11orf53 functions as a co-activator of POU2F3 by maintaining chromatin accessibility and enhancer activity
Authors: Szczepanski Aileen Patricia , Tsuboyama Natsumi , Zhao Zibo , Wang Lu
Published: March 2022

Small cell lung cancer (SCLC), accounting for around 13% of all lung cancers, often results in rapid tumor growth, early metastasis, and acquired therapeutic resistance. The POU class 2 homeobox 3 (POU2F3) is a master regulator of tuft cell identity and defines a subtype of SCLC tumors (SCLC-P subtype) that lack the expression of neuroendocrine (NE) markers. Here, we have identified a previously uncharacterized protein, C11orf53, which is co-expressed with POU2F3 in both SCLC cell lines and patient samples, defining the same SCLC-P subtype. C11orf53 is an essential gene for the cell survivability of SCLC-P subtype, based on genome-wide CRISPR screening and genetic depletion of C11orf53 or POU2F3, inducing apoptosis in different SCLC-P subtype cell lines. Furthermore, our biochemical and genome-wide studies have demonstrated that C11orf53 is a novel chromatin-bound protein that occupies broad active enhancer domains and regulates gene expression at active enhancers. Mechanistically, C11orf53 directly interacts with POU2F3 at the POU domain, and is recruited to chromatin by POU2F3. Consequently, depletion of C11orf53 dramatically reduced enhancer H3K27ac levels and chromatin accessibility, resulting in a reduction of the expression of POU2F3-dependent tuft cell markers. Based on the expression pattern and molecular function of C11orf53, we have renamed it as “POU Class 2 Homeobox Associating Factor 2” (POU2AF2). In summary, our study has identified a new co-activator of POU2 family transcription factors, and sheds light on the therapeutic potential of targeting POU2AF2/POU2F3 heterodimer in human SCLC.
Towards reliable quantification of cell state velocities
Authors: Marot-Lassauzaie Valérie , Bouman Brigitte Joanne , Donaghy Fearghal Declan , Haghverdi Laleh
Published: March 2022

A few years ago, it was proposed to use the simultaneous quantification of unspliced and spliced messenger RNA (mRNA) to add a temporal dimension to high-throughput snapshots of single cell RNA sequencing data. This concept can yield additional insight into the transcriptional dynamics of the biological systems under study. However, current methods for inferring cell state velocities from such data (known as RNA velocities) are afflicted by several theoretical and computational problems, hindering realistic and reliable velocity estimation. We discuss these issues and propose new solutions for addressing some of the current challenges in consistency of data processing, velocity inference and visualisation. We translate our computational conclusion in two velocity analysis tools: one detailed method κ-velo and one heuristic method eco-velo.

Author summary

Single cell transcriptomics has been used to study dynamical biological processes such as cell differentiation or disease progression. An ideal study of these systems would track individual cells in time but this is not directly feasible since cells are destroyed as part of the sequencing protocol. Because of asynchronous progression of cells, single cell snapshot datasets often capture cells at different stages of progression. The challenge is to infer both the overall direction of progression (pseudotime) as well as single cell specific variations in the progression. Computational methods development for inference of the overall direction are well advanced but attempts to address the single cell level variations of the dynamics are newer. La Manno et al. [1] proposed that simultaneous measurement of abundances of new (unspliced) and older (spliced) mRNA in the same single cell adds a temporal dimension to the data which can be used to infer the time derivative of single cells progression through the dynamical process. State-of-the-art methods for inference of cell state velocities from RNA-seq data (also known as RNA velocity) have multiple unaddressed issues. In this manuscript, we discuss these issues and propose new solutions. In previous works, agreement of RNA velocity estimations with pseudotime has been used as validation. We show that this in itself is not proof that the method works reliably and the overall direction of progression has to be distinguished from individual cells’ behaviour. We propose two new methods (one detailed and one cost efficient heuristic) for estimation and visualisation of RNA velocities and show that our methods faithfully capture the single-cell variances and overall trend on simulation. We further apply the methods to a dataset of developing mouse pancreas and show how the method can help us gain biological insight from real data.
Modeling of Aryl Hydrocarbon Receptor Pathway Intrinsic Immunometabolic Role using Glioblastoma Stem Cells and Patient-Derived Organoids
Authors: Watanabe Fumihiro , Hollingsworth Ethan W. , Banasavadi-Siddegowda Yeshavanth , Bartley Jenna M. , Sizemore Steven T. , Yu Lianbo , Pietrzak Maciej , Schiapparelli Paula , Puduvalli Vinay , Kaur Balveen , Quiñones-Hinojosa Alfredo , Imitola Jaime
Published: March 2022

The intrinsic genetic program of glioblastoma (GBM) stem cells is critical for tumor evolution and recurrence. We recently identified intrinsic phenotypes and immune-like genetic programs of GBM organoids (GBMO)1 from patient derived glioblastoma stem cells (GSCs), replicating genomic, metabolic, and cellular aspects of GBM in vivo. Aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor, is a key regulator of infiltrating immune cells in gliomas2, 3 and associated with poor prognosis, but its role in GSC biology is unknown2. Here, we show that AHR is a patient-specific regulator of the glioma intrinsic gene program in GSCs and GSC-derived GBMO that are enriched for AHR. We find that AHR is required for GSC self-renewal, GBMO expansion, radial glia-like cell proliferation, and expression of immune mediators seen in the mesenchymal subtype. CRISPR-Cas9 genetic ablation and pharmacological inhibition revealed that AHR regulates genes linked to intrinsic immunity, proliferation, and migration in GBMO. Genomic analysis of GBMO treated with AHR inhibitors identified expression signatures and candidate markers associated with survival of gliomas. Our work defines the glioma intrinsic function of AHR in a model of early GBM formation, offering a rationale for clinical exploration of a potential ‘two-hit’ target of both GBM cells and infiltrating immune cells in patients with GBM expressing high levels of AHR.
KAT5 regulates neurodevelopmental states associated with G0-like populations in glioblastoma
Authors: Mihalas Anca B. , Arora Sonali , O’Connor Samantha A. , Feldman Heather M. , Cucinotta Christine E. , Mitchell Kelly , Bassett John , Kim Dayoung , Jin Kang , Hoellerbauer Pia , Delegard Jennifer , Ling Melissa , Jenkins Wesley , Kufeld Megan , Corrin Philip , Carter Lucas , Tsukiyama Toshio , Aronow Bruce , Plaisier Christopher L. , Patel Anoop P. , Paddison Patrick J.
Published: March 2022

In solid tumors, G0-like states are likely critical for maintaining developmental hierarchies and cellular heterogeneity and promoting tumor growth/recurrence, yet little is known about tumor G0 states or regulation of their ingress/egress. To discover G0-like states and their regulators for glioblastoma (GBM), we analyzed G0 populations in an orthotopic model of GBM using single cell RNA-seq and performed a genome-wide CRISPR-Cas9 screen in patient-derived GBM stem-like cells (GSCs) for genes that trap cells in G0 when inhibited. We identify the protein acetyltransferase KAT5 as a key regulator of transcriptional, epigenetic, and proliferative heterogeneity impacting transitions into G0-like states. KAT5 activity suppresses the emergence of non-dividing subpopulations with oligodendrocyte progenitor and radial glial cell characteristics bothin vitroand in a human GSC brain tumor model. In primary gliomas, KAT5 activity is dynamic with KAT5lowtumor cells displaying quiescent properties, while KAT5 activity overall increases from low to high grade tumors and is associated with worse patient outcomes.
A novel class of long small RNAs associates with Argonaute1 and is up-regulated by nutrient deprivation in the alga Chlamydomonas
Authors: Li Yingshan , Kim Eun-Jeong , Voshall Adam , Moriyama Etsuko N. , Cerutti Heriberto
Published: March 2022

Small RNAs (sRNAs) associate with Argonaute (AGO) proteins forming effector complexes with key roles in gene regulation and defense responses against molecular parasites. In multicellular eukaryotes, extensive duplication and diversification of RNA interference (RNAi) components have resulted in intricate pathways for epigenetic control of gene expression. The unicellular alga Chlamydomonas reinhardtii also has a complex RNAi machinery, including three AGOs and three Dicer-like (DCL) proteins. However, little is known about the biogenesis and function of most endogenous sRNAs. We demonstrate here that Chlamydomonas contains uncommonly long sRNAs (>26 nt), which associate preferentially with AGO1. Somewhat reminiscent of animal PIWI-interacting RNAs, these long sRNAs are derived from moderately repetitive genomic clusters and their biogenesis appears to be Dicer-independent. Interestingly, long sRNA encoding sequences have been conserved and amplified in phylogenetically related Chlamydomonas species. Additionally, expression of several long sRNAs increases substantially under nutrient deprivation, correlating with the downregulation of predicted target transcripts. We hypothesize that the transposon-like sequences encoding long sRNAs might have been ancestrally targeted for silencing by the RNAi machinery but, during evolution, some long sRNAs might have fortuitously acquired endogenous target genes and become integrated into gene regulatory networks.
Avatar error in your favor: Embodied avatars can fix users’ mistakes without them noticing
Authors: Delahaye Mathias , Blanke Olaf , Boulic Ronan , Herbelin Bruno
Published: March 2022

In immersive Virtual Reality (VR), users can experience the subjective feeling of embodiment for the avatar representing them in a virtual world. This is known to be strongly supported by a high Sense of Agency (SoA) for the movements of the avatar that follows the user. In general, users do not self-attribute actions of their avatar that are different from the one they actually performed. The situation is less clear when actions of the avatar satisfies the intention of the user despite distortions and noticeable differences between user and avatar movements. A within-subject experiment was here conducted to determine whether a finger swap helping users achieve a task would be more tolerated than one penalizing them. In particular, in a context of fast-paced finger movements and with clear correct or incorrect responses, we swapped the finger animation of the avatar (e.g. user moves the index finger, the avatar moves the middle one) to either automatically correct for spontaneous mistakes or to introduce incorrect responses. Subjects playing a VR game were asked to report when they noticed the introduction of a finger swap. Results based on 3256 trials (~24% of swaps noticed) show that swaps helping users have significantly fewer odds of being noticed (and with higher confidence) than the ones penalizing users. This demonstrates how the context and the intention for motor action are important factors for the SoA and for embodiment, opening new perspectives on how to design and study interactions in immersive VR.
The motif composition of variable-number tandem repeats impacts gene expression
Authors: Lu Tsung-Yu , Smaruj Paulina N. , Fudenberg Geoffrey , Mancuso Nicholas , Chaisson Mark J.P.
Published: March 2022

Understanding the impact of DNA variation on human traits is a fundamental question in human genetics. Variable number tandem repeats (VNTRs) make up roughly 3% of the human genome but are often excluded from association analysis due to poor read mappability or divergent repeat content. While methods exist to estimate VNTR length from short-read data, it is known that VNTRs vary in both length and repeat (motif) composition. Here, we use a repeat-pangenome graph (RPGG) constructed on 35 haplotype-resolved assemblies to detect variation in both VNTR length and repeat composition. We align population scale data from the Genotype-Tissue Expression (GTEx) Consortium to examine how variations in sequence composition may be linked to expression, including cases independent of overall VNTR length. We find that 9,422 out of 39,125 VNTRs are associated with nearby gene expression through motif variations, of which only 23.4% associations are accessible from length. Fine-mapping identifies 174 genes to be likely driven by variation in certain VNTR motifs and not overall length. We highlight two genes,CACNA1CandRNF213that have expression associated with motif variation, demonstrating the utility of RPGG analysis as a new approach for trait association in multiallelic and highly variable loci.
A murine model of hnRNPH2-related neurodevelopmental disorder recapitulates clinical features of human disease and reveals a mechanism for genetic compensation ofHNRNPH2
Authors: Korff Ane , Yang Xiaojing , O’Donovan Kevin , Gonzalez Abner , Teubner Brett J. W. , Nakamura Haruko , Messing James , Yang Fen , Carisey Alex , Wang Yong-Dong , Patni Tushar , Tillman Heather , Zakharenko Stanislav S. , Chook Yuh Min , Taylor J. Paul , Kim Hong Joo
Published: March 2022

Mutations inHNRNPH2cause an X-linked neurodevelopmental disorder with a phenotypic spectrum that includes developmental delay, intellectual disability, language impairment, motor function deficits, and seizures. More than 90% of patients with this disorder have a missense mutation within or adjacent to the nuclear localization signal (NLS) of hnRNPH2, although the specific pathogenic consequences of these mutations have not been examined. Here we found that hnRNPH2 NLS mutations result in reduced interaction with the nuclear transport receptor Kapβ2 in vitro and in cultured human cells. These mutations also cause modest accumulation of hnRNPH2 in the cytoplasm, suggesting that mislocalization of the protein might contribute to pathogenesis. We generated two knock-in mouse models with human-equivalent mutations in the endogenous mouse geneHnrnph2, as well asHnrnph2knockout (KO) mice, and subjected them to extensive phenotyping. Mutant knock-in mice displayed a spectrum of phenotypes that recapitulated aspects of the human disorder, including reduced survival in males, craniofacial abnormalities, impaired motor and cognitive functions, and increased susceptibility to audiogenic seizures. Mutant knock-in male mice developed more severe phenotypes than female mice, likely due to differences in X-chromosome gene dosage. In contrast, two independent lines ofHnrnph2KO mice showed no detectable phenotypes. Notably, KO mice had upregulated expression ofHnrnph1, a close paralog ofHnrnph2, whereas mutantHnrnph2knock-in mice failed to upregulateHnrnph1.Thus, genetic compensation byHnrnph1might be sufficient to counteract the loss of hnRNPH2. These findings suggest that the pathogenesis ofHNRNPH2-related disorder in humans may be driven by a toxic gain of function or a complex loss ofHNRNPH2function with impaired compensation byHNRNPH1.The mutant knock-in mice described here are an important resource for preclinical studies to assess the potential benefit of either gene replacement or therapeutic knockdown of mutant hnRNPH2.
Temporal integration of rod signals in luminance and chromatic pathways
Authors: Méndez Iñaki Cormenzana , Martín Andrés , O’Donell Beatriz , Cao Dingcai , Barrionuevo Pablo A.
Published: March 2022

We assessed how rod excitation (R) affects luminance (L+M+S) and chromatic [L/(L+M)] reaction times (RTs). A four-primary display based on the overlapped images of two spectrally-modified monitors, which allowed specific or combined [L+M+S+R, L/(L+M) + R] photoreceptor stimulation, was used to present a C-target stimulus differing from the background only by the selected stimulation. For the luminance pathway, rod input increased RTs, suggesting a suppressive rod-cone interaction. The responses of the chromatic pathway were faster when rods were involved, suggesting a major role of rods in mesopic color perception.
Proteomic profiling identifies co-regulated expression of splicing factors as a characteristic feature of intravenous leiomyomatosis
Authors: Krasny Lukas , Wilding Chris P , Perkins Emma , Arthur Amani , Guljar Nafia , Jenks Andrew D , Fisher Cyril , Judson Ian , Thway Khin , Jones Robin L , Huang Paul H
Published: March 2022

Intravenous leiomyomatosis (IVLM) is a rare benign smooth muscle tumour that is characterised by intravenous growth in the uterine and pelvic veins. Previous DNA copy number and transcriptomic studies have shown that IVLM harbours unique genomic and transcriptomic alterations when compared to uterine leiomyoma (uLM), which may account for their distinct clinical behaviour. Here we undertake the first comparative proteomic analysis of IVLM and other smooth muscle tumours (comprising uLM, soft tissue leiomyoma and benign metastasising leiomyoma) utilising data-independent acquisition mass spectrometry. We show that, at the protein level, IVLM is defined by the unique co-regulated expression of splicing factors. In particular, IVLM is enriched in two clusters composed of co-regulated proteins from the hnRNP, LSm, SR and Sm classes of the spliceosome complex. One of these clusters (Cluster 3) is associated with key biological processes including nascent protein translocation and cell signalling by small GTPases. Taken together, our study provides evidence of co-regulated expression of splicing factors in IVLM compared to other smooth muscle tumours which suggests a possible role for alternative splicing in the pathogenesis of IVLM.
The link between autism and sex-specific neuroanatomy, and associated cognition and gene expression
Authors: Floris Dorothea L. , Peng Han , Warrier Varun , Lombardo Michael V. , Pretzsch Charlotte M. , Moreau Clara , Tsompanidis Alexandros , Gong Weikang , Mennes Maarten , Llera Alberto , van Rooij Daan , Oldehinkel Marianne , Forde Natalie , Charman Tony , Tillmann Julian , Banaschewski Tobias , Moessnang Carolin , Durston Sarah , Holt Rosemary J. , Ecker Christine , Dell’Acqua Flavio , Loth Eva , Bourgeron Thomas , Murphy Declan G. M. , Marquand Andre F. , Lai Meng-Chuan , Buitelaar Jan K. , Baron-Cohen Simon , Beckmann Christian F. ,   the EU-AIMS LEAP Group
Published: March 2022

Objectives

The male preponderance in autism spectrum conditions (ASC) prevalence is among the most pronounced sex ratios across different neurodevelopmental conditions. Here, we aimed to elucidate the relationship between autism and typical sex-differential neuroanatomy, cognition, and related gene expression.

Methods

Using a novel deep learning framework trained to predict biological sex, we compared sex prediction model performance across neurotypical and autistic males and females. Multiple large-scale datasets were employed at different stages of the analysis pipeline: a) Pre-training: the UK Biobank sample (>10.000 individuals); b) Transfer learning and validation: the ABIDE datasets (1,412 individuals, 5-56 years of age); c) Test and discovery: the EU-AIMS/AIMS-2-TRIALS LEAP dataset (681 individuals, 6-30 years of age) and d) Specificity: the Neuroimage and ADHD200 datasets (887 individuals, 7-26 years of age).

Results

Across both ABIDE and LEAP we showed that features positively predictive of neurotypical males were on average more predictive of autistic males (P=1.1e-23). Features positively predictive of neurotypical females were on average less predictive of autistic females (P=1.2e-22). These accuracy differences in autism were not observed in individuals with ADHD. In autistic females the male-shifted neurophenotype was further associated with poorer social sensitivity and emotional face processing while also with associated gene expression patterns of midgestational cell types.

Conclusions

Our results demonstrate a shift in both autistic male and female individuals’ neuroanatomy towards male-characteristic patterns associated with typically sex-differential, social cognitive features and related gene expression patterns. Findings hold promise for future research aimed at refining the quest for biological mechanisms underpinning the etiology of autism.
Selective Decontamination of the Digestive Tract in Invasively Ventilated Patients in an Intensive Care Unit: A protocol for a Systematic Review and Meta-Analysis
Authors: Hammond Naomi E , Myburgh John , Di Tanna Gian Luca , Garside Tessa , Vlok Ruan , Mahendran Sajeev , Adigbli Derick , Finfer Simon , Goodman Fiona , Guyatt Gordon , Venkatesh Balasubramanian , Seppelt Ian , Delaney Anthony
Published: March 2022

Introduction

The use of Selective Decontamination of the Digestive Tract (SDD) as a preventative infection-control strategy in invasively ventilated patients in the intensive care unit (ICU) remains low despite numerous randomised controlled trials (RCTs) consistently reporting reductions in interval mortality rates and shorter durations of mechanical ventilation. The Selective Decontamination of the Digestive Tract in the Intensive Care Unit (SuDDICU) cluster cross-over RCT, that includes over 5500 participants randomised to receive a standardised commercial grade SDD interventions or standard care, will be reported in 2022 and will add substantive weight to previous RCT data assessing the effect of SDD on interval mortality compared to standard care. We will conduct an updated systematic review and prospective aggregate data meta-analysis of previous conducted and published RCTs, developed using a protocol and statistical analysis plan completed prior to the completion of the SuDDICU RCT and including the SuDDICU data to present the most current evidence available to guide clinical practice.

Methods and analysis

We will include RCTs that compare the effect on hospital mortality and other patient-centred outcomes of treatment with SDD compared to standard care in invasively ventilated adults in the ICU. We will perform a search that includes the electronic databases MEDLINE and EMBASE and clinical trial registries. Two reviewers will independently screen titles and abstracts, perform full article reviews and extract study data, with discrepancies resolved by a third reviewer. We will report study characteristics and quantify risk of bias. We will perform random effects Bayesian meta-analyses to provide pooled estimates that SDD improves outcomes, whenever it is feasible to do so. We will evaluate overall certainty of evidence using the Grading of Recommendations Assessment, Development, and Evaluation framework.

Conclusion

This updated systematic review and prospective meta-analysis will provide clinicians with an expedited assessment of the totality of current evidence about the effect on mortality of using SDD in mechanically ventilated ICU patients.
Studies of a siderophore-producing cyclization domain: A refined proposal of substrate binding
Authors: Gnann Andrew D. , Xia Yuan , Soule Jess , Barthélemy Clara , Mawani Jayata , Musoke Sarah Nzikoba , Castellano Brian , Brignole Edward , Frueh Dominique , Dowling Daniel P.
Published: March 2022

Nonribosomal peptide synthetase (NRPS) heterocyclization (Cy) domains generate biologically important ox-/thiazoline modifications in natural products, including in production of compounds targeting disease or siderophores that are important for bacterial pathogenicity. Cy domains share the NRPS condensation domain fold but catalyze consecutive condensation and cyclodehydration reactions via an unknown mechanism. To further understanding of Cy domain catalysis, we report the crystal structure of the second Cy domain (Cy2) of yersiniabactin synthetase from the causative agent of the plague, Yersinia pestis. We find the high-resolution structure of Cy2 adopts a conformation enabling exploration of binding the extended, thiazoline-containing cyclodehydration intermediate for catalysis and the acceptor carrier protein to which it is tethered. We also report complementary electrostatic interfaces between Cy2 and its donor carrier protein that mediate donor binding. Lastly, we explore domain flexibility through the normal mode approximation and identify small-molecule fragment binding sites to inform antibiotic design targeting Cy function. Our results suggest how carrier protein binding may influence global conformation, with consequences for active site catalytic states and inhibitor development.
Efficient compartmentalization in insect bacteriomes protects symbiotic bacteria from host immune system
Authors: Ferrarini Mariana Galvão , Dell’Aglio Elisa , Vallier Agnès , Balmand Séverine , Vincent-Monégat Carole , Hughes Sandrine , Gillet Benjamin , Parisot Nicolas , Zaidman-Rémy Anna , Vieira Cristina , Heddi Abdelaziz , Rebollo Rita
Published: March 2022

Background

Many insects house symbiotic intracellular bacteria (endosymbionts) that provide them with essential nutrients, thus promoting usage of nutrient-poor habitats. Endosymbiont seclusion within host specialized cells, called bacteriocytes, often organized in a dedicated organ, the bacteriome, is crucial in protecting them from host immune defenses while avoiding chronic host immune activation. Previous evidence obtained in the cereal weevil Sitophilus oryzae has shown that bacteriome immunity is activated against invading pathogens, suggesting endosymbionts might be targeted and impacted by immune effectors during an immune challenge. To pinpoint any molecular determinants associated with such challenges, we conducted a dual transcriptomic analysis of S. oryzae’s bacteriome subjected to immunogenic peptidoglycan fragments.

Results

We show that upon immune challenge the bacteriome actively participates in the innate immune response via an induction of antimicrobial peptides (AMPs). Surprisingly, endosymbionts do not undergo any transcriptomic changes, indicating that this potential threat goes unnoticed. Immunohistochemistry showed that TCT-induced AMPs are located outside the bacteriome, excluding direct contact with the endosymbionts.

Conclusions

This work demonstrates that endosymbiont protection during an immune challenge is mainly achieved by efficient confinement within bacteriomes, which provides physical separation between host systemic response and endosymbionts.
Fragment libraries designed to be functionally diverse recover protein binding information more efficiently than standard structurally diverse libraries
Authors: Carbery Anna , Skyner Rachael , von Delft Frank , Deane Charlotte M.
Published: March 2022

Current fragment-based drug design relies on the efficient exploration of chemical space though the use of structurally diverse libraries of small fragments. However, structurally dissimilar compounds can exploit the same interactions on a target, and thus be functionally similar. Using 3D structures of many fragments bound to multiple targets, we examined if there exists a better strategy for selecting fragments for screening libraries. We show that structurally diverse fragments can be described as functionally redundant, often making the same interactions. Ranking fragments by the number of novel interactions they made, we show that functionally diverse selections of fragments substantially increase the amount of information recovered for unseen targets compared to other methods of selection. Using these results, we design small functionally efficient libraries that are able to give significantly more information about new protein targets than similarly sized structurally diverse libraries. By covering more functional space (rather than chemical space), more diverse sets of drug leads can be generated, increasing the chances of fragment screens resulting in viable drug candidates.
A deep learning classification task for brain navigation during functional ultrasound imaging
Authors: Lambert Théo , Brunner Clément , Kil Dries , Wuyts Roel , D’Hondt Ellie , Montaldo Gabriel , Urban Alan
Published: March 2022

Positioning and navigation are essential components of neuroimaging as they improve the quality and reliability of data acquisition, leading to advances in diagnosis, treatment outcomes, and fundamental understanding of the brain. Functional ultrasound (fUS) imaging is an emerging technology providing high-resolution images of the brain vasculature, allowing for the monitoring of brain activity. However, as the technology is relatively new, there is no standardized tool for inferring the position in the brain from the vascular images. This study presents a deep learning-based framework designed to address this challenge. Our approach uses an image classification task coupled with a regression on the resulting probabilities to determine the position of a single image. We conducted experiments using a dataset of 51 rat brain scans to evaluate its performance. The training positions were extracted at intervals of 375 µm, resulting in a positioning error of 176 µm. Further GradCAM analysis revealed that the predictions were primarily driven by subcortical vascular structures. Finally, we assessed the robustness of our method in a cortical stroke where the brain vasculature is severely impaired. Remarkably, no specific increase in the number of misclassifications was observed, confirming the method’s reliability in challenging conditions. Overall, our framework provides accurate and flexible positioning, not relying on a pre-registered reference but on conserved vascular patterns.
Integrated single-cell and spatial transcriptomic analyses unravel the heterogeneity of the prostate tumor microenvironment
Authors: Hirz Taghreed , Mei Shenglin , Sarkar Hirak , Kfoury Youmna , Wu Shulin , Verhoeven Bronte M. , Subtelny Alexander O. , Zlatev Dimitar V. , Wszolek Matthew W. , Salari Keyan , Murray Evan , Chen Fei , Macosko Evan Z. , Wu Chin-Lee , Scadden David T. , Dahl Douglas M. , Baryawno Ninib , Saylor Philip J. , Kharchenko Peter V. , Sykes David B.
Published: March 2022

The treatment of primary prostate cancer delicately balances an active surveillance approach for low-risk disease with multimodal treatment including surgery, radiation therapy, and hormonal therapy for high-risk disease. Recurrence and development of metastatic disease remains a clinical problem, without a clear understanding of what drives immune escape and tumor progression. Here, we sought to comprehensively describe the tumor microenvironment of localized prostate cancer contrasting this with adjacent normal samples and healthy controls. We performed single-cell RNA sequencing and high-resolution spatial transcriptomic analysis. This revealed tumor context dependent changes in gene expression. Our data point towards an immune suppressive tumor microenvironment associated with suppressive myeloid populations and exhausted T-cells, in addition to high stromal angiogenic activity. We inferred cell-to-cell relationships at an unprecedented scale for ligand-receptor interactions within undissociated tissue sections. Our work provides a highly detailed and comprehensive resource of the prostate tumor microenvironment as well as tumor-stromal cell interactions.

Highlights

Characterization of prostate cancer by combined scRNA-seq and spatial transcriptomic analysis

Primary prostate cancer establishes a suppressive immune microenvironment

The prostate tumor microenvironment exhibits a high angiogenic gene expression pattern

A new computational analysis pipeline to deconvolute context-specific differential gene expression
Mapping the Cell-Membrane Proteome to the Cancer Hallmarks
Authors: Lazar Iulia M. , Karcini Arba , Haueis Joshua R. S.
Published: March 2022

The hallmarks of biological processes that underlie the development of cancer have been long recognized, yet, existing therapeutic treatments cannot prevent cancer from continuing to be one of the leading causes of death worldwide. This work was aimed at exploring the extent to which the cell-membrane proteins are implicated in triggering cancer hallmark processes, and assessing the ability to pinpoint novel therapeutic targets through a combined membrane proteome/cancer hallmark perspective. By using GO annotations, a database of human proteins associated broadly with ten cancer hallmarks was created. Cell-membrane cellular subfractions of SKBR3/HER2+ breast cancer cells used as a model system were analyzed by high resolution mass spectrometry, and high-quality proteins (FDR<3 %) identified by at least two unique peptides were mapped to the cancer hallmark database. Over 1100 experimentally detected cell-membrane or cell-membrane associated proteins, representing ~8 % of the human cell-membrane proteome, were mapped to the hallmark database. Representative membrane constituents such as receptors, CDs, adhesion and transport proteins were distributed over the entire genome and present in every hallmark category. Sustained proliferative signaling/cell cycle, adhesion/tissue invasion, and evasion of immune destruction emerged as prevalent hallmarks represented by the membrane proteins. Construction of protein-protein interaction networks uncovered a high level of connectivity between the hallmark members, with some receptor (EGFR, ERBB2, FGFR, MTOR, CSF1R), antigen (CD44), and adhesion (MUC1) proteins being implicated in most hallmark categories. An illustrative subset of 116 hallmark proteins that included 44 oncogenes, 28 tumor suppressors, and 38 approved drug targets was subjected to a more in-depth analysis. The existing drug targets were implicated mainly in signaling processes. Network centrality analysis revealed that nodes with high degree, rather than betweenness, represent a good resource for informing the selection of putative novel drug targets. Through heavy involvement in supporting cancer hallmark processes, we show that the functionally diverse and networked landscape of cancer cell-membrane proteins fosters unique opportunities for guiding the development of novel therapeutic interventions, including multi-agent, immuno-oncology and precision medicine applications.
Disrupting the plastid-hosted iron-sulfur cluster biogenesis pathway in Toxoplasma gondii has pleiotropic effects irreversibly impacting parasite viability
Authors: Renaud Eléa A. , Pamukcu Sarah , Cerutti Aude , Berry Laurence , Lemaire-Vieille Catherine , Yamaryo-Botté Yoshiki , Botté Cyrille Y. , Besteiro Sébastien
Published: March 2022

Like many other apicomplexan parasites, Toxoplasma gondii contains a plastid harbouring key metabolic pathways, including the SUF pathway that is involved in the biosynthesis of iron-sulfur clusters. These cofactors are key for a variety of proteins involved in important metabolic reactions, potentially including plastidic pathways for the synthesis of isoprenoid and fatty acids. It was shown previously that impairing the NFS2 cysteine desulfurase, involved in the first step of the SUF pathway, leads to an irreversible killing of intracellular parasites. However, the metabolic impact of disrupting the pathway remained unexplored. We have generated another mutant of the pathway, deficient for the SUFC ATPase, and we have investigated in details the phenotypic consequences of TgNFS2 and TgSUFC depletion on parasite homeostasis. Our analysis confirms that Toxoplasma SUF mutants are severely and irreversibly impacted in growth: cell division and membrane homeostasis are particularly affected. Lipidomic analysis suggests a defect in apicoplast-generated fatty acids, along with a simultaneous increase in scavenging of host-derived lipids. However, addition of exogenous lipids did not allow full restauration of growth, suggesting other more important cellular functions were impacted in addition to fatty acid synthesis. For instance, we have shown that the SUF pathway is also key for generating isoprenoid-derived precursors necessary for the proper targeting of GPI-anchored proteins as well as for the parasite gliding motility. Thus, plastid-generated iron-sulfur clusters support the functions of proteins involved in several vital downstream cellular pathways, which implies the SUF machinery may be explored for discovering new potential anti-Toxoplasma targets.
Drivers of nematode diversity in forest soils across climatic zones
Authors: Shao Yuanhu , Wang Zuyan , Liu Tao , Kardol Paul , Ma Chengen , Hu Yonghong , Cui Yang , Zhao Cancan , Zhang Weixin , Guo Dali , Fu Shenglei
Published: March 2022

Nematodes are the most abundant multi-cellular animals in soil, influencing key processes and functions in terrestrial ecosystems. Yet, little is known about the drivers of nematode abundance and diversity in forest soils across climatic zones. This is despite forests cover approximately 30% of the earth’s land surface, provide many crucial ecosystem services but strongly vary in hydrothermal conditions and associated ecosystem properties across climatic zones.

Here, we collected nematode samples from 13 forests across a latitudinal gradient. We divided this gradient in temperate, warm-temperate, and tropical climatic zones. Using boosted regression trees, we showed that across the gradient, nematode abundance and diversity were mainly influenced by soil organic carbon. However, within climatic zones, other factors were more important in driving nematode alpha-diversity, nematode biomass and the abundance of different trophic groups: mean annual temperature and total soil phosphorus in temperate zones, soil pH in warm-temperate zones, and mean annual precipitation in tropical zones. Additionally, nematode beta-diversity was higher in temperate than in warm-temperate and tropical zones, and we did not find significant differences among climatic zones in nematode gamma diversity.

Together, our findings indicate a latitudinal shift in the main climatic variables controlling soil nematode communities and demonstrate that the drivers of soil nematode diversity in forested ecosystems are affected by the spatial scale and climatic conditions considered. This implies that high resolution studies are needed to accurately predict how soil functions respond if climate conditions move beyond the coping range of soil organisms. Our results also emphasize the importance of studying the area-diversity relationship of soil organisms under different climatic conditions.
VolumesJ: A new method and tool for volumetric estimation of brain structures after serial sectioning
Authors: Ferreiro Elisabete , Rodríguez-Iglesias Noelia , Cardoso João , Valero Jorge
Published: March 2022

Volume estimations are crucial for many neuroscience studies, allowing the evaluation of changes in the size of brain areas that may have relevant functional consequences. Classical histological methods and modern human brain imaging techniques rely on obtaining physical or digital sections, with a known thickness, of the organ to be analyzed. This “slicing” strategy is associated with an ineludible loss of information about the three-dimensional organization of the analyzed structures, especially affecting the precision of volumetric measurements. To overcome this problem, several methods have been developed. One of the most commonly used approaches for volume estimation is the classical Cavalieri’s method.

Within this book chapter, we provide first an overview of Cavalieri’s method and propose a new one, named the Truncated Cone Shape (TCS) method, for the estimation of volumes from tissue sections. Second, we compare the accuracy of both methods using computer-generated objects of different shapes and sizes. We conclude that, more frequently, the TCS method provides a better estimate of real volumes than Cavalieri’s method. And third, we describe a protocol to estimate volumes using a self-developed and freely available tool for ImageJ: VolumestJ (https://github.com/Jorvalgl/VolumestJ). This new tool helps to implement both Cavalieri’s and TCS methods using digital images of tissue sections. We consider that VolumestJ will facilitate the labor of researchers interested in volume estimations.
Dependence on a variable residue limits the breadth of an HIV MPER neutralizing antibody, despite convergent evolution with broadly neutralizing antibodies
Authors: Scheepers Cathrine , Kgagudi Prudence , Mzindle Nonkululeko , Gray Elin S. , Moyo-Gwete Thandeka , Lambson Bronwen E. , Oosthuysen Brent , Mabvakure Batsirai , Garrett Nigel J. , Abdool Karim Salim S. , Morris Lynn , Moore Penny L.
Published: March 2022

Broadly neutralizing antibodies (bNAbs) that target the membrane-proximal external region (MPER) of HIV gp41 envelope, such as 4E10 and VRC42.01, can neutralize >95% of viruses. These two MPER-directed monoclonal antibodies share germline antibody genes (IGHV1-69 and IGKV3-20) and form a bNAb epitope class. Furthermore, convergent evolution within these two lineages towards a 111.2GW111.3 motif in the CDRH3 is known to enhance neutralization potency. We have previously isolated an MPER neutralizing antibody, CAP206-CH12, that uses these same germline heavy and light chain genes but lacks breadth (neutralizing only 6% of heterologous viruses). Longitudinal sequencing of the CAP206-CH12 lineage over three years revealed similar convergent evolution towards 111.2GW111.3 among some lineage members. Mutagenesis of CAP206-CH12 from 111.2GL111.3 to 111.2GW111.3 modestly improved neutralization breadth (by 9%) and potency (3-fold), but did not reach the levels of breadth and potency of VRC42.01 and 4E10. To explore the lack of potency/breadth, viral mutagenesis was performed to map the CAP206-CH12 epitope. This indicated that CAP206-CH12 is dependent on D674, a highly variable residue at the solvent-exposed elbow of MPER. In contrast, VRC42.01 and 4E10 were dependent on highly conserved residues (W672, F673, T676, and W680) facing the hydrophobic patch of the MPER. Therefore, while CAP206-CH12, VRC42.01, and 4E10 share germline genes and show some evidence of convergent evolution, their dependence on different amino acids, which impacts orientation of binding to the MPER, result in differences in breadth and potency. These data have implications for the design of HIV vaccines directed at the MPER epitope.

Author Summary

Germline-targeting immunogens are a promising HIV vaccine design strategy. This approach is reliant on the identification of broadly neutralizing antibody (bNAb) classes, which use the same germline antibody genes to target the same viral epitopes. Here, we compare three HIV Envelope MPER-directed antibodies (4E10, VRC42.01 and CAP206-CH12) that despite having shared antibody genes, show distinct neutralization profiles. We show that CAP206-CH12 targets a single highly variable residue in the MPER, which results in low neutralization breadth. In contrast, the 4E10 and VRC42.01 mAbs are dependent on highly conserved residues in the MPER, resulting in exceptional neutralization breadth. Our data suggest that while shared germline genes within bNAb epitope classes are required, in some cases these are not sufficient to produce neutralization breadth, and MPER immunogens will need to trigger responses to conserved sites.
Dynamic control of metabolic zonation and liver repair by endothelial cell Wnt2 and Wnt9b revealed by single cell spatial transcriptomics using Molecular Cartography
Authors: Hu Shikai , Liu Silvia , Bian Yu , Poddar Minakshi , Singh Sucha , Cao Catherine , McGaughey Jackson , Bell Aaron , Blazer Levi L , Adams Jarret J , Sidhu Sachdev S , Angers Stephane , Monga Satdarshan P.
Published: March 2022

The conclusive identity of Wnt proteins regulating liver zonation (LZ) and regeneration (LR) remains unclear despite an undisputed role of β-catenin. Using single-cell analysis of liver cells from various species, a conserved Wnt2 and Wnt9b expression in endothelial cells (ECs) in zone 3 shown to be the major Wnt cell source, was identified. Conditional EC-elimination of Wnt2 and Wnt9b led to perturbation of LZ with not only loss of β-catenin targets in zone 3, but also re-appearance of zone 1 genes in zone 3, unraveling dynamicity as revealed by single-cell spatial transcriptomics using Molecular Cartography. Defective LR observed in the knockouts phenocopied other models of defective hepatic Wnt signaling. Administration of a tetravalent antibody to activate Wnt signaling rescued LZ and LR in the knockouts. Molecular Cartography on the livers of the agonist-treated animal revealed changes in LZ. Administration of the agonist also promoted LR in acetaminophen overdose acute liver failure (ALF) fulfilling an unmet clinical need. Overall, we report an unequivocal role of EC-Wnt2 and Wnt9b in LZ and LR and show the role of Wnt activators as regenerative therapy for ALF.