bluuebunny/crossref_metadata_2025 huggingface dataset:
Negative autoregulation mitigates collateral RNase activity of repeat-targeting CRISPR-Cas13d in mammalian cells
Authors: Kelley Chase P , Haerle Maja C , Wang Eric T
Published: December 2021

Cas13 is a unique family of CRISPR endonucleases exhibiting programmable binding and cleavage of RNAs and is a strong candidate for eukaryotic RNA knockdown in the laboratory and the clinic. However, sequence-specific binding of Cas13 to the target RNA unleashes non-specific bystander RNA cleavage, or collateral activity, which may confound knockdown experiments and raises concerns for therapeutic applications. Although conserved across orthologs and robust in cell-free and bacterial environments, the extent of collateral activity in mammalian cells remains disputed. Here, we investigate Cas13d collateral activity in the context of an RNA-targeting therapy for myotonic dystrophy type 1, a disease caused by a transcribed long CTG repeat expansion. We find that when targeting CUGn RNA in HeLa and other cell lines, Cas13d depletes endogenous and transgenic RNAs, interferes with critical cellular processes, and activates stress response and apoptosis pathways. We also observe collateral effects when targeting other repetitive and unique transgenic sequences, and we provide evidence for collateral activity when targeting highly expressed endogenous transcripts. To minimize collateral activity for repeat-targeting Cas13d therapeutics, we introduce gRNA excision for negative-autoregulatory optimization (GENO), a simple strategy that leverages crRNA processing to control Cas13d expression and is easily integrated into an AAV gene therapy. We argue that thorough assessment of collateral activity is necessary when applying Cas13d in mammalian cells and that implementation of GENO illustrates the advantages of compact and universally robust regulatory systems for Cas-based gene therapies.
Geminivirus C4 proteins inhibit GA signaling via prevention of NbGAI degradation, to promote viral infection and symptom development in N. benthamiana
Authors: Li Pengbai , Guo Liuming , Lang Xinyuan , Li Mingjun , Wu Gentu , Wu Rui , Wang Lvxin , Zhao Meisheng , Qing Ling
Published: December 2021

Phytohormone gibberellin (GA) is an important plant signaling molecule that regulates plant growth and defense against abiotic and biotic stresses. To date, the molecular mechanism of the plant responses to viral infection mediated by GA is still undetermined. DELLA is a repressor of GA signaling and is recognized by the F-box protein, a component of the SCFSLY1/GID2 complex. The recognized DELLA is degraded by the ubiquitin-26S proteasome, leading to the activation of the GA signaling. Here, we report that ageratum leaf curl Sichuan virus (ALCScV)-infected N. benthamiana plants showed dwarfing symptom and abnormal flower development. The infection of ALCScV alters the expressions of GA pathway-related genes and decreases the content of endogenous GA significantly in N. benthamiana. Further, ALCScV-encoded C4 protein interacts with the DELLA protein NbGAI, and interferes with the interaction between NbGAI and NbGID2 to prevent the degradation of NbGAI, leading to the inhibition of the GA signaling pathway. Silencing of NbGAI or exogenous GA3 treatment significantly reduces viral accumulation and disease symptoms in N. benthamiana plants. The same results were proved by the experiments with C4 protein encoded by tobacco curly shoot virus (TbCSV). Therefore, we propose a novel mechanism of geminivirus C4 proteins controling virus infection and disease symptom development through interfering GA signaling pathway.

Author Summary

Gibberellins (GAs) are plant hormones that are essential for many developmental processes in plants. It has indicated that plant virus infection can induce abnormal flower development and influence GA pathway resulting the plant dwarfing symptom, but the underlying mechanisms is still not well described. Here, we demonstrate that geminivirus - encoded C4 protein regulates the GA signaling pathway to promote viral accumulation and disease symptom development. Through directly interacting with NbGAI, the C4 protein interferes with the interaction between NbGAI and NbGID2, which inhibits the degradation of NbGAI. As a result, the GA signaling pathway is blocked, and the infected plants display symptoms of typical dwarfing and delayed flowering. Our results reveal a novel mechanism by which geminivirus C4 proteins influence viral pathogenicity via interfering the GA signaling pathway, and provide new insights into the interaction between virus and host.
hapCon: Estimating contamination of ancient genomes by copying from reference haplotypes
Authors: Huang Yilei , Ringbauer Harald
Published: December 2021

Motivation

Human ancient DNA (aDNA) studies have surged in recent years, revolutionizing the study of the human past. Typically, aDNA is preserved poorly, making such data prone to contamination from other human DNA. Therefore, it is important to rule out substantial contamination before proceeding to downstream analysis. As most aDNA samples can only be sequenced to low coverages (<1x average depth), computational methods that can robustly estimate contamination in the low coverage regime are needed. However, the ultra low-coverage regime (0.1x and below) remains a challenging task for existing approaches.

Results

We present a new method to estimate contamination in aDNA for male individuals. It utilizes a Li&Stephen’s haplotype copying model for haploid X chromosomes, with mismatches modelled as genotyping error or contamination. We assessed an implementation of this new approach, hapCon, on simulated and down-sampled empirical aDNA data. Our results demonstrate that hapCon outperforms a commonly used tool for estimating male X contamination (ANGSD), with substantially lower variance and narrower confidence intervals, especially in the low coverage regime. We found that hapCon provides useful contamination estimates for coverages as low as 0.1x for SNP capture data (1240k) and 0.02x for whole genome sequencing data (WGS), substantially extending the coverage limit of previous male X chromosome based contamination estimation methods.

Availability and Implementation

A implementation of our software (hapCON) using Python and C has been deposited at https://github.com/hyl317/hapROH. We make hapCon available as part of a python package (hapROH), which is available at the Python Package Index (https://pypi.org/project/hapROH) and can be installed via pip. The documentation provides example use cases as blueprints for custom applications (https://haproh.readthedocs.io).
“Prefrontal cortical contributions to working memory loading, maintenance and recall are parsed by hippocampal-prefrontal oscillatory assembly dynamics”
Authors: Domanski Aleksander P.F. , Kucewicz Michal T. , Russo Eleonora , Tricklebank Mark D. , Robinson Emma S.J. , Durstewitz Daniel , Jones Matt W.
Published: December 2021

Working memory enables incorporation of recent experience into subsequent decision-making. This processing recruits both prefrontal cortex and hippocampus, where neurons encode task cues, rules and outcomes. However, precisely which information is carried when, and by which neurons, remains unclear. Using population decoding of activity in rat medial prefrontal cortex (mPFC) and dorsal hippocampal CA1, we confirm that mPFC populations lead in maintaining sample information across delays of an operant nonmatch to sample task, despite individual neurons firing only transiently. During sample encoding, distinct mPFC subpopulations joined distributed CA1-mPFC cell assemblies hallmarked by 4-5Hz rhythmic modulation; CA1-mPFC assemblies re-emerged during choice episodes, but were not 4-5Hz modulated. Delay-dependent errors arose when attenuated rhythmic assembly activity heralded collapse of sustained mPFC encoding; pharmacological disruption of CA1-mPFC assembly rhythmicity impaired task performance. Our results map component processes of memory-guided decisions onto heterogeneous CA1-mPFC subpopulations and the dynamics of physiologically distinct, distributed cell assemblies.
Different excitation-inhibition correlations between spontaneous and tone-evoked activity in primary auditory cortex neurons
Authors: Chew Katherine C. M. , Kumar Vineet , Tan Andrew Y. Y.
Published: December 2021

Tone-evoked synaptic excitation and inhibition are highly correlated in many neurons with V-shaped tuning curves in the primary auditory cortex of pentobarbital-anesthetized rats. In contrast, there is less correlation between spontaneous excitation and inhibition in visual cortex neurons under the same anesthetic conditions. However, it was not known whether the primary auditory cortex resembles visual cortex in having spontaneous excitation and inhibition that is less correlated than tone-evoked excitation and inhibition. Here we report whole-cell voltage-clamp measurements of spontaneous excitation and inhibition in primary auditory cortex neurons of pentobarbital-anesthetized rats. The larger excursions of both spontaneous excitatory and inhibitory currents appeared to consist of distinct events, with the inhibitory event rate typically lower than the excitatory event rate. We use the ratio of the excitatory event rate to the inhibitory event rate, and the assumption that the excitatory and inhibitory synaptic currents can each be reasonably described as a filtered Poisson process, to estimate the maximum spontaneous excitatory-inhibitory correlation for each neuron. In a subset of neurons, we also measured tone-evoked excitation and inhibition. In neurons with V-shaped tuning curves, although tone-evoked excitation and inhibition were highly correlated, the spontaneous inhibitory event rate was typically sufficiently lower than the spontaneous excitatory event rate to indicate a lower excitatory-inhibitory correlation for spontaneous activity than for tone-evoked responses.
Genetic and compound screens uncover factors modulating cancer cell response to indisulam
Authors: Pogacar Ziva , Groot Kelvin , Jochems Fleur , Dos Santos Dias Matheus , Morris Ben , Roosen Mieke , Wardak Leyma , De Conti Giulia , Velds Arno , Lieftink Cor , Beijersbergen Roderick L. , Bernards René , de Oliveira Rodrigo Leite
Published: December 2021

Discovering biomarkers of drug response and finding powerful drug combinations can support the reuse of previously abandoned cancer drugs in the clinic. Indisulam is an abandoned drug that acts as a molecular glue, inducing degradation of splicing factor RBM39 through interaction with CRL4DCAF15. Here, we performed genetic and compound screens to uncover factors mediating indisulam sensitivity and resistance. First, a dropout CRISPR screen identified SRPK1 loss as a synthetic lethal interaction with indisulam that can be exploited therapeutically by the SRPK1 inhibitor SPHINX31. Moreover, a CRISPR resistance screen identified components of the degradation complex that mediate resistance to indisulam: DCAF15, DDA1, and CAND1. Lastly, we show that cancer cells readily acquire spontaneous resistance to indisulam. Upon acquiring indisulam resistance, pancreatic cancer (Panc10.05) cells still degrade RBM39 and are vulnerable to BCL-xL inhibition. The better understanding of the factors that influence the response to indisulam can assist rational reuse of this drug in the clinic.
Single cell RNA sequencing and lineage tracing confirm mesenchyme to epithelial transformation (MET) contributes to repair of the endometrium at menstruation
Authors: Kirkwood Phoebe M , Gibson Douglas A , Shaw Isaac , Dobie Ross , Kelepouri Olympia , Henderson Neil C , Saunders Philippa TK
Published: December 2021

The human endometrium experiences repetitive cycles of tissue wounding characterised by piecemeal shedding of the surface epithelium and rapid restoration of tissue homeostasis. In this study we used a mouse model of endometrial repair and three transgenic lines of mice to investigate whether epithelial cells that become incorporated into the newly formed luminal epithelium have their origins in one or more of the mesenchymal cell types present in the stromal compartment of the endometrium. Using scRNAseq we identified a novel population of PDGFRb+ mesenchymal stromal cells that developed a unique transcriptomic signature in response to endometrial breakdown/repair. These cells expressed genes usually considered specific to epithelial cells and in silico trajectory analysis suggested they were stromal fibroblasts in transition to becoming epithelial cells. To confirm our hypothesis we used a lineage tracing strategy to compare the fate of stromal fibroblasts (PDGFRa+) and stromal perivascular cells (NG2/CSPG4+). We demonstrated that stromal fibroblasts can undergo a mesenchyme to epithelial transformation and become incorporated into the re-epithelialised luminal surface of the repaired tissue. This study is the first to discover a novel population of wound-responsive, plastic endometrial stromal fibroblasts that contribute to the rapid restoration of an intact luminal epithelium during endometrial repair. These findings form a platform for comparisons both to endometrial pathologies which involve a fibrotic response (Ashermans syndrome, endometriosis) as well as other mucosal tissues which have a variable response to wounding.

Highlights

Single cell RNA sequencing identified a population of PDGFRβ+ mesenchymal cells with a unique transcriptomic signature that arises in response to induction of a menses-like wound

The repair-specific mesenchymal cells express genes considered specific to both mesenchymal and epithelial lineages indicative of an intermediate phenotype and a mesenchymal to epithelial transition (MET)

in silicotrajectory analysis revealed that repair-specific cells originate from the fibroblast cell clusters, were distinct from perivascular cells, and had a predicted trans-differentiation trajectory towards definitive epithelial cells

in vivolineage tracing in adult mice provides definitive evidence that PDGFRα+ endometrial fibroblasts, and not NG2+ perivascular cells, undergo MET and can become incorporated into the luminal epithelium of the post repair tissue
The adolescent pattern of sleep spindle development revealed by HD-EEG polysomnography
Authors: Bocskai Gábor , Pótári Adrián , Gombos Ferenc , Kovács Ilona
Published: December 2021

Sleep spindles are developmentally relevant cortical oscillatory patterns; however, they have mostly been studied by considering the entire spindle frequency range (11 to 15 Hz) without a distinction between the functionally and topographically different slow and fast spindles, using relatively few electrodes and analysing wide age-ranges. Here, we employ HD-EEG polysomnography in three age-groups between 12 to 20 years of age, with an equal distribution between the two genders, and analyse the adolescent developmental pattern of the four major parameters of slow and fast sleep spindles. Most of our findings corroborate those very few previous studies that also make a distinction between slow and fast spindles in their developmental analysis. We find spindle frequency increasing with age, although spindle density change is not obvious in our study. We confirm the declining tendencies for amplitude and duration, although within narrower, more specific age-windows than previously. Spindle frequency seems to be higher in females in the oldest age-group. Based on the pattern of our findings, we suggest that HD-EEG, specifically targeting slow and fast spindle ranges and relatively narrow age-ranges would advance the understanding of both adolescent development and the functional relevance of sleep spindles in general.
Informeasure: an R/Bioconductor package to quantify nonlinear dependence between variables in biological networks from an information theory perspective
Authors: Pan Chu , Jing Limei , Luo Jiawei , Zeng Xiangxiang
Published: December 2021

Using information measures to infer biological regulatory networks can observe nonlinear relationship between variables, but it is computationally challenging and there is currently no convenient tool available. We here describe an information theory R package named Informeasure that devotes to quantifying nonlinear dependence between variables in biological regulatory networks from an information theory perspective. This package compiles most of the information measures currently available: mutual information, conditional mutual information, interaction information, partial information decomposition and part mutual information. The first estimator is used to infer bivariate networks while the last four estimators are dedicated to analysis of trivariate networks. The base installation of this turn-key package allows users to approach these information measures out of the box. Informeasure is implemented in R program and is available as an R/Bioconductor package at https://bioconductor.org/packages/Informeasure.
Structural and biophysical characterization of HNF-1A as a tool to study MODY3 diabetes variants
Authors: Kind Laura , Raasakka Arne , Molnes Janne , Aukrust Ingvild , Bjørkhaug Lise , Njølstad Pål Rasmus , Kursula Petri , Arnesen Thomas
Published: December 2021

Hepatocyte nuclear factor 1A (HNF-1A) is a transcription factor expressed in several embryonic and adult tissues, modulating expression of numerous target genes. Pathogenic variants in theHNF1Agene cause maturity-onset diabetes of the young 3 (MODY3 orHNF1AMODY), characterized by dominant inheritance, age of onset before 25-35 years of age, and pancreatic β-cell dysfunction. A precise diagnosis alters management as insulin can be exchanged with sulfonylurea tablets and genetic counselling differs from polygenic forms of diabetes. More knowledge on mechanisms of HNF-1A function and the level of pathogenicity of the numerousHNF1Avariants identified by exome sequencing is required for precise diagnostics. Here, we have structurally and biophysically characterized an HNF-1A protein containing both the DNA binding domain and the dimerization domain. We also present a novel approach to characterize HNF-1A variants. The folding and DNA binding capacity of two established MODY3 HNF-1A variant proteins (P112L, R263C) and one variant of unknown significance (N266S) were determined. All three variants showed reduced functionality compared to the wild-type protein. While the R263C and N266S variants displayed reduced binding to an HNF-1A target promoter, the P112L variant was unstablein vitroand in cells. Our results support and mechanistically explain disease causality for all investigated variants and allow for the dissection of structurally unstable and DNA binding defective variants. This points towards structural and biochemical investigation of HNF-1A being a valuable aid in reliable variant classification needed for precision diagnostics and management.
BISoN: A Bayesian Framework for Inference of Social Networks
Authors: Hart Jordan D. A. , Weiss Michael N. , Franks Daniel W. , Brent Lauren J. N.
Published: December 2021

Social networks are often constructed from point estimates of edge weights. In many contexts, edge weights are inferred from observational data, and the uncertainty around estimates can be affected by various factors. Though this has been acknowledged in previous work, methods that explicitly quantify uncertainty in edge weights have not yet been widely adopted, and remain undeveloped for many common types of data. Furthermore, existing methods are unable to cope with some of the complexities often found in observational data, and do not propagate uncertainty in edge weights to subsequent statistical analyses.

We introduce a unified Bayesian framework for modelling social networks based on observational data. This framework, which we call BISoN, can accommodate many common types of observational social data, can capture confounds and model effects at the level of observations, and is fully compatible with popular methods used in social network analysis.

We show how the framework can be applied to common types of data and how various types of downstream statistical analyses can be performed, including non-random association tests and regressions on network properties.

Our framework opens up the opportunity to test new types of hypotheses, make full use of observational datasets, and increase the reliability of scientific inferences. We have made example R scripts available to enable adoption of the framework.
Profiles and Dynamics of the Transcriptome of Microglial Cells Reveal their Inflammatory Status
Authors: Zohar Keren , Lezmi Elyad , Reichert Fanny , Eliyahu Tsiona , Rotshenker Shlomo , Weinstock Marta , Linial Michal
Published: December 2021

The primary role of microglia in the maintenance of brain homeostasis is to respond to disturbances in the microenvironment. In this study, we cultured murine neonatal microglia and activated them with lipopolysaccharide (LPS) and benzoyl ATP (bzATP) to characterize changes in the transcriptome in response to variousin vivostimuli caused by pathogens, injury, or toxins. Activation by bzATP, an agonist of purinergic receptors, induces a transient wave of transcriptional changes. However, a long-lasting transcriptional profile affecting thousands of genes occurs only following a combination of bzATP and LPS. This profile is dominated by a coordinated induction of cytokines (e.g., IL1-α and IL1-β), chemokines, and their direct regulators. Many of these inflammatory-related genes are up-regulated by several orders of magnitude. We identified the TNF-α and NF-κB pathways as the principal hubs for signaling of interleukin and chemokine induction in this cell system. We propose that primary microglia under controlled activation paradigms can be used for testing reagents that could attenuate their activated state. Such a microglial system could serve as a model for changes occurring in the aging brain and neurodegenerative diseases.

Highlight points

*

Primary murine microglia cultures release cytokines following activation with bzATP and LPS

*

The wave of changes in gene expression by bzATP is transient.

*

bzATP+LPS causes a transcription program dominated by the induction of interleukins and chemokines.
Background selection under evolving recombination rates
Authors: Booker Tom R. , Payseur Bret A. , Tigano Anna
Published: December 2021

Background selection (BGS), the effect that purifying selection exerts on sites linked to deleterious alleles, is expected to be ubiquitous across eukaryotic genomes. The effects of BGS reflect the interplay of the rates and fitness effects of deleterious mutations with recombination. A fundamental assumption of BGS models is that recombination rates are invariant over time. However, in some lineages recombination rates evolve rapidly, violating this central assumption. Here, we investigate how recombination rate evolution affects genetic variation under BGS. We show that recombination rate evolution modifies the effects of BGS in a manner similar to a localised change in the effective population size, potentially leading to an underestimation of the genome-wide effects of selection. Furthermore, we find evidence that recombination rate evolution in the ancestors of modern house mice may have impacted inferences of the genome-wide effects of selection in that species.
Insight into the mechanism of H+-coupled nucleobase transport
Authors: Weng Jun , Zhou Xiaoming , Wiriyasermkul Pattama , Ren Zhenning , Yan Xiuwen , Chen Kehan , Iturbe Eva Gil , Zhou Ming , Quick Matthias
Published: December 2021

Members of the nucleobase/ascorbic acid transporter (NAT) gene family are found in all kingdoms of life. In mammals, the concentrative uptake of ascorbic acid (vitamin C) by members of the NAT family is driven by the Na+ gradient, while the uptake of nucleobases in bacteria is powered by the H+ gradient. Here we report the structure and function PurTCp, a NAT family member from Colwellia psychrerythraea. The structure of PurTCp was determined to 2.80 Å resolution by X-ray crystallography. PurTCp forms a homodimer and each protomer has 14 transmembrane segments folded into a substrate-binding domain (core domain) and an interface domain (gate domain) A purine base is present in the structure and defines the location of the substrate binding site. Functional studies reveal that PurTCp transports purines but not pyrimidines, and that purine binding and transport is dependent on the pH. Mutation of a conserved aspartate residue close to the substrate binding site reveals the critical role of this residue in H+-dependent transport of purines. Comparison of the PurTCp structure with transporters of the same structural fold suggests that rigid-body motions of the substrate-binding domain are central for substrate translocation across the membrane.
Interpersonal brain synchronization during face-to-face economic exchange between acquainted dyads
Authors: Kikuchi Yuto , Tanioka Kensuke , Hiroyasu Tomoyuki , Hiwa Satoru
Published: December 2021

Interpersonal brain synchronization (IBS) has been observed during social interactions and involves various factors, such as familiarity with the partner and type of social activity. A previous study has shown that face-to-face interactions in pairs of strangers increase IBS. However, it is unclear whether this can be observed when the nature of the interacting partners is different. Herein, we aimed to extend these findings to pairs of acquaintances. Neural activity in the frontal and temporal regions was recorded using functional near-infrared spectroscopy hyperscanning. Participants played an ultimatum game that required virtual economic exchange in two experimental settings: the face-to-face and face-blocked conditions. Random pair analysis confirmed whether IBS was induced by social interaction. Contrary to the aforementioned study, our results did not show any cooperative behavior or task-induced IBS increase. Conversely, the random pair analysis results revealed that the pair-specific IBS was significant only in the task condition at the left and right superior frontal, middle frontal, orbital superior frontal, right superior temporal, precentral, and postcentral gyri. Our results revealed that face-to-face interaction in acquainted pairs did not increase IBS and supported the idea that IBS is affected by “with whom we interact and how.”
Fine mapping of goat polledness variant in six Chinese native breeds
Authors: Li Yong , Chen Tao , Yang Man-Man , Han Hu , jiang Dan , Wei Qiang , Zhang Xing-Ju , Ao Yan , Zhang Qingfeng , Miao Ze-Pu , Wang Ran , Li Yuan-Lun , Chao Sheng-Yu , Li Lin , Zhang Ting-Ting , Fang Ming
Published: December 2021

Background

The genetic mechanism of goat polledness has been studied for decades, but identifying causative variants and functional genes remains challenging.

Results

Using a genome-wide association study (GWAS), we identified a significant striking locus for polledness in two different goat breeds. To reduce the linkage disequilibrium among variants for localizing causative variants in the finer region, we sequenced 79 goats from six Chinese native breeds (Jining Gray, Matou, Guizhou black, Yunnan black bone, Chaidamu, and Ujumqin) and identified 483.5 kb CNV (150,334,567-150,818,099) translocated into the previously identified 11.7 kb polled intersex syndrome region, which was consistent with previous research using intersex goat populations. Within the 483.5 kb CNV, a ~322 bp horn-specific element, similar to the superfamily of tRNA-derived families of SINEs, located at the first intron of theERGgene was identified. The results of the GO enrichment analysis showed that the Horn-SINE element-associated genes were involved in both nervous system and head development. Finally, we used RNA sequencing to investigate gene expression profiles in the horn bud and skin tissues of horned and polled goats. We identified 1077 and 1222 differentially expressed genes between the horn bud and skin tissue in polled and horned goats, respectively. We also identified 367 differentially expressed genes in horn buds between polled and horned animals, and found that the two CNV-related genes,ERGandFOXL2, were upregulated in the horn bud of polled goats. Gene functional enrichment analysis demonstrated that the downregulated genes in the horn bud of polled goats were enriched in skeletal system development, whereas the upregulated genes were significantly overexpressed in muscle tissue development.

Conclusions

Broadly, this study describes a novel structural variant responsible for polledness/intersex traits and contributes to the discovery of molecular mechanisms underlying the development and regulation of the polledness trait.
Overactive STAT3 drives accumulation of disease-associated CD21lowB cells
Authors: Masle-Farquhar Etienne , Peters Timothy , Jackson Katherine JL , Singh Mandeep , Ma Cindy S , Suan Daniel , Uzel Gulbu , Chua Ignatius , Leiding Jennifer W , Heiskanen Kaarina , Preece Kahn , Kainulainen Leena , O’Sullivan Michael , Cooper Megan A , Seppänen Mikko RJ , Mustjoki Satu , Brothers Shannon , Vogel Tiphanie P , Brink Robert , Tangye Stuart G , Reed Joanne H , Goodnow Christopher C
Published: December 2021

Dysregulated STAT3 signalling is correlated with antibody-mediated autoimmunity and B- cell neoplasia, but its effect on B cells is underexplored. Here we address this in children with STAT3 gain-of-function (GOF) syndrome and in mice with STAT3T716M, the most common STAT3 GOF syndrome human mutation, or STAT3K658N, a dimerization interface mutation responsible for STAT3 GOF syndrome in two children. The main B cell consequence of overactive STAT3 was accumulation of CD19highCD21lowatypical memory B cells in humans and of CD21lowCD23lowB cells in mice resembling age-associated B cells expressing T-bet, CD11c and plasma cell differentiation genes. Overactive STAT3 within B cells increased expression of many genes in the B cell receptor and T cell help pathways, increased the tolerogenic receptor CD22, but opposed B cell tolerance checkpoints and increased formation of T-bet+ B cells upon BCR and CD40 stimulation. These results reveal overactive STAT3 as a central driver of a key class of disease- associated B-lymphocytes in humans and mice.
High-Resolution Structure of the Nuclease Domain of the Human Parvovirus B19 Main Replication Protein NS1
Authors: Sanchez Jonathan L. , Ghadirian Niloofar , Horton Nancy C.
Published: December 2021

Two new structures of the N-terminal domain of the main replication protein, NS1, of Human Parvovirus B19 (B19V) are presented. This domain (NS1-nuc) plays an important role in the “rolling hairpin” replication of the single-stranded B19V DNA genome, recognizing origin of replication sequences in double-stranded DNA, and cleaving (i.e. nicking) single-stranded DNA at a nearby site known as the trs. One structure of NS1-nuc is solved to 2.4 Å and shows the positions of two bound phosphate ions. A second structure shows the position of a single divalent cation in the DNA nicking active site. The threedimensional structure of NS1-nuc is well conserved between the two forms, as well as with a previously solved structure of a sequence variant of the same domain, however shown here at significantly higher resolution. Using structures of NS1-nuc homologues bound to single- and double-stranded DNA, models for DNA recognition and nicking by B19V NS1-nuc are presented which predict residues important for DNA cleavage and for sequence specific recognition at the viral origin of replication.
Evolution of Enterococcus faecium to A Combination of Daptomycin and Fosfomycin Reveals Distinct and Diverse Adaptive Strategies
Authors: Supandy Adeline , Mehta Heer H. , Tran Truc T. , Miller William R. , Zhang Rutan , Xu Libin , Arias Cesar A. , Shamoo Yousif
Published: December 2021

Infections caused by vancomycin-resistant Enterococcus faecium (VREfm) are an important public health threat. VREfm have become increasingly resistant to the front-line antibiotic, daptomycin (DAP). As such, the use of DAP combination therapies (like fosfomycin [FOS]), has received increased attention. Antibiotic combinations could extend the efficacy of current available antibiotics and potentially delay the onset of further resistance. We investigated the potential for E. faecium HOU503, a clinical VREfm isolate that is DAP and FOS susceptible, to develop resistance to a DAP-FOS combination. Of particular interest was whether the genetic drivers for DAP-FOS resistance might be epistatic and, thus, potentially decrease the efficacy of a combinatorial approach in either inhibiting VREfm or in delaying the onset of resistance. We show that resistance to DAP-FOS could be achieved by independent mutations to proteins responsible for cell wall synthesis for FOS and in altering membrane dynamics for DAP. However, we did not observe genetic drivers that exhibited substantial cross-drug epistasis that could undermine DAP-FOS combination. Of interest was that FOS resistance in HOU503 was largely mediated by changes in phosphoenolpyruvate (PEP) flux as a result of mutations in pyruvate kinase (pyk). Increasing PEP flux could be a readily accessible mechanism for FOS resistance in many pathogens. Importantly, we show that HOU503 were able to develop DAP resistance through a variety of biochemical mechanisms and were able to employ different adaptive strategies. Finally, we showed that the addition of FOS can prolong the efficacy of DAP, significantly extending the timeline to resistance in vitro.

Importance

While the discovery of antibiotics was one of the greatest health care advances in history, its success is being challenged by the emergence of multidrug-resistant pathogens, including vancomycin-resistant enterococci (VRE). Daptomycin (DAP), a lipopeptide antibiotic that targets cell membrane, is currently prescribed as a frontline drug to treat VRE infections. However, emergence of daptomycin-resistant VRE is concerning. Consequently, DAP-Fosfomycin (FOS) combination (DF) has been proposed as a potential method to maintain DAP efficacy. Here, we provide evidence that DF successfully delayed the emergence of resistance in vitro. Genetic data indicates that resistance was acquired independently, with little evidence of significant cross-drug epistasis that could undermine a combinatorial approach. We also uncovered a novel FOS resistance mechanism, through changes in phosphoenolpyruvate (PEP) flux, that may potentially be shared with other bacterial species. Additionally, we also have evidence showing that E. faecium was able to employ different resistance mechanisms.
Helminth infection modulates number and function of adipose tissue Tregs in high fat diet-induced obesity
Authors: Queiroz-Glauss Camila P. , Vieira Mariana S. , Gonçalves-Pereira Marcela Helena , Almeida Stephanie S. , Freire Rachel H. , Gomes Maria Aparecida , Alvarez-Leite Jacqueline Isaura , Santiago Helton C.
Published: December 2021

Background

Epidemiological and experimental studies have shown a protective effect of helminth infections in weight gain and against the development of metabolic dysfunctions in the host. However, the mechanisms induced by the parasite that regulate the development of metabolic diseases in the host are unclear. The present study aimed to verify the influence ofHeligmosomoides polygyrusinfection in early stages of high fat diet-induced obesity.

Principal Findings

The presence of infection was able to prevent exacerbated weight gain in mice fed with high fat diet when compared to non-infected controls. In addition, infected animals displayed improved insulin sensitivity and decreased fat accumulation in the liver. Obesity-associated inflammation was reduced in the presence of infection, demonstrated by higher levels of IL10 and adiponectin, increased infiltration of Th2 and eosinophils in adipose tissue of infected animals. Of note, the parasite infection was associated with increased Treg frequency in adipose tissue which showed higher expression of cell surface markers of function and activation, like LAP and CD134. The infection could also revert the loss of function in Tregs associated with high fat diet.

Conclusion

These data suggest thatH. polygyrusinfection can prevent weight gain and metabolic syndrome in animals fed with high fat diet associated with modulations of adipose tissue Treg cells.

Author summary

Helminth infections are known to modulate the immune system being responsible for protecting the host from developing allergic and autoimmune disorders (Hygiene Hypothesis). We hypothesized that the same immunomodulatory effect can have an impact on immunometabolic diseases, such as obesity and its linked diseases such as type 2 diabetes. Weight disorders have reached epidemic levels, nearly tripling since 1975 and being responsible for almost 5 million premature deaths each year. To test our hypothesis C57BL/6 male mice were fed control or high fat diet, for five weeks, in the presence or not ofHeligmosomoides polygyrusinfection. Weight gain, development of metabolic disorders, inflammation and cellular migration to the adipose tissue were evaluated. In accordance with our hypothesis, we found that the presence of infection prevented the exacerbated weight gain and also improved metabolic parameters in animals fed a high fat diet. This was associated with the infection’s ability to modulate parameters of a cell responsible for regulatory functions: Tregs. In the light of these findings, helminth infection could be protective against weight gain and metabolic disturbances.
Diminished neutralization responses towards SARS-CoV-2 Omicron VoC after mRNA or vector-based COVID-19 vaccinations
Authors: Jacobsen Henning , Strengert Monika , Maaß Henrike , Ynga Durand Mario Alberto , Kessel Barbora , Harries Manuela , Rand Ulfert , Abassi Leila , Kim Yeonsu , Lüddecke Tatjana , Hernandez Pilar , Ortmann Julia , Heise Jana-Kristin , Castell Stefanie , Gornyk Daniela , Glöckner Stephan , Melhorn Vanessa , Kemmling Yvonne , Lange Berit , Dulovic Alex , Häring Julia , Junker Daniel , Schneiderhan-Marra Nicole , Hoffmann Markus , Pöhlmann Stefan , Krause Gérard , Cicin-Sain Luka
Published: December 2021

SARS-CoV-2 variants accumulating immune escape mutations provide a significant risk to vaccine-induced protection. The novel variant of concern (VoC) Omicron (B.1.1.529) has the largest number of amino acid alterations in its Spike protein to date. Thus, it may efficiently escape recognition by neutralizing antibodies, allowing breakthrough infections in convalescent and vaccinated individuals. We analyzed neutralization activity of sera from individuals after vaccination with all mRNA-, vector- or heterologous immunization schemes currently available in Europe by in vitro neutralization assay at peak response towards SARS-CoV-2 B.1, Omicron, Beta and Delta pseudotypes and also provide longitudinal follow-up data from BNT162b2 vaccinees. All vaccines apart from Ad26.CoV2.S showed high levels of responder rates (93-100%) towards SARS-CoV-2 wild-type, but some reductions in neutralizing Beta and Delta VoC pseudotypes. The novel Omicron variant had the biggest impact, both in terms of response rates and neutralization titers. Only mRNA-1273 showed a 100% response rate to Omicron and induced the highest level of neutralizing antibody titers, followed by heterologous prime-boost approaches. Homologous BNT162b2 vaccination or vector-based AZD1222 or Ad26.CoV2.S performed less well with peak responder rates of 33%, 50% and 9%, respectively. However, Omicron responder rates in BNT162b2 recipients were maintained in our six month longitudinal follow-up indicating that individuals with cross-protection against Omicron maintain it over time. Overall, our data strongly argues for urgent booster doses in individuals who were previously vaccinated with BNT162b2, or a vector-based immunization scheme.
Immunity acquired by a minority active fraction of the population could explain COVID-19 spread in Greater Buenos Aires (June-November 2020)
Authors: Fabricius Gabriel , Borzi Rodolfo A. , Caminos José , Grigera Tomás S.
Published: December 2021

The COVID-19 pandemic had an uneven development in different countries. In Argentina, the pandemic began in march 2020 and, during the first 3 months, the vast majority of cases were concentrated in a densely populated region that includes the city of Buenos Aires (country capital) and the Greater Buenos Aires area that surrounds it. This work focuses on the spread of COVID-19 between June and November 2020 in Greater Buenos Aires. Within this period of time there was no vaccine, basically only the early wild strain of SARS-CoV-2 was present, and the official restriction and distancing measures in this region remained more or less constant. Under these particular conditions, the incidences show a sharp rise from June 2020 and begin to decrease towards the end of August until the end of November 2020. In this work we study, through mathematical modelling and available epidemiological information, the spread of COVID-19 in this region and period of time. We show that a coherent explanation of the evolution of incidences can be obtained assuming that only a minority fraction of the population got involved in the spread process, so that the incidences decreased as this group of people was becoming immune. The observed evolution of the incidences could then be a consequence at the population level of lasting immunity conferred by SARS-CoV-2.
GB0139, an inhaled small molecule inhibitor of galectin-3, in COVID-19 pneumonitis: a randomised, controlled, open-label, phase 2a experimental medicine trial of safety, pharmacokinetics, and potential therapeutic value
Authors: Gaughan Erin , Sethi Tariq , Quinn Tom , Hirani Nikhil , Mills Andrew , Bruce Annya M. , MacKinnon Alison , Aslanis Vassilios , Li Feng , O’Connor Richard , Parker Richard A. , Norrie John , Dear James , Akram Ahsan R. , Koch Oliver , Wang-Jairaj Jie , Slack Robert J. , Gravelle Lise , Lindmark Bertil , Dhaliwal Kevin
Published: December 2021

Rationale

High galectin-3 levels predict poor outcomes in patients with COVID-19. Galectin-3 activates monocytes and macrophages which are directly implicated in COVID-19 immunopathology and the cytokine storm. GB0139 is a potent thiodigalactoside galectin-3 inhibitor and may reduce the severe effects of the disease. We report safety and pharmacokinetics and pharmacodynamics of the inhaled galectin-3 inhibitor, GB0139, and assess clinical outcomes and key systemic inflammatory biomarkers in hospitalised patients with COVID-19 (ClinicalTrials.gov/EudraCT identifier: NCT04473053/2020-002230-32).

Methods

Adults with COVID-19 requiring oxygen, and with pneumonitis on x-ray, were randomised to receive standard of care (SOC; including dexamethasone; n=21) or SOC plus 10 mg GB0139 twice daily for 48 hours, then once daily for ≤14 days (n=20).

Results

Patients aged 27–87 years were enrolled from July 2020; the final patient completed the 90-day follow-up in April 2021. GB0139+SOC was well tolerated with no treatment-related serious adverse events reported. Incidences of adverse events were similar between treatment arms (40 with GB0139+SOC vs 35 with SOC). Plasma GB0139 was measurable in all patients after inhaled exposure, with moderate interpatient variability, and demonstrated target engagement with decreased circulating galectin (overall treatment effect post-hoc over days 2–7: p=0·0099 vs SOC). Rate of decline in fraction of inspired oxygen (%) requirement was significantly greater in the GB0139+SOC arm with a posterior mean difference of -1·51 (95% highest posterior density: -2·90, -0·189) versus SOC. Plasma levels of biomarkers associated with inflammation, coagulopathy, major organ function and fibrosis showed a downward trend versus SOC.

Conclusions

GB0139+SOC was well tolerated and achieved clinically relevant plasma concentrations and target engagement. This, and the reduction in markers associated with inflammatory, coagulation, fibrosis, and reduction in inspired oxygen (%) over SOC alone, indicates the therapeutic potential for inhaled GB0139 in hospitalised patients with COVID-19.
The impact of gastric acid suppression on the developing intestinal microbiome of a child
Authors: Andrew Spurr , Wei Zhu , Wong Wendy S. W. , Bernadette Diez , Ian Leibowitz , Otto Louis-Jacques , Thierry Vilboux , John Niederhuber , Levy Shira L , Hourigan Suchitra K
Published: December 2021

Gastric acid suppressing medications have been associated with an increased risk of Clostridioides difficile (C. difficile) infection, hypothesized to be due to underlying intestinal microbiome changes. Our goal is to characterize these changes in children as their microbiome is undergoing critical development. Our study included 5 children (< 3 years old) who were started on clinically indicated gastric acid suppression and 15 healthy age-matched controls. Stool samples were collected before and after 2 months of treatment. We analyzed the microbiome using 16S rRNA sequencing. Quantitative-PCR was used to detect C. difficile toxins. Subjects and controls had similar alpha and beta diversity. We found no significant change in alpha or beta diversity of subjects after treatment. C. difficile toxins were not found and there was no increase in C. difficile carriage after treatment. A significant increase in Lactobacillus was found after treatment, which has been associated with C. difficile in adults.
An automated heart rate-based algorithm for sleep stage classification: validation using conventional PSG and innovative wearable ECG device
Authors: Pini Nicolò , Ong Ju Lynn , Yilmaz Gizem , Chee Nicholas I. Y. N. , Siting Zhao , Awasthi Animesh , Biju Siddharth , Kishan Kishan , Patanaik Amiya , Fifer William P. , Lucchini Maristella
Published: December 2021

Study Objectives

Validate a HR-based deep-learning algorithm for sleep staging named Neurobit-HRV (Neurobit Inc., New York, USA).

Methods

The algorithm can perform classification at 2-levels (Wake; Sleep), 3-levels (Wake; NREM; REM) or 4-levels (Wake; Light; Deep; REM) in 30-second epochs. The algorithm was validated using an open-source dataset of PSG recordings (Physionet CinC dataset, n=994 participants) and a proprietary dataset (Z3Pulse, n=52 participants), composed of HR recordings collected with a chest-worn, wireless sensor. A simultaneous PSG was collected using SOMNOtouch. We evaluated the performance of the models in both datasets using Accuracy (A), Cohen’s kappa (K), Sensitivity (SE), Specificity (SP).

Results

CinC - The highest value of accuracy was achieved by the 2-levels model (0.8797), while the 3-levels model obtained the best value of K (0.6025). The 4-levels model obtained the lowest SE (0.3812) and the highest SP (0.9744) for the classification of Deep sleep segments. AHI and biological sex did not affect sleep scoring, while a significant decrease of performance by age was reported across the models. Z3Pulse - The highest value of accuracy was achieved by the 2-levels model (0.8812), whereas the 3-levels model obtained the best value of K (0.611). For classification of the sleep states, the lowest SE (0.6163) and the highest SP (0.9606) were obtained for the classification of Deep sleep segment.

Conclusions

Results demonstrate the feasibility of accurate HR-based sleep staging. The combination of the illustrated sleep staging algorithm with an inexpensive HR device, provides a cost-effective and non-invasive solution easily deployable in the home.