bluuebunny/crossref_metadata_2025 huggingface dataset:
An updated review of the marine ornamental fish trade in the European Union
Authors: Biondo Monica V. , Burki Rainer P. , Aguayo Francisco , Calado Ricardo
Published: March 2024

Wild-caught fishes from coral reefs, one of the most threatened ecosystems on the planet, continue to supply the marine aquarium trade. Despite customs and veterinary checks during imports, comprehensive data on this global industry remains scarce. This study provides an updated review on one of its largest import markets, the European Union (EU): 24-million-euro annual trade value, detailed exporting and importing countries also diversity of species and families of the 26 million specimens imported between 2014 and 2021. We then show how a watchlist alert system based on the number of specimens traded, import trends, and vulnerability index according to FishBase and IUCN Red List conservation status can provide key information on which species should require a closer scrutiny. The stark decline in trade of the most traded species,Chromis viridis,may warrant monitoring through the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). While the European TRAde Control and Expert System (TRACES) electronically monitors the movement of live animals to respond quickly to biosecurity risks, one-third of marine ornamental fishes imported lack species-level information. With minor adjustments, TRACES could enhance the monitoring of wildlife trade, with marine ornamental fish being an interesting case study.
Expanded methylome and quantitative trait loci detection by long-read profiling of personal DNA
Authors: Groza Cristian , Ge Bing , Cheung Warren , Pastinen Tomi , Bourque Guillaume
Published: March 2024

Structural variants (SVs) are omnipresent in human DNA, yet their genotype and methylation status is rarely characterized due to previous limitations in genome assembly and detection of modified nucleotides. Because of this, the extent to which these regions act as quantitative-trait loci is also largely unknown.

Here, we generated a pangenome graph summarizing the SVs in 782 de novo assembled genomes obtained from the Genomic Answers for Kids rare disease cohort, that captures 14.6 million CpGs in DNA segments that are absent from the CHM13v2 assembly (SV-CpGs), expanding their number by 43.6%. Next, using 435 methylomes from the same samples, we genotyped a total of 7.99 million SV-CpGs, of which 5.18 million (64.8%) were found to be methylated (SV-5mCpGs) in at least one sample.

To understand the provenance and impact of these novel SV-CpGs, we noted that non-repeat sequences were the leading contributor of SV-CpGs (3.3 × 106), followed by centromeric satellites (1.58 × 106), simple repeats (1.19 × 106), Alus (0.67 × 106), satellites (0.39 × 106), L1s (0.27 × 106), and SVAs (0.19 × 106). Meanwhile, the methylation rate of SV-CpGs was the highest in repeat sequences. Moreover, in contrast to Alus and L1s, centromeric satellites, simple repeats and SVA sequences were overrepresented in SV-5mCpGs compared to reference CpGs. Similarly, we established that non-reference CpGs were more than twice (37% vs. 15%) as likely to be variable, showing intermediate methylation levels in the population.

Lastly, to explore if SVs detected in this pangenome are potentially causal for functional variation in population we measured methylation quantitative trait loci (SV-mQTLs) using CHM13v2 as a backbone. This revealed over 230,464 methylation bins within 100 kbp of a common SV (>5% MAF) showing significant association (at 5% FDR) with methylation variation. Finally, we assessed how many of these SVs-mQTLs were the leading QTL variant compared to SNVs and identified 65,659 methylation bins (28.5%) where the leading variant was an SV.

In conclusion, our results demonstrate that graph genome references providing full SV structures in combination with the associated methylation variation reveal tens-of-thousands of QTLs that are more accurately mapped in personal genomes, underscoring the importance of assembly-based analyses of human traits.
DNA metabarcoding revealed how time and space do matter -sex does not- in the dietary variation of the endangered Dupont’s Lark
Authors: Zurdo Julia , Bustillo-de la Rosa Daniel , Barrero Adrián , Gómez-Catasús Julia , Reverter Margarita , Pérez-Granados Cristian , García Jesús T. , Viñuela Javier , Domínguez Julio C. , Morales Manuel B. , Traba Juan
Published: March 2024

A species’ diet is highly dependent on the availability of food resources in space and time, as well as on intrinsic factors such as sex or age. Accurate assessments of variations in the diet composition of bird populations across spatial scales, seasons and demographic groups are essential not only for understanding the basic ecology of species, but also for the conservation of endangered ones. However, our current knowledge about how birds’ diet change according to spatio-temporal variations or intrinsic factors is very limited. Here, we used a multi-marker metabarcoding approach to characterize the diet of a declining shrub-steppe passerine, the Dupont’s Lark (Chersophilus duponti), throughout a large part of its global distribution range. We also investigated spatial, phenological and sexual variations in its diet. Using markers from two genomic regions (18S and COI), we analyzed fecal samples from 303 adult Dupont’s larks from Morocco and Spain during the breeding and non-breeding seasons. Overall, arthropods from the orders Coleoptera, Lepidoptera, Julida and Orthoptera were the main prey consumed by Dupont’s Larks. We found that Dupont’s Lark diet varied spatially, as well as temporally, reflecting dietary plasticity in response to changes in prey availability across landscapes and the species’ phenological periods. High dietary overlap and no differences between sexes were observed, suggesting similar foraging behavior and nutritional requirements in both sexes. This is the first study providing detailed information on Dupont’s Lark food ecology over much of its distribution, which is fundamental for the management and conservation of this declining steppe species.
Endocrine, immune and disease dynamics in a patient with rheumatoid arthritis during flare and medication change
Authors: Seizer Lennart , Gostner Johanna , Garbers Christoph , Licht Melina , Sager Sebastian , Schubert Christian
Published: March 2024

Objective

Rheumatoid arthritis (RA) is a chronic autoimmune disease of widely unknown etiology and pathophysiology. In this integrative single-case study on a patient with RA, we had the unique opportunity to closely monitor the individual dynamics of endocrine, immune and disease variables during a naturally occurring flare-up and subsequent medication change.

Methods

The female RA patient collected her entire urine over 30 days in 12-h intervals (60 consecutive measurements in total). Subsequently, interleukin-6 (IL-6), orosomucoid-2, cortisol (ELISA), neopterin and creatinine (HPLC) levels were determined in the urine samples. Further, each morning and evening, the patient completed the DIARI, a set of questionnaires on variables such as subjective pain, subjective RA disease activity and emotional states. Once a week, besides an online video interview, the patient had an appointment at her rheumatologist, in which several indices of RA disease activity were determined: SDAI, CDAI and DAS28. From these data various time series were constructed for statistical analysis.

Results

RA disease state increased from low to high activity during the first 12 study days. Thereupon, the medication was changed, which proved effective in reducing RA disease activity. However, the levels of urinary neopterin, urinary orosomucoid-2, and urinary IL-6 did not show any response, neither to the increasing disease activity nor the medication change. The patient’s daily reports on pain, RA disease activity, and emotional states, however, mirrored the course of the rheumatologic indices.

Conclusion

This integrative single-case study clearly demonstrated the importance of process analysis for the evaluation of therapeutic measures in RA. In the patient studied, urinary levels of neopterin, orosomucoid-2 and urinary IL-6 were not found to be appropriate biomarkers of short-term fluctuations in RA disease activity. Instead, the results reported by the patient proved to be a useful tool for ambulatory and longitudinal monitoring of RA.
CORACLE (COVID-19 liteRAture CompiLEr): A platform for efficient tracking and extraction of SARS-CoV-2 and COVID-19 literature, with examples from post-COVID with respiratory involvement
Authors: Piontkovskaya Kristina , Luo Yulian , Lindberg Pia , Gao Jing , Runold Michael , Kolosenko Iryna , Li Chuan-Xing , Wheelock Åsa M.
Published: March 2024

Background

During the COVID-19 pandemic there emerged a need to efficiently monitor and process large volumes of scientific literature on the subject. Currently, as the pandemic is winding down, the clinicians encountered a novel syndrome - Post-acute Sequelae of COVID- 19 (PASC) - that affects over 10% of those who contract SARS-CoV-2 and presents a significant and growing challenge in the medical field. The continuous influx of new research publications underscores a critical need for efficient tools for navigating the literature.

Objectives

We aimed to develop an application which will allow monitoring and categorizing COVID-19-related literature through building publication networks and medical subject headings (MeSH) maps to be able to quickly identify key publications and publication networks.

Methods

We introduce CORACLE (COVID-19 liteRAture CompiLEr), an innovative web application designed for the analysis of COVID-19-related scientific articles and the identification of research trends. CORACLE features three primary interfaces: The “Search” interface, which displays research trends and citation links; the “Citation Map” interface, allowing users to create tailored citation networks from PubMed Identifiers (PMIDs) to uncover common references among selected articles; and the “MeSH” interface, highlighting current MeSH trends and associations between MeSH terms.

Results

Our web application, CORACLE, leverages regularly updated PubMed data to aggregate and categorize the extensive literature on COVID-19 and PASC, aiding in the identification of relevant research publication hubs. Using lung function in PASC patients as a search example, we demonstrate how to identify and visualize the interactions between the relevant publications.

Conclusion

CORACLE proves to be an effective tool for the extraction and analysis of literature. Its functionalities, including the MeSH trends and customizable citation mapping, facilitate the discovery of relevant information and emerging trends in COVID-19 and PASC research.
Gene editing ofCD3 epsilongene to redirect regulatory T cells for adoptive T cell transfer
Authors: Du Weijie , Noyan Fatih , McCallion Oliver , Drosdek Vanessa , Kath Jonas , Glaser Viktor , Fuster-Garcia Carla , Yang Mingxing , Stein Maik , Weber Olaf , Polansky Julia K. , Cathomen Toni , Jaeckel Elmar , Hester Joanna , Issa Fadi , Volk Hans-Dieter , Schmueck-Henneresse Michael , Reinke Petra , Wagner Dimitrios L.
Published: March 2024

I.

Adoptive transfer of regulatory T cells (Tregs) is a promising strategy to combat immunopathologies in transplantation and autoimmune diseases. Antigen-specific Tregs are more effective in modulating undesired immune reactions, but their low frequency in peripheral blood poses challenges for manufacturing and their clinical application. Chimeric antigen receptors (CARs) have been used to redirect the specificity of Tregs, employing retroviral vectors. However, retroviral gene transfer is costly, time consuming, and raises safety issues. Here, we explored non-viral gene editing to redirect Tregs with CARs, using HLA-A2-specific constructs for proof-of-concept studies in transplantation models. We introduce a virus-free CRISPR-Cas12a approach to integrate an antigen-binding domain into theCD3 epsilon(CD3ε) gene, generating Tregs expressing a T cell receptor fusion construct (TruC). TheseCD3ε-TruC Tregs exhibit potent antigen-dependent activation while maintaining responsiveness to TCR/CD3 stimulation. This enables preferential enrichment of TruC-redirected Tregs via repetitive CD3/CD28-stimulation in a GMP-compatible expansion system. Non-viral gene editedCD3ε-TruC Tregs retained their phenotypic, epigenetic, and functional identity. In a humanized mouse model, HLA-A2-specificCD3ε-TruC Tregs demonstrate superior protection of allogeneic HLA-A2+skin grafts from rejection compared to polyclonal Tregs. This approach provides a pathway for developing clinical-gradeCD3ε-TruC-based Treg cell products for transplantation immunotherapy and other immunopathologies.
VLK drives extracellular phosphorylation of EphB2 to govern the EphB2-NMDAR interaction and injury-induced pain
Authors: Srikanth Kolluru D. , Elahi Hajira , Chander Praveen , Washburn Halley R. , Hassler Shayne , Mwirigi Juliet M. , Kume Moeno , Loucks Jessica , Arjarapu Rohita , Hodge Rachel , Shiers Stephanie I. , Sankaranarayanan Ishwarya , Erdjument-Bromage Hediye , Neubert Thomas A. , Campbell Zachary T. , Paik Raehum , Price Theodore J. , Dalva Matthew B.
Published: March 2024

Phosphorylation of hundreds of protein extracellular domains is mediated by two kinase families, yet the significance of these kinases is underexplored. Here, we find that the presynaptic release of the tyrosine directed-ectokinase, Vertebrate Lonesome Kinase (VLK/Pkdcc), is necessary and sufficient for the direct extracellular interaction between EphB2 and GluN1 at synapses, for phosphorylation of the ectodomain of EphB2, and for injury-induced pain.Pkdccis an essential gene in the nervous system, and VLK is found in synaptic vesicles, and is released from neurons in a SNARE-dependent fashion. VLK is expressed by nociceptive sensory neurons where presynaptic sensory neuron-specific knockout renders mice impervious to post-surgical pain, without changing proprioception. VLK defines an extracellular mechanism that regulates protein-protein interaction and non-opioid-dependent pain in response to injury.

One-Sentence Summary

Synaptic protein-protein interactions and pain are regulated by the presynaptic release of the extracellular kinase VLK in the spinal cord.
Granzyme B-based CAR T cells block metastasis by eliminating circulating tumor cells
Authors: Sun Bin , Guo Jing , Yang Dong , Hu Qiancheng , Ma Haiyan , Tian Panwen , Liu Nan , Lv Longbao , Yan Lanzhen , Ding Hao , Fu Maoyong , Gou Hongfeng , Cao Dan , Liu Dan , Chen Nianyong , Shi Peng , Li Weimin , Zhao Xudong
Published: March 2024

Chimeric antigen receptor (CAR) T cells have limited efficacy against solid tumors due to the hostile microenvironment. Circulating tumor cells (CTCs) are essential to metastasis, which is the cause of most of cancer-related death. Here, we generated GrB-CAR T cells targeting membrane-bound HSP70 (mHSP70), a highly tumor-specific antigen detected in numerous cancers. GrB-CAR T cells exhibited potent cytotoxicity against a broad spectrum of cancer cell lines and stem-like cancer cellsin vitroand effectively inhibited xenograft tumor growthin vivo. Importantly, GrB-CAR T cells markedly decreased the number of CTCs and, therefore, hindered cancer metastasis in spontaneous metastasis models with uncontrollable primary tumor growth, a scenario commonly encountered in clinical trials of CAR T therapies for solid tumors. Furthermore, despite the 100% homology between human and macaque HSP70 protein, the autotransplantation of macaque T cells expressing human GrB-CAR did not cause any obvious toxic effects. These results not only demonstrate GrB-CAR T cells as a safe and effective tactic with broad-spectrum anticancer activity, but also offer strong experimental evidence and proof-of-concept validation for CAR T cell-mediated metastasis inhibition by targeting CTCs.
Myc upregulates Ggct, γ-glutamylcyclotransferase to promote development ofp53-deficient osteosarcoma
Authors: Ueno Tomoya , Otani Shohei , Date Yuki , Katsuma Yu , Nagayoshi Yuma , Ito Tomoko , Ii Hiromi , Kageyama Susumu , Nakata Susumu , Ito Kosei
Published: March 2024

Osteosarcoma (OS) in humans is characterized by alterations in theTP53gene. In mice, loss of p53 triggers OS development, for which c-Myc (Myc) oncogenicity is indispensable. However, little is known about which genes are targeted by Myc to promote tumorigenesis. Here, we examined the role of Ggct, γ-glutamylcyclotransferase which is a component enzyme of γ-glutamyl cycle essential for glutathione homeostasis, in human and mouse OS development. We found thatGGCTis a poor prognostic factor for human OS, and that deletion ofGgctsuppressesp53-deficient osteosarcomagenesis in mice. Myc upregulates Ggct directly by binding to theGgctpromoter, and deletion of a Myc binding site therein by genome editing attenuated the tumorigenic potential ofp53- deficient OS cells. Taken together, these results show a rationale that GGCT is widely upregulated in cancer cells and solidify its suitability as a target for anticancer drugs.
Altered co-stimulatory and inhibitory receptors on monocyte subsets in patients with visceral leishmaniasis
Authors: Adem E. , Yizengaw E. , Mulaw T. , Nibret E. , Müller I. , Takele Y. , Kropf P.
Published: March 2024

Visceral leishmaniasis (VL) is a neglected tropical disease caused by parasites from theLeishmania(L.)donovanicomplex. VL is characterised by uncontrolled parasite replication in spleen, liver and bone marrow, and by an impaired immune response and high systemic levels of inflammation. Monocytes have been poorly characterised in VL patients. The aim of this study was to evaluate the expression levels of markers involved in the regulation of T cell responses on different subsets of monocytes from the blood of VL patients and healthy non-endemic controls (HNEC). Monocytes can broadly be divided into three subsets: classical, intermediate and non-classical monocytes. Our results show that the percentages of all three subsets stay similar at the time of VL diagnosis (ToD) and at the end of anti-leishmanial treatment (EoT). We first looked at co-stimulatory receptors: the expression levels of CD40 were significantly increased on classical and intermediate, but not non-classical monocytes, at ToD as compared to EoT and HNEC. CD80 expression levels were also increased on intermediate monocytes at ToD as compared to EoT and HNEC, and on classical monocytes only as compared to HNEC. The levels of CD86 were similar at EoT and ToD and in HNEC on classical and intermediate monocytes, but significantly higher at EoT on non-classical monocytes. We also looked at an inhibitory molecule, PD-L1. Our results show that the expression levels of PD-L1 is significantly higher on all three monocyte subsets at ToD as compared to HNEC, and to EoT on classical and intermediate monocytes.

These results show that monocytes from the blood of VL patients upregulate both co-stimulatory and inhibitory receptors and that their expression levels are restored at EoT.
In vivooptogenetics using a Utah Optrode Array with enhanced light output and spatial selectivity
Authors: McAlinden Niall , Reiche Christopher F. , Clark Andrew M. , Scharf Robert , Cheng Yunzhou , Sharma Rohit , Rieth Loren , Dawson Martin D. , Angelucci Alessandra , Mathieson Keith , Blair Steve
Published: March 2024

Optogenetics allows manipulation of neural circuitsin vivowith high spatial and temporal precision. However, combining this precision with control over a significant portion of the brain is technologically challenging (especially in larger animal models). Here, we have developed, optimised, and tested in vivo, the Utah Optrode Array (UOA), an electrically addressable array of optical needles and interstitial sites illuminated by 181 µLEDs and used to optogenetically stimulate the brain. The device is specifically designed for non-human primate studies. Thinning the combined µLED and needle backplane of the device from 300 µm to 230 µm improved the efficiency of light delivery to tissue by 80%, allowing lower µLED drive currents, which improved power management and thermal performance. The spatial selectivity of each site was also improved by integrating an optical interposer to reduce stray light emission. These improvements were achieved using an innovative fabrication method to create an anodically bonded glass/silicon substrate with through-silicon vias etched, forming an optical interposer. Optical modelling was used to demonstrate that the tip structure of the device had a major influence on the illumination pattern. The thermal performance was evaluated through a combination of modelling and experiment, in order to ensure that cortical tissue temperatures did not rise by more than 1°C. The device was testedin vivoin the visual cortex of macaque expressing ChR2-tdTomato in cortical neurons. It was shown that the strongest optogenetic response occurred in the region surrounding the needle tips, and that the extent of the optogenetic response matched the predicted illumination profile based on optical modelling – demonstrating the improved spatial selectivity resulting from the optical interposer approach. Furthermore, different needle illumination sites generated different patterns of low-frequency potential (LFP) activity.
Post-transcriptional RNA stabilization of telomere-proximal RNAs FRG2, DBET, D4Z4 at human 4q35 in response to genotoxic stress and D4Z4 macrosatellite repeat length
Authors: Salsi Valentina , Losi Francesca , Salani Monica , Kaufman Paul D. , Tupler Rossella
Published: March 2024

Background

Reduced copy number of the D4Z4 macrosatellite at human chromosome 4q35 is associated with facioscapulohumeral muscular dystrophy (FSHD). A pervasive idea is that chromatin alterations at the 4q35 locus following D4Z4 repeat unit deletion lead to disease via inappropriate expression of nearby genes. Here, we sought to analyze transcription and chromatin characteristics across 4q35 and how these are affected by D4Z4 deletions and exogenous stresses.

Results

We found that the 4q subtelomere is subdivided into discrete domains, each with characteristic chromatin features associated with distinct gene expression profiles. Centromere-proximal genes within 4q35 (ANT1,FAT1andFRG1)display active histone marks at their promoters. In contrast, poised or repressed markings are present at telomere-proximal loci includingFRG2, DBE-TandD4Z4. We discovered that these discrete domains undergo region-specific chromatin changes upon treatment with chromatin enzyme inhibitors or genotoxic drugs. We demonstrated that the 4q35 telomere-proximalFRG2, DBE-TandD4Z4-derived transcripts are induced upon DNA damage to levels inversely correlated with the D4Z4 repeat number, are stabilized through post-transcriptional mechanisms upon DNA damage, and are bound to chromatin.

Conclusion

Our study reveals unforeseen biochemical features of RNAs from clustered transcription units within the 4q35 subtelomere. Specifically, theFRG2, DBE-TandD4Z4-derived transcripts are chromatin-associated and are stabilized post-transcriptionally after induction by genotoxic stress. Remarkably, the extent of this response is modulated by the copy number of the D4Z4 repeats, raising new hypotheses about their regulation and function in human biology and disease.
Fibroblast diversification is an embryonic process dependent on muscle contraction
Authors: Coren Lavi , Zaffryar-Eilot Shelly , Odeh Anas , Kaganovsky Anna , Hasson Peleg
Published: March 2024

Fibroblasts are the most common cell type found in connective tissues, known to play major roles in development, homeostasis, regeneration and disease. Although specific fibroblast subpopulations were associated with different biological processes, the mechanisms and unique activities underlying their diversity has not been thoroughly examined. Turning to skeletal muscle development, we set to dissect the variation of muscle-resident fibroblasts (mrFibroblasts). Our results demonstrate mrFibroblasts diversify following the transition from embryonic to fetal myogenesis prior to birth. We find mrFibroblast segregate into two major subpopulations occupying distinct niches, with interstitial fibroblasts residing between the muscle fibers, and delineating fibroblasts sheathing the muscle mass. We further show these subpopulations entail distinct cellular dynamics and transcriptomes. Notably, we find mrFibroblast subpopulations exert distinct regulatory roles on myoblast proliferation and differentiation. Finally, we demonstrate this diversification depends on muscle contraction. Altogether, these findings establish mrFibroblast diversify in a spatio-temporal embryonic process into distinct cell types, entailing different characteristics and roles.
CACNA1Ahaploinsufficiency leads to reduced synaptic function and increased intrinsic excitability
Authors: Hommersom Marina P. , Doorn Nina , Puvogel Sofía , Lewerissa Elly I. , Mordelt Annika , Ciptasari Ummi , Kampshoff Franziska , Dillen Lieke , van Beusekom Ellen , Oudakker Astrid , Kogo Naoki , Frega Monica , Schubert Dirk , van de Warrenburg Bart P.C. , Nadif Kasri Nael , van Bokhoven Hans
Published: March 2024

Haploinsufficiency of theCACNA1Agene, encoding the pore-forming α1 subunit of P/Q-type voltage-gated calcium channels, is associated with a clinically variable phenotype ranging from cerebellar ataxia, to neurodevelopmental syndromes with epilepsy and intellectual disability.

To understand the pathological mechanisms ofCACNA1Aloss-of-function variants, we characterized a human neuronal model forCACNA1Ahaploinsufficiency, by differentiating isogenic induced pluripotent stem cell lines into glutamatergic neurons, and investigated the effect ofCACNA1Ahaploinsufficiency on mature neuronal networks through a combination of electrophysiology, gene expression analysis, andin silicomodeling.

We observed an altered network synchronization inCACNA1A+/−networks alongside synaptic deficits, notably marked by an augmented contribution of GluA2-lacking AMPA receptors. Intriguingly, these synaptic perturbations coexisted with increased non-synaptically driven activity, as characterized by inhibition of NMDA and AMPA receptors on micro-electrode arrays. Single-cell electrophysiology and gene expression analysis corroborated this increased intrinsic excitability through reduced potassium channel function and expression. Moreover, we observed partial mitigation of theCACNA1A+/−network phenotype by 4-aminopyridine, a therapeutic intervention for episodic ataxia type 2.

In summary, our study pioneers the characterization of a human induced pluripotent stem cell-derived neuronal model forCACNA1Ahaploinsufficiency, and has unveiled novel mechanistic insights. Beyond showcasing synaptic deficits, this neuronal model exhibited increased intrinsic excitability mediated by diminished potassium channel function, underscoring its potential as a therapeutic discovery platform with predictive validity.
NFIB influences progenitor competence in maturation of GABAergic neurons in mice
Authors: Bright Ann Rose , Kotlyarenko Yana , Neuhaus Florian , Rodrigues Diana , Feng Chao , Peters Christian , Vitali Ilaria , Dönmez Elif , Myoga Michael H. , Dvoretskova Elena , Mayer Christian
Published: March 2024

Diverse types of GABAergic projection neurons and interneurons of the telencephalon derive from progenitors in a ventral germinal zone, called the ganglionic eminence. Using single-cell transcriptomics, chromatin accessibility profiling, lineage tracing, birthdating, heterochronic transplantation, and perturbation sequencing in mouse embryos, we investigated how progenitor competence influences the maturation and differentiation of these neurons. We found that the progression of neurogenesis over developmental time shapes maturation competence in ganglionic eminence progenitors, influencing how they progress into mature states. In contrast, differentiation competence, which defines the ability to produce diverse transcriptomic identities, remains largely unaffected by the stages of neurogenesis. Chromatin remodeling alongside a NFIB-driven regulatory gene module influences maturation competence in late-born neurons. These findings provide key insights into how transcriptional programs and chromatin accessibility govern neuronal maturation and the diversification of GABAergic neuron subtypes during neurodevelopment.
A Developmental Mechanism to Regulate Alternative Polyadenylation in an Adult Stem Cell Lineage
Authors: Gallicchio Lorenzo , Matias Neuza R. , Morales-Polanco Fabian , Nava Iliana , Stern Sarah , Zeng Yi , Fuller Margaret T.
Published: March 2024

Alternative Cleavage and Polyadenylation (APA) often results in production of mRNA isoforms with either longer or shorter 3’UTRs from the same genetic locus, potentially impacting mRNA translation, localization and stability. Developmentally regulated APA can thus make major contributions to cell-type-specific gene expression programs as cells differentiate. DuringDrosophilaspermatogenesis, approximately 500 genes undergo APA when proliferating spermatogonia differentiate into spermatocytes, producing transcripts with shortened 3’ UTRs, leading to profound stage-specific changes in the proteins expressed. The molecular mechanisms that specify usage of upstream polyadenylation sites in spermatocytes are thus key to understanding the changes in cell state. Here, we show that upregulation of PCF11 and Cbc, the two components of Cleavage Factor II (CFII), orchestrates APA duringDrosophilaspermatogenesis. Knock down ofPCF11orcbcin spermatocytes caused dysregulation of APA, with many transcripts normally cleaved at a proximal site in spermatocytes now cleaved at their distal site, as in spermatogonia. Forced overexpression of CFII components in spermatogonia switched cleavage of some transcripts to the proximal site normally used in spermatocytes. Our findings reveal a developmental mechanism where changes in expression of specific cleavage factors can direct cell-type-specific APA at selected genes.
Natural history of eukaryotic DNA viruses with double jelly-roll major capsid proteins
Authors: Krupovic Mart , Kuhn Jens H. , Fischer Matthias G. , Koonin Eugene V.
Published: March 2024

The phylumPreplasmiviricota(kingdomBamfordvirae, realmVaridnaviria) is a broad assemblage of diverse viruses with comparatively short double-stranded DNA genomes (<50 kbp) that produce icosahedral capsids built from double jelly-roll major capsid proteins. Preplasmiviricots infect hosts from all cellular domains, testifying to their ancient origin and, in particular, are associated with six of the seven supergroups of eukaryotes. Preplasmiviricots comprise four major groups of viruses, namely, polintons, polinton-like viruses (PLVs), virophages, and adenovirids. We employed protein structure modeling and analysis to show that protein-primed DNA polymerases (pPolBs) of polintons, virophages, and cytoplasmic linear plasmids encompass an N-terminal domain homologous to the terminal proteins (TPs) of prokaryotic PRD1-like tectivirids and eukaryotic adenovirids that are involved in protein-primed replication initiation, followed by a viral ovarian tumor-like cysteine deubiquitinylase (vOTU) domain. The vOTU domain is likely responsible for the cleavage of the TP from the large pPolB polypeptide and is inactivated in adenovirids, in which TP is a separate protein. Many PLVs and transpovirons encode a distinct derivative of polinton-like pPolB that retains the TP, vOTU and pPolB polymerization palm domains but lacks the exonuclease domain and instead contains a supefamily 1 helicase domain. Analysis of the presence/absence and inactivation of the vOTU domains, and replacement of pPolB with other DNA polymerases in eukaryotic preplasmiviricots enabled us to outline a complete scenario for their origin and evolution.

Significance

Structural modeling of protein domains using advanced artificial intelligence-based methods such as AlphaFold2 may lead to insights into evolutionary relationships among proteins that are unreachable by sequence analysis. We applied this approach to elucidate the evolutionary relationships of four major groups of eukaryotic viruses: polintons, polinton-like viruses (PLVs), virophages, and adenovirids. We identified previously uncharacterized protein domains predicted to be essential for virus genome replication. Analysis of the presence/absence and inactivation of these domains suggests a complete scenario for the origin and evolution of this major part of the eukaryotic virosphere.
Single-cell genomics and regulatory networks for 388 human brains
Authors: Emani Prashant S. , Liu Jason J. , Clarke Declan , Jensen Matthew , Warrell Jonathan , Gupta Chirag , Meng Ran , Lee Che Yu , Xu Siwei , Dursun Cagatay , Lou Shaoke , Chen Yuhang , Chu Zhiyuan , Galeev Timur , Hwang Ahyeon , Li Yunyang , Ni Pengyu , Zhou Xiao ,   PsychENCODE Consortium, Bakken Trygve E. , Bendl Jaroslav , Bicks Lucy , Chatterjee Tanima , Cheng Lijun , Cheng Yuyan , Dai Yi , Duan Ziheng , Flaherty Mary , Fullard John F. , Gancz Michael , Garrido-Martín Diego , Gaynor-Gillett Sophia , Grundman Jennifer , Hawken Natalie , Henry Ella , Hoffman Gabriel E. , Huang Ao , Jiang Yunzhe , Jin Ting , Jorstad Nikolas L. , Kawaguchi Riki , Khullar Saniya , Liu Jianyin , Liu Junhao , Liu Shuang , Ma Shaojie , Margolis Michael , Mazariegos Samantha , Moore Jill , Moran Jennifer R. , Nguyen Eric , Phalke Nishigandha , Pjanic Milos , Pratt Henry , Quintero Diana , Rajagopalan Ananya S. , Riesenmy Tiernon R. , Shedd Nicole , Shi Manman , Spector Megan , Terwilliger Rosemarie , Travaglini Kyle J. , Wamsley Brie , Wang Gaoyuan , Xia Yan , Xiao Shaohua , Yang Andrew C. , Zheng Suchen , Gandal Michael J. , Lee Donghoon , Lein Ed S. , Roussos Panos , Sestan Nenad , Weng Zhiping , White Kevin P. , Won Hyejung , Girgenti Matthew J. , Zhang Jing , Wang Daifeng , Geschwind Daniel , Gerstein Mark
Published: March 2024

Single-cell genomics is a powerful tool for studying heterogeneous tissues such as the brain. Yet, little is understood about how genetic variants influence cell-level gene expression. Addressing this, we uniformly processed single-nuclei, multi-omics datasets into a resource comprising >2.8M nuclei from the prefrontal cortex across 388 individuals. For 28 cell types, we assessed population-level variation in expression and chromatin across gene families and drug targets. We identified >550K cell-type-specific regulatory elements and >1.4M single-cell expression-quantitative-trait loci, which we used to build cell-type regulatory and cell-to-cell communication networks. These networks manifest cellular changes in aging and neuropsychiatric disorders. We further constructed an integrative model accurately imputing single-cell expression and simulating perturbations; the model prioritized ∼250 disease-risk genes and drug targets with associated cell types.

Summary Figure
The putative Type 4 secretion system effector BspD is involved in maintaining envelope integrity of the pathogenBrucella
Authors: Ketterer Maren , Chiquet Petra , Sedzicki Jaroslaw , Québatte Maxime , Dehio Christoph
Published: March 2024

Brucellosis is a debilitating disease caused by the Gram-negative, facultative intracellular zoonotic pathogenBrucella.En routeto its intracellular replicative niche,Brucellaencounters various stressful environments that may compromise envelop integrity. Here we show, that the proposed Type 4 secretion system (T4SS) effector BspD is a conserved protein of the Rhizobiales, which does not show signs of co-evolution with the presence of a T4SS or a certain lifestyle. We further present data indicating that BspD is critical forBrucella abortusenvelope integrity in the stationary phase and in the presence of EDTA, a compound known to destabilize the outer membrane. Deletion ofbspDresulted in abnormal bacterial morphologies, indicating its involvement in maintaining envelope integrity. Additionally, the absence of BspD led to the formation of fewer and smaller intracellular microcolonies in a macrophage infection model. From our observations, we propose that BspD ofB. abortusis critical for preserving the integrity of the bacterial envelope, particularly under stressful conditions, which may enhanceBrucella’s ability to survive within host cells.

Importance

Brucellosis, caused by the intracellular pathogenBrucella, poses a significant health threat. Understanding howBrucellaadapts to stressful environments is crucial. This study unveils BspD, a conserved protein within the Rhizobiales order, as a key player in maintainingBrucella’s envelope integrity. Remarkably, BspD’s function appears independent of the presence of a Type 4 Secretion System (T4SS) or specific lifestyle. Deletion ofbspDresulted in compromised envelope integrity, abnormal bacterial morphologies, and reduced intracellular microcolony formation. These findings underscore BspD’s critical role, particularly in stressful conditions like the stationary phase and EDTA exposure, and highlight its significance inBrucella’s survival within host cells. This elucidation deepens our understanding ofBrucellapathogenesis and may inform future therapeutic strategies against brucellosis.
GGCX promotes Eurasian avian-like H1N1 swine influenza virus adaption to interspecies receptor binding
Authors: Zou Jiahui , Jiang Meijun , Xiao Rong , Sun Huimin , Liu Hailong , Peacock Thomas , Tu Shaoyu , Chen Tong , Guo Jinli , Zhao Yaxin , Barclay Wendy , Xie Shengsong , Zhou Hongbo
Published: March 2024

The Eurasian avian-like (EA) H1N1 swine influenza virus (SIV) possesses the capacity to instigate the next influenza pandemic, owing to its heightened affinity for the human-type α-2,6 sialic acid (SA) receptor. Nevertheless, the molecular mechanisms underlying the switch in receptor binding preferences of EA H1N1 SIV remain elusive. In this study, we conducted a comprehensive genome-wide CRISPR/Cas9 knockout screen utilizing EA H1N1 SIV in porcine kidney cells. Knocking out the enzyme gamma glutamyl carboxylase (GGCX) reduced virus replicationin vitroandin vivoby inhibiting the carboxylation modification of viral haemagglutinin (HA) and the adhesion of progeny viruses, ultimately impeding the replication of EA H1N1 SIV. Furthermore, GGCX was revealed to be the determinant of the D225E substitution of EA H1N1 SIV, and GGCX-medicated carboxylation modification of HA 225E contributed to the receptor binding adaption of EA H1N1 SIV to the α-2,6 SA receptor. Taken together, our CRISPR screen has elucidated a novel function of GGCX in the support of EA H1N1 SIV adaption for binding to α-2,6 SA receptor. Consequently, GGCX emerges as a prospective antiviral target against the infection and transmission of EA H1H1 SIV.
Structural analysis of genetic variants of the human tumor suppressor Palb2 coiled-coil domain
Authors: Reddy Pothula Purushotham , Phale Apurva , Das Ranabir
Published: March 2024

Homologous recombination (HR) is an obligate pathway to repair DNA lesions and maintain genome integrity. The tumor suppressor Palb2 is an integral part of the HR pathway that functionally connects BRCA proteins at the site of DNA damage. The N-terminal coiled-coil domain in Palb2 switches from an inactive to an active complex during HR. In the inactive form, Palb2 forms homodimers, and during HR, it assumes the active state, forming a heterodimeric complex with BRCA1. However, the structural details of the human Palb2 coiled-coil domain are unknown. About 600 missense variants have been reported in Palb2, and several are in the coiled-coil domain. The structure-function relationship of such variants is poorly understood, which challenges genetic counseling. Here, we report the solution structure of the human Palb2 coiled-coil domain, which forms an antiparallel homodimer. We then use the structure to study the effect of a few well-characterized missense mutations on the fold and interactions of the Palb2 coiled-coil domain. The structural impact of mutations correlates excellently with their efficiency in homologous recombination, signifying that the approach can be exploited to study other genetic variations in the Palb2.
A packaging signal-binding protein regulates the assembly checkpoint of integrative filamentous phages
Authors: Yeh Ting-Yu , Feehley Michael C. , Feehley Patrick J. , Ooi Vivian Y. , Hung Yi-Yung , Wang Shao-Cheng , Contreras Gregory P.
Published: March 2024

Many integrative filamentous phages not only lack Ff coliphage homologues essential for assembly but also have distinct packaging signals (PS). Their encapsidation remains completely uncharacterized to date. Here we report the first evidence of a PS-dependent checkpoint for integrative filamentous phage assembly. Suppressor screening of PS-deficient phages identified an unknown protein, PSB15 (PS-binding15kDa), crucial for encapsidation. The WAGFXF motif of the PSB15 N-terminus directly binds to PS DNA with conformational change, while suppressor mutations relieve DNA binding specificity constraints to rescue assembly arrest. PSB15 interacts with phospholipid cardiolipin via its basic helix and C-terminus, and recruits PS DNA to the inner membrane (IM). The PSB15-PS complex is released from the IM by interaction between its hydrophobic linker and thioredoxin (Trx), a host protein that is required for Ff assembly but whose mechanisms are still unclear. Live cell imaging shows that thioredoxin and DNA binding regulate the dwelling time of PSB15 at cell poles, suggesting that they both facilitate the dissociation of PSB15 from the IM. Loss of PSB15 or its PS-binding and IM-targeting/dissociation activity compromised virus egress, indicating that the PS/PSB15/Trx complex establishes a regulatory phage assembly checkpoint critical for integrative phage infection and life cycles.
Energy landscapes of peptide-MHC binding
Authors: Collesano Laura , Łuksza Marta , Lässig Michael
Published: March 2024

Molecules of the Major Histocompatibility Complex (MHC) present short protein fragments peptides on the cell surface, an important step in T cell immune recognition. MHC-I molecules process peptides from intracellular proteins; MHC-II molecules act in antigen-presenting cells and present peptides derived from extracellular proteins. Here we show that the sequence-dependent energy landscapes of MHC-peptide binding encode class-specific nonlinearities (epistasis). MHC-I has a smooth landscape with global epistasis; the binding energy is a simple deformation of an underlying linear trait. This form of epistasis enhances the discrimination between strong-binding peptides. In contrast, MHC-II has a rugged landscape with idiosyncratic epistasis: binding depends on detailed amino acid combinations at multiple positions of the peptide sequence. The form of epistasis affects the learning of energy landscapes from training data. For MHC-I, a low-complexity problem, we derive a simple matrix model of binding energies that outperforms current models trained by machine learning. For MHC-II, higher complexity prevents learning by simple regression methods. Epistasis also affects the energy and fitness effects of mutations in antigen-derived peptides (epitopes). In MHC-I, large-effect mutations occur predominantly in anchor positions of strong-binding epitopes. In MHC-II, large effects depend on the background epitope sequence but are broadly distributed over the epitope, generating a bigger target for escape mutations from T cell immunity than for MHC-I.

Author Summary

T cell immunity involves the binding of short peptides to the intracellular MHC recognition machinery. Understanding how the binding energy depends on the peptide sequence is key to computationally predict immune recognition and immune escape evolution, for example, of pathogens and cancer cells. We find nonlinear energy landscapes that depend on the recognition pathway: smooth and easy to learn for MHC class I, rugged and difficult to learn for class II. Together, this work establishes links between biophysical origin, nonlinear structure, learnability from data, and biological implications for protein interaction landscapes.
Adaptation ofPseudomonas aeruginosato repeated invasion into a commensal competitor
Authors: Wheatley Rachel M. , Shaw Liam P. , Shah Sarah , Lindon Selina , MacLean R. Craig
Published: March 2024

The host-associated microbiome is an important barrier to bacterial pathogen colonization and can mediate protection through a variety of mechanisms. We wanted to investigate the potential consequences of selection imposed by commensal bacterial competitors on an invading bacterial pathogen. To do this, we tested the ability of the opportunistic pathogenPseudomonas aeruginosato invade pre-established communities of an abundant commensal bacterium in the human microbiome,Staphylococcus epidermidis. We passaged ten independent lines ofP. aeruginosathrough daily invasion into a pre-establishedS. epidermidispopulation (coculture evolved lines), alongside daily passage through monoculture conditions (monoculture evolved lines). The monoculture evolved lines showed strong parallel evolution in the Wsp (Wrinkly spreader phenotype) signal transducing system involved in biofilm formation, and significantly elevated biofilm formation. On the other hand, adaptation toS. epidermidisoccurred via mutations in a diverse set of genes, and the coculture evolved lines showed much weaker evidence for parallel evolution, suggesting that the selective pressure imposed by competition withS. epidermidisis more complex than the pressure imposed by culture conditions. Interestingly, the elevated biofilm formation phenotype seen in the monoculture evolved lines was not observed in the lines evolved in the presence ofS. epidermidis, raising the question of whether enhanced biofilm formation did not evolve withS. epidermidispresent because it was not beneficial, or becauseS. epidermidismay be able to restrict this evolutionary path by inhibiting biofilm formation.
Concepts and methods for predicting viral evolution
Authors: Meijers Matthijs , Ruchnewitz Denis , Eberhardt Jan , Karmakar Malancha , Łuksza Marta , Lässig Michael
Published: March 2024

The seasonal human influenza virus undergoes rapid evolution, leading to significant changes in circulating viral strains from year to year. These changes are typically driven by adaptive mutations, particularly in the antigenic epitopes, the regions of the viral surface protein haemagglutinin targeted by human antibodies. Here we describe a consistent set of methods for data-driven predictive analysis of viral evolution. Our pipeline integrates four types of data: (1) sequence data of viral isolates collected on a worldwide scale, (2) epidemiological data on incidences, (3) antigenic characterization of circulating viruses, and (4) intrinsic viral phenotypes. From the combined analysis of these data, we obtain estimates of relative fitness for circulating strains and predictions of clade frequencies for periods of up to one year. Furthermore, we obtain comparative estimates of protection against future viral populations for candidate vaccine strains, providing a basis for pre-emptive vaccine strain selection. Continuously updated predictions obtained from the prediction pipeline for influenza and SARS-CoV-2 are available on the websiteprevir.app.