bluuebunny/crossref_metadata_2025 huggingface dataset:
Altered GM1 catabolism affects NMDAR-mediated Ca2+signaling at ER-PM junctions and increases synaptic spine formation
Authors: Weesner Jason A. , Annunziata Ida , van de Vlekkert Diantha , Robinson Camenzind G. , Campos Yvan , Mishra Ashutosh , Fremuth Leigh E. , Gomero Elida , Hu Huimin , d’Azzo Alessandra
Published: July 2023

Endoplasmic reticulum–plasma membrane (ER-PM) junctions mediate Ca2+flux across neuronal membranes. The properties of these membrane contact sites are defined by their lipid content, but little attention has been given to glycosphingolipids (GSLs). Here, we show that GM1-ganglioside, an abundant GSL in neuronal membranes, is integral to ER-PM junctions; it interacts with synaptic proteins/receptors and regulates Ca2+signaling. In a model of the neurodegenerative lysosomal storage disease, GM1-gangliosidosis, pathogenic accumulation of GM1 at ER-PM junctions due to β-galactosidase deficiency drastically alters neuronal Ca2+homeostasis. Mechanistically, we show that GM1 interacts with the phosphorylated NMDAR Ca2+channel, thereby increasing Ca2+flux, activating ERK signaling, and increasing the number of synaptic spines without increasing synaptic connectivity. Thus, GM1 clustering at ER-PM junctions alters synaptic plasticity and exacerbates the generalized neuronal cell death characteristic of GM1-gangliosidosis.
Position-Specific Enrichment Ratio Matrix scores predict antibody variant properties from deep sequencing data
Authors: Smith Matthew D. , Case Marshall A. , Makowski Emily K. , Tessier Peter M.
Published: July 2023

Motivation

Deep sequencing of antibody and related protein libraries after phage or yeast-surface display sorting is widely used to identify variants with increased affinity, specificity and/or improvements in key biophysical properties. Conventional approaches for identifying optimal variants typically use the frequencies of observation in enriched libraries or the corresponding enrichment ratios. However, these approaches disregard the vast majority of deep sequencing data and often fail to identify the best variants in the libraries.

Results

Here, we present a method, Position-Specific Enrichment Ratio Matrix (PSERM) scoring, that uses entire deep sequencing datasets from pre- and post-selections to score each observed protein variant. The PSERM scores are the sum of the site-specific enrichment ratios observed at each mutated position. We find that PSERM scores are much more reproducible and correlate more strongly with experimentally measured properties than frequencies or enrichment ratios, including for multiple antibody properties (affinity and non-specific binding) for a clinical-stage antibody (emibetuzumab). We expect that this method will be broadly applicable to diverse protein engineering campaigns.

Availability

All deep sequencing datasets and code to do the analyses presented within are available via GitHub.

Contact

Peter Tessier,ptessier@umich.edu

Supplementary information

Supplementary data are available at Bioinformatics online.
Estimation of indirect genetic effects and heritability under assortative mating
Authors: Young Alexander Strudwick
Published: July 2023

Both direct genetic effects (effects of alleles in an individual on that individual) and indirect genetic effects — effects of alleles in an individual (e.g. parents) on another individual (e.g. offspring) — can contribute to phenotypic variation and genotype-phenotype associations. Here, we consider a phenotype affected by direct and parental indirect genetic effects under assortative mating at equilibrium. We generalize classical theory to derive a decomposition of the equilibrium phenotypic variance in terms of direct and indirect genetic effect components. We extend this theory to show that popular methods for estimating indirect genetic effects or ‘genetic nurture’ through analysis of parental and offspring polygenic predictors (called polygenic indices or scores — PGIs or PGSs) are substantially biased by assortative mating. We propose an improved method for estimating indirect genetic effects while accounting for assortative mating that can also correct heritability estimates for bias due to assortative mating. We validate our method in simulations and apply it to PGIs for height and educational attainment (EA), estimating that the equilibrium heritability of height is 0.699 (S.E. = 0.075) and finding no evidence for indirect genetic effects on height. We estimate a very high correlation between parents’ underlying genetic components for EA, 0.755 (S.E. = 0.035), which is inconsistent with twin based estimates of the heritability of EA, possibly due to confounding in the EA PGI and/or in twin studies. We implement our method in the software packagesnipar, enabling researchers to apply the method to data including observed and/or imputed parental genotypes. We provide a theoretical framework for understanding the results of PGI analyses and a practical methodology for estimating heritability and indirect genetic effects while accounting for assortative mating.
Algal kainoid synthases exhibit substrate-dependent hydroxylation and cyclization activities
Authors: Hopiavuori Austin R. , McKinnie Shaun M. K.
Published: July 2023

FeII/α-ketoglutarate-dependent dioxygenases (Fe/αKG) are a large enzyme family that functionalize C-H bonds on diverse organic substrates. Although Fe/αKG homologs catalyze an array of chemically useful reactions, hydroxylation typically predominates. Microalgal DabC uniquely forms a novel C-C bond to construct the bioactive pyrrolidine ring in domoic acid biosynthesis. However, this kainoid synthase exclusively performs a stereospecific hydroxylation reaction on itscissubstrate regioisomer. Mechanistic and kinetic analyses with native and alternative substrates identified a 20-fold rate increase in DabC radical cyclization over β–hydroxylation, with no observable 1,5-hydrogen atom transfer. Moreover, this dual activity was conserved among macroalgal RadC1 and KabC homologs and provided insight into substrate recognition and reactivity trends. Investigation of this substrate-dependent chemistry improves our understanding of Fe/αKG enzymes and their biocatalytic application.
Association between diabetes and levels of micro-nutrients in a Middle Eastern population - A case-control study
Authors: Soliman Nada , Almishal Ruba , Elsayed Basant , Ahmed Ayaaz , Al-Amri Sara , Al-Kuwari Aisha , Al-Muhannadi Shaikha , Nadeer Muhammed , Chivese Tawanda
Published: July 2023

Objective

To investigate associations between levels of micronutrients and diabetes, controlled and uncontrolled diabetes.

Methods

A case-control study, matched on age and gender, was performed on participants with (cases) and without diabetes (controls), who were Qatari or long-term residents. Participants with diabetes were divided into those with controlled and uncontrolled diabetes using an HbA1c cutoff of 7%. Levels of micronutrients were measured from serum and categorized into normal and abnormal levels.

Results

A total of 1118 participants (374 cases and 744 controls) were included with a mean age of 41.7 years (SD 9.9), of whom 53.9% were female. Of those with diabetes, 229 had controlled diabetes and 145 had uncontrolled diabetes. Compared to those without diabetes, participants with diabetes had significantly lower mean magnesium (0.80 mmol/L (SD 0.07) vs 0.84 mmol/L (SD 0.06), respectively, p<0.001). Lower magnesium and iron were observed in participants with uncontrolled compared to participants with controlled diabetes. After multivariable logistic regression, diabetes was associated with hypomagnesemia (OR 3.2, 95% CI 3.4-213.9), and low iron (OR 1.49, 95% CI 1.03-2.15). Uncontrolled diabetes showed stronger odds of association with hypomagnesemia (OR 5.57, 95% CI 3.65-8.52).

Conclusion

In an affluent setting in the Middle East and North Africa (MENA) region, diabetes was associated with low magnesium and low iron, and this association was worse in individuals with uncontrolled diabetes.
Anatomy-informed multimodal learning for myocardial infarction prediction
Authors: Sievering Ivan-Daniel , Senouf Ortal , Mahendiran Thabo , Nanchen David , Fournier Stephane , Muller Olivier , Frossard Pascal , Abbé Emmanuel , Thanou Dorina
Published: July 2023

In patients with coronary artery disease the prediction of future cardiac events such as myocardial infarction (MI) remains a major challenge. In this work, we propose a novel anatomy-informed multimodal deep learning framework to predict future MI from clinical data and Invasive Coronary Angiography (ICA) images. The images are analyzed by Convolutional Neural Networks (CNNs) guided by anatomical information, and the clinical data by an Artificial Neural Network (ANN). Embeddings from both sources are then merged to provide a patient-level prediction. The performance of our framework on a clinical study of 445 patients admitted with acute coronary syndromes confirms that multimodal learning increases the predictive power and achieves good performance, which outperforms the prediction obtained by each modality independently as well as that of interventional cardiologists. To the best of our knowledge, this is the first attempt towards combining multimodal data through a deep learning framework for future MI prediction.
Impact of environmental factors on calling behavior in midshipman fish across ocean basins
Authors: Kok Annebelle C.M. , Kim Ella B. , Rowell Timothy J. , Margolina Tetyana , Joseph John E. , Reeves Lindsey E. Peavey , Hatch Leila T. , Baumann-Pickering Simone
Published: July 2023

Chorusing is widespread across the animal kingdom. Animal calling behavior is often driven by phenological and environmental factors such as seasonality, lunar period, and temperature. Now, in the Anthropocene, factors such as increased anthropogenic noise levels are also affecting calling behavior. Many fish call in choruses to attract mates, but the dynamics that drive fish calling behavior have rarely been studied in the field. We investigated how seasonality, lunar period, ambient noise, and temperature influenced the calling behavior of two species of toadfish, the plainfin midshipman (Porichthys notatus) and putatively, the Atlantic midshipman (Porichthys plectrodon). Acoustic recordings from a two-year period in twelve different locations, spanning two ocean basins showed that midshipman chorus presence was driven by seasonality and lunar period. Furthermore, chorus frequency increased with increasing temperature. Chorus levels were strongly influenced by seasonality and increased somewhat with increasing noise levels. Taken together, these results indicate that midshipman calling behavior was strongly influenced by interacting environmental conditions. Understanding the various impacts of each driver will facilitate predictions of changes in midshipman calling due to future changes in environmental conditions.
Specialized mesenteric lymphatic capillaries by-pass the mesenteric lymph node chain to transport peritoneal antigens directly into mediastinal lymph nodes
Authors: Redder Esther , Kirschnick Nils , Fang Shentong , Kuhlmann Michael , González-Loyola Alejandra , Wyss Tania , Stehling Martin , Adams Ralf H. , Petrova Tatiana V. , Alitalo Kari , Rot Antal , Kiefer Friedemann
Published: July 2023

Lymphatic vessels (LVs) are indispensable for tissue fluid homeostasis and immune cell trafficking. The network of LVs that channel fluids from the gut into mesenteric lymph nodes (MLN) has been recognized as the sole lymphatic system in the mesentery. Here we describe an alternative, functionally autonomous set of capillary mesenteric LVs (capMLVs) that by-pass the MLNs and drain directly into mediastinal LNs. CapMLVs develop perinatally from valves of collective mesenteric lymphatic vessels (colMLVs) in response to arterial endothelial cell-derived VEGF-C. Once extended, capMLVs detach from colMLVs to form an independent elongated network comprised of LYVE1+, CCL21+ endothelial cells. Avascular areas of the mesentery juxtaposed to capMLVs contain cell islets that express ACKR4. This CCL21-scavenging atypical receptor facilitates the migration of mesenteric phagocytes into capMLVs to be channeled directly into mediastinal LNs. This allows peritoneum-derived ominous antigens to be processed separately from alimentary antigens.
Oncogene EVI1 Drives Acute Myeloid Leukemia Via a Targetable Interaction with CTBP2
Authors: Pastoors Dorien , Havermans Marije , Mulet-Lazaro Roger , Brian Duncan , Noort Willy , Grasel Julius , Hoogenboezem Remco , Smeenk Leonie , Demmers Jeroen A.A. , Milsom Michael D. , Enver Tariq , Groen Richard WJ , Bindels Eric , Delwel Ruud
Published: July 2023

Acute myeloid leukemia (AML) driven by the activation of EVI1 due to chromosome 3q26/MECOM rearrangements is incurable. Since transcription factors like EVI1 are notoriously hard to target, insight into the mechanism by which EVI1 drives myeloid transformation could provide alternative avenues for therapy. Applying protein folding predictions combined with proteomics technologies, we demonstrate that interaction with CTBP1 and CTBP2 via a single PLDLS motif in EVI1 is indispensable for leukemic transformation. Furthermore, we show that a 4x PLDLS repeat construct outcompetes binding of EVI1 to CTBP1 and CTBP2 and thereby inhibits proliferation of 3q26/MECOM rearranged AML both in in vitro and in xenotransplant models. This proof-of-concept study opens the possibility to therapeutically target one of the most incurable forms of AML with specific EVI1-CTBP inhibitors. This has important implications for other tumour types with aberrant expression of EVI1 as well as for cancers transformed by distinct CTBP-dependent oncogenic transcription factors.
Effects of noise and metabolic cost on cortical task representations
Authors: Stroud Jake P. , Wójcik Michał , Jensen Kristopher T. , Kusunoki Makoto , Kadohisa Mikiko , Buckley Mark J. , Duncan John , Stokes Mark G. , Lengyel Máté
Published: July 2023

Cognitive flexibility requires both the encoding of task-relevant and the ignoring of task-irrelevant stimuli. While the neural coding of task-relevant stimuli is increasingly well understood, the mechanisms for ignoring task-irrelevant stimuli remain poorly understood. Here, we study how task performance and biological constraints jointly determine the coding of relevant and irrelevant stimuli in neural circuits. Using mathematical analyses and task-optimized recurrent neural networks, we show that neural circuits can exhibit a range of representational geometries depending on the strength of neural noise and metabolic cost. By comparing these results with recordings from primate prefrontal cortex (PFC) over the course of learning, we show that neural activity in PFC changes in line with a minimal representational strategy. Specifically, our analyses reveal that the suppression of dynamically irrelevant stimuli is achieved by activity-silent, sub-threshold dynamics. Our results provide a normative explanation as to why PFC implements an adaptive, minimal representational strategy.
Salmonella entericaserovar Typhimurium ST313 sublineage 2.2 has emerged in Malawi with a characteristic gene expression signature and a fitness advantage
Authors: Kumwenda Benjamin , Canals Rocío , Predeus Alexander V. , Zhu Xiaojun , Kröger Carsten , Pulford Caisey , Wenner Nicolas , Lora Lizeth Lacharme , Li Yan , Owen Siân V. , Everett Dean , Hokamp Karsten , Heyderman Robert S. , Ashton Philip M. , Gordon Melita A. , Msefula Chisomo L. , Hinton Jay C. D.
Published: July 2023

Invasive non-typhoidalSalmonella(iNTS) disease is a serious bloodstream infection that targets immune-compromised individuals, and causes significant mortality in sub-Saharan Africa.Salmonella entericaserovar Typhimurium ST313 causes the majority of iNTS in Malawi, and we performed an intensive comparative genomic analysis of 608 isolates obtained from fever surveillance at the Queen Elizabeth Hospital, Blantyre between 1996 and 2018. We discovered that following the upsurge of the well-characterisedS.Typhimurium ST313 lineage 2 from 1999 onwards, two new multidrug-resistant sublineages designated 2.2 and 2.3, emerged in Malawi in 2006 and 2008, respectively. The majority ofS.Typhimurium isolates from human bloodstream infections in Malawi now belong to sublineage 2.2 or 2.3. To identify factors that characterised the emergence of the prevalent ST313 sublineage 2.2, we performed genomic and functional analysis of two representative strains, D23580 (lineage 2) and D37712 (sublineage 2.2). Comparative genomic analysis showed that the chromosome of ST313 lineage 2 and sublineage 2.2 were broadly similar, only differing by 29 SNPs and small indels and a 3kb deletion in the Gifsy-2 prophage region that spanned thesseIpseudogene. Lineage 2 and sublineage 2.2 have unique plasmid profiles that were verified by long read sequencing. The transcriptome was initially explored in 15 infection-relevant conditions and within macrophages. Differential gene expression was subsequently investigated in depth in the four most importantin vitrogrowth conditions. We identified up-regulation of SPI2 genes in non-inducing conditions, and down-regulation of flagellar genes in D37712, compared to D23580. Following phenotypic confirmation of transcriptional differences, we discovered that sublineage 2.2 had increased fitness compared with lineage 2 during mixed-growth in minimal media. We speculate that this competitive advantage is contributing to the continuing presence of sublineage 2.2 in Malawi.
Potency of CRISPR-Cas Antifungals Is Enhanced by Co-targeting DNA Repair and Growth Regulatory Machinery at the Genetic Level
Authors: Mendoza Brian J. , Zheng Xianliang , Clements Jared C. , Cotter Christopher , Trinh Cong T.
Published: July 2023

Fungal pathogens are virulent and resistant to antibiotic treatment. Due to their adaptability, specificity, and efficiency, CRISPR-Cas systems can be harnessed for neutralizing these fungal pathogens. However, the conventional design of CRISPR-Cas antimicrobials, based on induction of DNA double-strand-breaks (DSBs), is potentially less effective in fungi due to robust eukaryotic DNA repair machinery. Here, we report a novel design principle to formulate more effective CRISPR-Cas antifungals by co-targeting essential genes with DNA repair defensive genes that remove the fungi’s ability to repair the DSB sites of essential genes. By evaluating this design on the model fungusSaccharomyces cerevisiae, we demonstrated that essential and defensive gene co-targeting is more effective than either essential or defensive gene targeting alone. The top-performing CRISPR-Cas antifungals performed as effectively as the antibiotic Geneticin. Fast growth kinetics ofS. cerevisiaeinduced resistance to CRISPR-Cas antifungals where genetic mutations mostly occurred in defensive genes and guide RNA sequences.

Significance

The emergence of virulent, resistant, and rapidly evolving fungal pathogens poses a significant threat to public health, agriculture, and the environment. Targeting cellular processes with standard small-molecule intervention may be effective but requires long development times and is prone to antibiotic resistance. To overcome the current limitation of antibiotic development and treatment, this study harnesses CRISPR-Cas systems as antifungals by capitalizing on their adaptability, specificity, and efficiency in target design. Simultaneous co-targeting of both essential and defensive genes is shown to be a novel design principle for formulating effective CRISPR-Cas antimicrobials that can be rapidly tuned to adapt to inevitable escapee events.
The pelvic organs receive no parasympathetic innervation
Authors: Sivori Margaux , Dempsey Bowen , Chettouh Zoubida , Boismoreau Franck , Ayerdi Maïlys , Eymael Annaliese Nucharee , Baulande Sylvain , Lameiras Sonia , Coulpier Fanny , Delattre Olivier , Rohrer Hermann , Mirabeau Olivier , Brunet Jean-François
Published: July 2023

The pelvic organs (bladder, rectum and sex organs) have been represented for a century as receiving autonomic innervation from two pathways — lumbar sympathetic and sacral parasympathetic — by way of a shared relay, the pelvic ganglion, conceived as an assemblage of sympathetic and parasympathetic neurons. Using single cell RNA sequencing, we find that the mouse pelvic ganglion is made of four classes of neurons, distinct from both sympathetic and parasympathetic ones, albeit with a kinship to the former, but not the latter, through a complex genetic signature. We also show that spinal lumbar preganglionic neurons synapse in the pelvic ganglion onto equal numbers of noradrenergic and cholinergic cells, both of which therefore serve as sympathetic relays. Thus, the pelvic viscera receive no innervation from parasympathetic or typical sympathetic neurons, but instead from a divergent tail end of the sympathetic chains, in charge of its idiosyncratic functions.
Core enhancers of the 3’RR optimizeIgHnuclear position and loop conformation for oriented CSR
Authors: Bruzeau Charlotte , Pollet Justine , Thomas Morgane , Ba Zhaoqing , Roulois David , Pinaud Eric , Le Noir Sandrine
Published: July 2023

Class switch recombination is an essential process which enabling B cells to adapt immunoglobulin subtypes to antigens. Transcription plays a crucial role in regulating CSR in which theIgH 3’Regulatory Region(3’RR) was identified as a key player. The3’RRstands at the 3’ end ofIgHlocus and is composed of four core enhancers surrounded by inverted repeated sequences, forming a quasi-palindrome. In addition to transcriptional control, nuclear organization appears to be an important level in CSR regulation. Furthermore, the chromatin loops atIgHlocus facilitate an efficient CSR recombination by bringing the donor and acceptor switch regions closer together. However, the precise control mechanisms governing both of these processes remain partially understood. Here, using the reference DNA 3D-FISH technique combined with various high throughput approaches, we showed that 3’RR core enhancers are necessary and sufficient to preorganize resting B cell nuclei to facilitate a deletional CSR mechanism at activated stage. We demonstrated that the 3’RR core enhancers regulateIgHlocus addressing in the nuclei, controlIgHlocus accessibility and orchestrateIgHloops formation. Our findings pinpointed an additional regulation level of mechanisms underlying B cell diversification.

Graphical abstract

Position ofIgHloci through B cell development (from transitional to stimulated stages) is represented by the red spots. Inwtandc3’RRcontext,IgHloci get closer from each other and from nucleus center during evolution from transitional to mature resting stage and they relocates more at nuclear periphery, away one from each other, uponin vitrostimulation. InΔ3’RRmodel, this dynamic is lost and, moreover,IgHloci are more localized to pericentromeric heterochromatin (represented by green area) since the mature resting B cell stage and remain in afterin vitrostimulation.
Cross-modal interactions between auditory attention and oculomotor control
Authors: Zhao Sijia , Contadini-Wright Claudia , Chait Maria
Published: July 2023

Microsaccades are small, involuntary eye movements that occur during fixation. Their role is debated with recent hypotheses proposing a contribution to automatic scene sampling. Microsaccade inhibition (MSI) refers to the abrupt suppression of microsaccades, typically evoked within 0.1 seconds after new stimulus onset. The functional significance and neural underpinnings of MSI are subjects of ongoing research. It has been suggested that MSI is a component of the brain’s attentional re-orienting network which facilitates the allocation of attention to new environmental occurrences by reducing disruptions or shifts in gaze that could interfere with processing.

The extent to which MSI is reflexive or influenced by top-down mechanisms remains debated. We developed a task that examines the impact of auditory top-down attention on MSI, allowing us to disentangle ocular dynamics from visual sensory processing. Participants (N=24 and 27; both sexes) listened to two simultaneous streams of tones and were instructed to attend to one stream while detecting specific task “targets.” We quantified MSI in response to occasional task-irrelevant events presented in both the attended and unattended streams (frequency steps in Experiment 1, omissions in Experiment 2).

The results show that initial stages of MSI are not affected by auditory attention. However, later stages (∼0.25s post event-onset), affecting the extent and duration of the inhibition, are enhanced for sounds in the attended stream compared to the unattended stream. These findings provide converging evidence for the reflexive nature of early MSI stages and robustly demonstrate the involvement of auditory attention in modulating the later stages.

Significance Statement

Microsaccades are rapid eye movements occurring during fixation. Their precise role is not known but a major hypothesis is that they reflect automatic sampling of the environment. A feature of microsaccades is that they exhibit abrupt suppression (MSI) after the presentation of new stimuli. This is thought to be part of attentional re-orienting. To understand the neural circuit that controls MSI, and by extension, the brain’s response to novel events in the environment, it is essential to determine which factors affect MSI. We show, for the first time, that auditory attention affects the latter (but not initial) stages of MSI. Thus, the early stages of MSI are automatic, but subsequent phases are affected by the perceptual state of the individual.
Tolerance toHaemophilus influenzaeinfection in human epithelial cells: insights from a primary cell-based model
Authors: Kappler Ulrike , Henningham Anna , Nasreen Marufa , Buultjens Andrew H. , Stinear Timothy P. , Sly Peter , Fantino Emmanuelle
Published: July 2023

Haemophilus influenzaeis a human respiratory pathogen and inhabits the human respiratory tract as its only niche. Despite this, the molecular mechanisms that allowH. influenzaeto establish persistent infections of human epithelia are not well understood.

Here, we have investigated howH. influenzaeadapts to the host environment and triggers the host immune response using a human primary cell-based infection model that closely resembles human nasal epithelia (NHNE).

Physiological assays combined with dualRNAseq revealed that NHNE from five healthy donors all responded toH. influenzaeinfection with an initial, ‘unproductive’ inflammatory response that included a strong hypoxia signature but did not produce pro-inflammatory cytokines. Subsequently, an apparent tolerance to large extra- and intracellular burdens ofH. influenzaedeveloped, with NHNE transcriptional profiles resembling the pre-infection state. This occurred in parallel with the development of intracellular bacterial populations, and appears to involve interruption of NFkB signalling. This is the first time that large-scale, persistence-promoting immunomodulatory effects ofH. influenzaeduring infection have been u. Interestingly, NHNE were able to re-activate pro-inflammatory responses towards the end of the 14-day infection resulting in release of pro-inflammatory cytokines (IL8, TNFα). Our data further indicate the presence of infection stage-specific gene expression modules, highlighting fundamental similarities between immune responses in NHNE and canonical immune cells, which merit further investigation.
Patterns of recombination in snakes reveal a tug of war between PRDM9 and promoter-like features
Authors: Hoge Carla , de Manuel Marc , Mahgoub Mohamed , Okami Naima , Fuller Zachary , Banerjee Shreya , Baker Zachary , McNulty Morgan , Andolfatto Peter , Macfarlan Todd S. , Schumer Molly , Tzika Athanasia C. , Przeworski Molly
Published: July 2023

In vertebrates, there are two known mechanisms by which meiotic recombination is directed to the genome: in humans, mice, and other mammals, recombination occurs almost exclusively where the protein PRDM9 binds, while in species lacking an intactPRDM9, such as birds and canids, recombination rates are elevated near promoter-like features. To test if PRDM9 also directs recombination in non-mammalian vertebrates, we focused on an exemplar species, the corn snake (Pantherophis guttatus). Unlike birds, this species possesses a single, intactPRDM9ortholog. By inferring historical recombination rates along the genome from patterns of linkage disequilibrium and identifying crossovers in pedigrees, we found that PRDM9 specifies the location of recombination events outside of mammals. However, we also detected an independent effect of promoter-like features on recombination, which is more pronounced on macrothan microchromosomes. Thus, our findings reveal that the uses of PRDM9 and promoter-like features are not mutually-exclusive, and instead reflect a tug of war, which varies in strength along the genome and is more lopsided in some species than others.

One sentence summary

While the localization of meiotic recombination in vertebrates was previously thought to occur using one of two distinct mechanisms, our analysis of recombination in corn snakes reveals that they and likely other vertebrates use both of these mechanisms.
A fluorescent assay for cryptic transcription inSaccharomyces cerevisiaereveals novel insights into factors that stabilize chromatin structure on newly replicated chromatin
Authors: Gao Ellia , Jung Stephanie , Howe LeAnn J.
Published: July 2023

The disruption of chromatin structure can result in transcription initiating from cryptic promoters. A well-characterized, chromatin-destabilizing stress is the passage of RNA polymerase, and numerous factors function to stabilize chromatin on transcribed genes, suppressing cryptic transcription from sites within gene bodies. DNA replication is also inherently disruptive to chromatin, and multiple replication-coupled histone chaperones suppress cryptic transcription. However, these factors also have documented roles in transcription, and thus whether DNA replication per se can activate cryptic promoters has not been directly examined. In this study, we tested the hypothesis that, in the absence of chromatin-stabilizing factors, DNA replication can promote cryptic transcription inS. cerevisiae. Using a novel fluorescent reporter assay, we show that multiple factors, including Asf1, Rtt106, Spt6, and Spt16, suppress transcription from a cryptic promoter, but are entirely or partially dispensable in G1-arrested cells, suggesting a requirement for DNA replication in chromatin disruption. Additionally, for the first time, we demonstrate modest cryptic transcription following the depletion of Rlf2/Cac1, a CAF-1 chromatin assembly complex component. Collectively, these results suggest that transcription fidelity is dependent on numerous factors that function to assemble chromatin on nascent DNA.
Whole-genome Omics delineates the function of CCM1 within the CmPn networks
Authors: Croft Jacob , Grajeda Brian , Aguirre Luis A , Gao Liyuan , Abou-Fadel Johnathan , Sheng Victor , Zhang Jun
Published: July 2023

Introduction

Cerebral cavernous malformations (CCMs) are abnormal dilations of brain capillaries that increase the risk of hemorrhagic strokes. Mutations in the KRIT1, MGC4607, and PDCD10 genes cause CCMs, with mutations in CCM1 accounting for about 50% of familial cases. The disorder exhibits incomplete penetrance, meaning that individuals with CCM may appear normal initially, but once symptoms manifest, their brains have already suffered irreversible damage. Compromised blood-brain barrier (BBB) is crucial in regulating the flow of substances between the blood and the central nervous system, which can result in hemorrhagic CCMs. Progesterone and its derivatives have been studied for their impact on maintaining BBB integrity. CCM2 interacts with CCM1 and CCM3, forming the CCM signaling complex (CSC), which connects classic and non-classic progesterone signaling to establish the CmPn signaling network, vital in preserving BBB integrity.

Methods

The study aimed to explore the relationship between CCM1 and key pathways of the CmPn signaling network, utilizing a toolset comprising three mouse embryonic fibroblast lines (MEFs) with distinct CCM1 expression levels. Omics and systems biology analysis were performed to investigate Ccm1-mediated signaling within the CmPn signaling network.

Results

The findings suggest that CCM1 plays a critical role in controlling cellular processes in response to different progesterone-mediated actions within CmPn/CmP signaling networks, partly by regulating gene transcription. This function is crucial for preserving the integrity of microvessels, indicating that targeting CCM1 could hold promise as a therapeutic approach for this condition.
LC3B conjugation machinery promotes autophagy-independent HIV-1 entry in CD4+ T lymphocytes
Authors: Pradel Baptiste , Deffieu Maïka S. , Robert-Hebmann Véronique , Cantaloube Guilhem , Faure Mathias , Chazal Nathalie , Gaudin Raphaël , Espert Lucile
Published: July 2023

HIV-1 entry into CD4+ T lymphocytes relies on the viral and cellular membranes’ fusion, leading to viral capsid delivery in the cytoplasm of target cells. The conjugation of ATG8/LC3B protein, process referred to as ATG8ylation and mainly studied in the context of autophagy, occurs transiently in the early stages of the HIV-1 replication cycle in CD4+ T lymphocytes. Despite numerous studies investigating the interplays of HIV-1 with autophagy machinery, the impact of ATG8ylation in the early stages of HIV-1 infection remains unknown. Here we found that HIV-1 exposure leads to the rapid enrichment of LC3B towards the target cell plasma membrane, in close proximity with the incoming viral particles. Furthermore, we demonstrated that ATG8ylation is a key event that facilitates HIV-1 fusion with target CD4+ T cells. Interestingly, this effect is independent of the canonical autophagy pathway as ATG13 silencing does not prevent HIV-1 entry. Together, our results provide an unconventional role of LC3B conjugation subverted by HIV-1 to achieve a critical early step of its replication cycle.

Teaser

HIV-1 induces LC3B enrichment towards its target cell entry site and uses the conjugation of this protein to favor its entry step.
Exercise on cytokine responses in males and females: Effect of leucine, HMB and BCAA
Authors: Brown Alexander D. , Grammenou Maria , Tee Chris C.L. , Chong Mee Chee , Stewart Claire E.
Published: July 2023

Purpose

To identify the effects of leucine, β-hydroxy β-methylbutyrate (HMB) and branched chain amino acid (BCAA) on post-exercise cytokine responses in females and males.

Methods

Males (n=53) and females (n=37) completed 100 drop jumps and consumed either no supplement, leucine (3g/d), HMB (3g/d) or BCAA (4.5g/d) from 1d pre to 14d post-exercise. Muscle soreness, squat jumps, chair rises and creatine kinase (CK) were measured at pre, post, 24h, 48h, 7 and 14d. Blood lactate (pre, post), 10 cytokines (pre, 24h, 48h, 7d) and oestradiol (pre, 7d) were also measured.

Results

Without supplementation post-exercise, soreness was induced in both males (6-fold) and females (5-fold). With supplementation, there were no increases in CK or oestradiol in females and no impact on muscle soreness, performance, or function in both sexes. In males, CK was elevated in untreated (48%) and leucine (69%) conditions vs baseline, but these were suppressed with HMB and BCAA. IL-7 was elevated in females vs males at baseline (6.3-fold), leucine increased IL-7 concentrations in females at 24h (17.0-fold), 48h (5.1-fold) vs males. With HMB, TNFr1-α increased in females at 24h (2.2-fold), 48h (2.3-fold) and 7d (2.3-fold) vs males. In males with BCAA, TNFr1-α decreased (P=0.06) from pre to 24h (6.8-fold), then increased (P<0.05) from 24 to 48h (8.0-fold).

Conclusion

Although supplements were without effect on soreness following exercise, the cytokine response was evoked by exercise and impacted significantly by leucine, HMB and BCAA in females vs males. This improved cytokine response in females could lead to improved resistance to damage.
Seasonal influenza viruses decay more rapidly at intermediate humidity in droplets containing saliva compared to respiratory mucus
Authors: Rockey Nicole C. , Le Sage Valerie , Marr Linsey C. , Lakdawala Seema S.
Published: July 2023

Expulsions of virus-laden aerosols or droplets are an important source of onward respiratory virus transmission and can originate from both the oral and nasal cavity of an infected host. However, the presence of infectious influenza virus in the oral cavity during infection has not been widely considered, and thus little work has explored the environmental persistence of influenza virus in oral cavity expulsions that may facilitate transmission. Using the ferret model, we detected infectious virus in the nasal and oral cavities, suggesting that virus can be expelled into the environment from either anatomical site. We also assessed the stability of two influenza A viruses (H1N1 and H3N2) in droplets of human saliva or respiratory mucus over a range of relative humidities. We observed that influenza virus infectivity decays rapidly in saliva droplets at intermediate relative humidity, while viruses in airway surface liquid droplets retain infectivity. Virus inactivation was not associated with bulk protein content, salt content, or droplet drying time. Instead, we found that saliva droplets exhibited distinct inactivation kinetics during the wet and dry phases at intermediate relative humidity and that droplet residue morphology may lead to the elevated first-order inactivation rate observed during the dry phase. Additionally, distinct differences in crystalline structure and nanobead localization were observed between saliva and airway surface liquid droplets. Together, our work demonstrates that different respiratory fluids exhibit unique virus persistence profiles and suggests that influenza viruses expelled from the oral cavity may contribute to virus transmission in low and high humidity environments.

Importance

Determining how viruses persist in the environment is important for mitigating transmission risk. Expelled infectious droplets and aerosols are composed of respiratory fluids, including saliva and complex mucus mixtures, but how influenza viruses survive in such fluids is largely unknown. Here, we find that infectious influenza virus is present in the oral cavity of infected ferrets, suggesting that saliva-containing expulsions can play a role in onward transmission. Additionally, influenza virus in droplets composed of saliva degrades more rapidly than virus within respiratory mucus. Droplet composition impacts the crystalline structure and virus localization in dried droplets. These results suggest that viruses from distinct sites in the respiratory tract could have variable persistence in the environment, which will impact viral transmission fitness.
The length of haplotype blocks and signals of structural variation in reconstructed genealogies
Authors: Ignatieva Anastasia , Favero Martina , Koskela Jere , Sant Jaromir , Myers Simon R.
Published: July 2023

Recent breakthroughs have enabled the accurate inference of large-scale genealogies. Through modelling the impact of recombination on the correlation structure between genealogical local trees, we evaluate how this structure is reconstructed by leading approaches. Despite identifying pervasive biases, we show that applying a simple correction recovers the desired distributions for one algorithm, Relate. We develop a statistical test to identify clades spanning unexpectedly long genomic regions, likely reflecting regional suppression of recombination in some individuals. Our approach allows a systematic scan for inter-individual recombination rate variation at an intermediate scale, between genome-wide differences and individual hotspots. Using genealogies reconstructed with Relate for 2 504 human genomes, we identify 50 regions possessing clades with unexpectedly long genomic spans (p <1·10−12). The strongest signal corresponds to a known inversion on chromosome 17. The second strongest uncovers a novel 760kb inversion on chromosome 10, common (21%) in S. Asians and correlated with GWAS hits for a range of phenotypes. Other regions indicate additional genomic rearrangements: inversions (8), copy number changes (2), or other variants (12). The remaining regions appear to reflect recombination suppression by previously unevidenced mechanisms. They are enriched for precisely spanning single genes (p= 5·10−10), specifically those expressed in male gametogenesis, and for eQTLs (p= 2·10−3). This suggests an extension of previously hypothesised crossover suppression within meiotic genes, towards a model of suppression varying across individuals with different expression levels. Our methods can be readily applied to other species, showing that genealogies offer previously un-tapped potential to study structural variation and other phenomena impacting evolution.
Efficient PCR-based gene targeting in isolates of the non-conventional yeastDebaryomyces hansenii
Authors: Alhajouj Sondos , Turkolmez Selva , Abalkhail Tarad , Alwan Zeena Hadi Obaid , Gilmour D. James , Mitchell Phil J. , Hettema Ewald H.
Published: July 2023

Debaryomyces hanseniiis a yeast with considerable biotechnological potential as an osmotolerant, stress tolerant oleaginous microbe. However, targeted genome modification tools are limited and require a strain with auxotrophic markers. Gene targeting by homologous recombination has been reported to be inefficient but here we describe a set of reagents and a method that allows gene targeting at high efficiency in wild type isolates. It uses a simple PCR-based amplification that extends a completely heterologous selectable marker with 50 bp flanks identical to the target site in the genome. Transformants integrate the PCR product through homologous recombination at high frequency (>75%). We illustrate the potential of this method by disrupting genes at high efficiency and by expressing a heterologous protein from a safe chromosomal harbour site. These methods should stimulate and facilitate further analysis ofD. hanseniistrains and open the way to engineer strains for biotechnology.
Improved Protocol for Reproducible Human Cortical Organoids Reveals Early Alterations in Metabolism withMAPTMutations
Authors: Bertucci Taylor , Bowles Kathryn R. , Lotz Steven , Qi Le , Stevens Katherine , Goderie Susan K. , Borden Susan , Oja Laura Maria , Lane Keith , Lotz Ryan , Lotz Hailey , Chowdhury Rebecca , Joy Shona , Arduini Brigitte L. , Butler David C. , Miller Michael , Baron Heide , Sandhof Carl Alexander , Silva M. Catarina , Haggarty Stephen J. , Karch Celeste M. , Geschwind Daniel H. , Goate Alison M. , Temple Sally
Published: July 2023

Cerebral cortical-enriched organoids derived from human pluripotent stem cells (hPSCs) are valuable models for studying neurodevelopment, disease mechanisms, and therapeutic development. However, recognized limitations include the high variability of organoids across hPSC donor lines and experimental replicates. We report a 96-slitwell method for efficient, scalable, reproducible cortical organoid production. When hPSCs were cultured with controlled-release FGF2 and an SB431542 concentration appropriate for theirTGFBR1/ALK5expression level, organoid cortical patterning and reproducibility were significantly improved. Well-patterned organoids included 16 neuronal and glial subtypes by single cell RNA sequencing (scRNA-seq), frequent neural progenitor rosettes and robust BCL11B+ and TBR1+ deep layer cortical neurons at 2 months by immunohistochemistry. In contrast, poorly-patterned organoids contain mesendoderm-related cells, identifiable by negative QC markers includingCOL1A2. Using this improved protocol, we demonstrate increased sensitivity to study the impact of differentMAPTmutations from patients with frontotemporal dementia (FTD), revealing early changes in key metabolic pathways.
Viral infection arrests coccolithophore calcification and nutrient consumption, and triggers shifts in organic stoichiometry
Authors: Dikstein Tamar , Antler Gilad , Pellerin Andre , Sharoni Shlomit , Frada Miguel J.
Published: July 2023

Blooms of the coccolithophoreEmiliania huxleyiare routinely infected by a specific lytic virus (EhV), which rapidly kills host cells triggering bloom termination and organic and inorganic carbon export. However, the impact of EhV on the dynamic of resource acquisition and cellular stoichiometry remains unknown, limiting the current understanding of the ecological and biogeochemical significance ofE. huxleyiblooms. To tackle this knowledge gap, we used algal and EhV cultures to determine over the course of infections the dynamics of alkalinity, modulated by calcification, nitrate and phosphate consumption and organic matter stoichiometry. We found that within 24hr alkalinity concentration stabilized and nutrient uptake declined to background levels. In parallel, the stoichiometric ratio of carbon to nitrogen was about 15% higher and the nitrogen to phosphorus ratio was about 12% lower during infections relative to controls. These variations likely resulted from lipid accumulation required for viral replication and the differential retention of phosphorus-rich macromolecular pools in decaying cells, respectively. Finally, after host population decay a progressive enrichment in phosphorus relative to nitrogen and carbon was detected in the remaining cell lysates. We estimate that this stoichiometric shift post-infection was driven by the progressive accumulation of heterotrophic bacteria involved in the degradation of organic material. Viral-mediated cell remodeling and consequent shifts in biomass stoichiometry likely impacts the patterns of nutrient cycling and biological carbon pump efficiency during large-scale blooms in the oceans.
The arginine/ornithine binding protein ArgT plays an essential role inBrucellato prevent intracellular killing and contribute to chronic persistence in the host
Authors: Mallik Sushree Rekha , Joshi Kiranmai , Radhakrishnan Girish K.
Published: July 2023

Brucellaspecies are facultative intracellular bacterial pathogens that cause the contagious zoonotic disease, brucellosis.Brucellaspp. infect a wide range of animals, including livestock, wild animals, and marine mammals. Brucellosis remains endemic to various parts of the world, affecting the economic growth of many countries because of its impact on public health and livestock productivity. There are no human vaccines for brucellosis, and controlling the disease in susceptible animals is crucial for limiting human infections. Although the available live-attenuated vaccines have protective efficacy in animals, they have many disadvantages, including infectivity in humans. Compared with other invasive bacterial pathogens, minimal information is available on the virulence factors ofBrucellathat enable them to survive in the host. Here, we performed transposon-based random mutagenesis ofB. neotomaeand identified the arginine/ornithine binding protein, ArgT, as the crucial virulence determinant ofBrucella. DeletingArgTfromB. melitensisresulted in its attenuation in macrophages, which was restored upon complementation with anArgTexpression plasmid. We observed that macrophages infected with ΔArgT-Brucellaproduced elevated levels of NO due to the inability of ΔArgT Brucellato deplete the host intracellular arginine through its importer. Furthermore, defective survival of ΔArgT B. melitensiswas observed in the infected mice, which correlated with enhanced NO production in the mice. Our studies revealed thatArgTinBrucellaplays a vital role in preventing intracellular killing and contributes to the chronic persistence ofBrucellain the host. This study highlights the essential role of arginine in clearing intracellular infections and the subversion of this host defense mechanism by intracellular pathogens for their chronic persistence.