bluuebunny/crossref_metadata_2025 huggingface dataset:
Spatial Multi-omics of Arterial Regions from Cardiac Allograft Vasculopathy Rejected Grafts
Authors: Nevarez-Mejia Jessica , Pickering Harry C. , Sosa Rebecca A. , Fishbein Gregory A. , Baldwin William M. , Fairchild Robert L. , Reed Elaine F.
Published: April 2023

2. Abstract

Background

Cardiac allograft vasculopathy (CAV) is a major cause of late-graft failure and mortality following heart transplantation. The mechanisms underlying vascular remodeling are poorly understood. A major immune risk factor associated with the development of CAV is the presence of donor-specific antibodies (DSA) that induce chronic endothelial cell (EC) injury, leukocyte recruitment and inflammation resulting in thickening of arterial intima. Here, we molecularly characterized innate and adaptive immune cells present in the arteries of rejected cardiac allografts with DSA and identified protein and transcriptomic signatures distinguishing early and late CAV lesions.

Methods

Arterial areas of interest (AOIs) from CAV+DSA+ rejected cardiac allografts (N=3; 2 females, 1 male) were subjected to GeoMx digital spatial profiling (DSP). AOIs were scored on the level of CAV progression/neointimal thickening (22 AOIs total; 11 high and 11 low neointima) and were subjected to whole transcriptome and protein profiling.

Results

AOIs with low neointima significantly increased markers for activated inflammatory infiltrates, transcripts of EC activation and gene modules involved in activating metalloproteinases and TP53 regulation of caspases. Inflammatory and apoptotic protein markers significantly correlated with inflammatory modules in AOIs with low neointima. AOIs with high neointima increased TGFβ-regulated transcripts and modules enriched for platelet activation/aggregation. Proteins encoding SMCs and growth factors/survival correlated with modules enriched for proliferation/repair in AOIs with high neointima. Key transcripts in promoting proliferation, migration, and EndoMT were significantly associated with increasing neointima scores.

Conclusion

Our results reveal new protein and transcriptomic signatures associated with CAV progression. Lesions exhibiting inflammatory profiles appear to be early lesions that transition to later proliferative/pro-fibrotic phenotype CAV lesions. These findings should form the foundation for the identification of improved biomarkers to guide CAV treatment.
A novel mouse allele of the DNA/RNA helicase senataxin (Setxspcar3) causing meiotic arrest of spermatocytes and male infertility
Authors: Fujiwara Yasuhiro , Saito Kouhei , Sun Fengyun , Petri Sabrina , Inoue Erina , Schimenti John , Okada Yuki , Handel Mary Ann
Published: April 2023

An unbiased screen for discovering novel genes for fertility identified thespcar3, spermatocyte arrest 3,mutant phenotype. Thespcar3mutation identified a new allele of theSetxgene, encoding senataxin, a DNA/RNA helicase that regulates transcription termination by resolving DNA/RNA hybrid R-loop structures. Although mutations in the humanSETXgene cause neural disorders,Setxspcar3mutant mice do not show any apparent neural phenotype, but instead exhibit male infertility and female subfertility. Histology of theSetxspcar3mutant testes revealed absence of spermatids and mature spermatozoa in the seminiferous tubules. Cytological analysis of chromosome spread preparations of theSetxspcar3mutant spermatocytes revealed normal synapsis, but aberrant DNA damage in the autosomes, and defective formation of the sex body. Furthermore,Setxspcar3testicular cells exhibited abnormal accumulation of R-loops compared to wild type testicular cells. Transient expression assays identified regions of the senataxin protein required for sub-nuclear localization. Together, these results not only confirm that senataxin is required for normal meiosis and spermatogenesis but also provide a new resource for determination of its role in maintaining R-loop formation and genome integrity.
Population ageing, incarceration and the growing digital divide: understanding the effects of digital literacy inequity experienced by older people leaving prison
Authors: Hwang Ye In (Jane) , Hagos Amanuel , Withall Adrienne , Hampton Stephen , Snoyman Phillip , Butler Tony
Published: April 2023

Background

Digital inequity refers to the inequality and exclusion experienced by those who lack the same opportunities or circumstances to support the development of digital skills as the rest of modern society. One rapidly growing and highly vulnerable group to digital inequity is older people attempting to reintegrate into society after release from prison, where technology access is limited. Inadequate support for digital skills in this population entails widespread consequences for public health, human rights, social welfare and recidivism. This qualitative study is the first to: examine digital inequity experienced by older people who have been incarcerated, understand the effects of this on reintegration to society, and begin informing appropriate solutions.

Method

Semi-structured interviews were conducted with N=15 older people (mean age= 57) who had been released from an Australian prison in the last two years, regarding their experiences of digital literacy since leaving prison. Reflexive thematic analysis was conducted under a critical realist lens.

Results

The analysis resulted in six themes that illustrated the extent of digital inequity experienced by this population, and key challenges for improving digital literacy: ‘surviving in a digital world’, ‘stranger in a foreign world’, ‘questioning the digital divide’, ‘overcoming your “old” self’, ‘don’t like what you don’t know’, and ‘seeking versus finding help’.

Conclusions

The digital inequity that older people experience during and after incarceration creates additional challenges for a growing group who are already medically and socially marginalised. Prioritisation of this group for digital literacy initiatives both during incarceration and in the community will have benefits for their health, social and financial reintegration. Their unique life experiences should be considered in designing and delivering these programs.
Co-immobilization of ciprofloxacin and chlorhexidine as a long-term, broad-spectrum antimicrobial dual-drug coating for polyvinyl chloride (PVC)-based endotracheal tubes
Authors: Alves Diana , Pereira Maria Olívia , Lopes Susana Patrícia
Published: April 2023

The endotracheal tube (ETT) affords support for intubated patients, but the rising incidence of ventilator-associated pneumonia (VAP) is jeopardizing its application. ETT surfaces promote (poly)microbial colonization and biofilm formation, with a heavy burden for VAP. Devising safe, broad-spectrum antimicrobial materials to tackle ETT bioburden is needful. Herein, we immobilized ciprofloxacin (CIP) and/or chlorhexidine (CHX), through polydopamine (pDA)-based functionalization, onto polyvinyl chloride (PVC) surfaces. These surfaces were characterized and challenged with single and polymicrobial cultures of VAP-relevant bacteria (Pseudomonas aeruginosa;Acinetobacter baumannii;Klebsiella pneumoniae;Staphylococcus aureus;Staphylococcus epidermidis) and fungi (Candida albicans). The coatings imparted PVC surfaces with homogeneous morphology, varied wettability, and low roughness. Coated surfaces exhibited sustained CIP/CHX release, retaining long-term (10 days) stability. Surfaces evidencing no A549 lung cell toxicity exhibited broad-spectrum anti-biofilm activity. CIP/CHX co-immobilization resulted in better outcomes than CIP or CHX coatings, reducing bacteria up to >7 Log10, and modestly distressing (ca. 1 Log10)C. albicans. The anti-biofilm effectiveness of coated surfaces endured for dual biofilms, substantially preventing bacterial populations and fungi (ca. 2.7 Log10) inP. aeruginosa/C. albicansconsortia. A less pronounced antifungal effect (ca. 1 Log10reduction) was found in triple-species communities, but fully preventingP. aeruginosaandS. aureuspopulations. CIP/CHX co-immobilization holds a safe and robust broad-spectrum antimicrobial coating for PVC-ETTs, with the promise laying in reducing VAP incidence.
Recruitment of PI4KIIIβ to the Golgi by ACBD3 is dependent on an upstream pathway of a SNARE complex and golgins
Authors: Stalder Danièle , Yakunin Igor , Gershlick David C.
Published: April 2023

ACBD3 is a protein localised to the Golgi apparatus and recruits other proteins, such as PI4KIIIβ, to the Golgi. However, the mechanism through which ACBD3 itself is recruited to the Golgi is poorly understood. This study demonstrates there are two mechanisms for ACBD3 recruitment to the Golgi. First, we identified that an MWT374-376motif in the unique region upstream of the GOLD domain in ACBD3 is essential for Golgi localisation. Second, we use unbiased proteomics to demonstrate that ACBD3 interacts with SCFD1, a Sec1/Munc-18 (SM) protein, and a SNARE protein, SEC22B. CRISPR-KO of SCFD1 causes ACBD3 to become cytosolic. We also found that ACBD3 is redundantly recruited to the Golgi apparatus by two golgins: golgin-45 and giantin, which bind to ACBD3 through interaction with the MWT374-376motif. Taken together, our results demonstrate that ACBD3 is recruited to the Golgi in a two-step sequential process, with the SCFD1-mediated interaction occurring upstream of the interaction with the golgins.
Genome editing in the mouse brain with minimally immunogenic Cas9 RNPs
Authors: Stahl Elizabeth C. , Sabo Jennifer K. , Kang Min Hyung , Allen Ryan , Applegate Elizabeth , Kim Shin Eui , Kwon Yoonjin , Seth Anmol , Lemus Nicholas , Salinas-Rios Viviana , Soczek Katarzyna , Trinidad Marena , Vo Linda T. , Jeans Chris , Wozniak Anna , Morris Timothy , Kimberlin Athen , Foti Thomas , Savage David F. , Doudna Jennifer A.
Published: April 2023

Transient delivery of CRISPR-Cas9 ribonucleoproteins (RNPs) into the central nervous system (CNS) for therapeutic genome editing could avoid limitations of viral vector-based delivery including cargo capacity, immunogenicity, and cost. Here we tested the ability of cell penetrant Cas9 RNPs to edit the mouse striatum when introduced using a convection enhanced delivery system. These transient Cas9 RNPs showed comparable editing of neurons and reduced adaptive immune responses relative to one formulation of Cas9 delivered using AAV serotype 9. The production of ultra-low-endotoxin Cas9 protein manufactured at scale further improved innate immunity. We conclude that injection-based delivery of minimally immunogenic CRISPR genome editing RNPs into the CNS provides a valuable alternative to virus-mediated genome editing.
Ensembles for improved detection of invasive breast cancer in histological images
Authors: Solorzano Leslie , Robertson Stephanie , Hartman Johan , Rantalainen Mattias
Published: April 2023

Accurate detection of invasive breast cancer (IC) can provide decision support to pathologists as well as improve downstream computational analyses, where detection of IC is a first step. Tissue containing IC is characterized by the presence of specific morphological features, which can be learned by convolutional neural networks (CNN). Here, we compare the use of a single CNN model versus an ensemble of several base models with the same CNN architecture, and we evaluate prediction performance as well as variability across ensemble based model predictions.

Two in-house datasets comprising 587 WSI are used to train an ensemble of ten InceptionV3 models whose consensus is used to determine the presence of IC. A novel visualization strategy was developed to communicate ensemble agreement spatially. Performance was evaluated in an internal test set with 118 WSIs, and in an additional external dataset (TCGA breast cancer) with 157 WSI.

We observed that the ensemble-based strategy outperformed the single CNN-model alternative with respect to accuracy on tile level in 89% of all WSIs in the test set. The overall accuracy was 0.92 (DICE coefficient, 0.90) for the ensemble model, and 0.85 (DICE coefficient, 0.83) for the single CNN alternative in the internal test set. For TCGA the ensemble outperformed the single CNN in 96.8% of the WSI, with an accuracy of 0.87 (DICE coefficient 0.89), the single model provides an accuracy of 0.75 (DICE coefficient 0.78)

The results suggest that an ensemble-based modeling strategy for breast cancer invasive cancer detection consistently outperforms the conventional single model alternative. Furthermore, visualization of the ensemble agreement and confusion areas provide direct visual interpretation of the results. High performing cancer detection can provide decision support in the routine pathology setting as well as facilitate downstream computational analyses.
Separate attentional processes in the two visual systems of jumping spiders
Authors: Ferrante Federico , Loconsole Maria , Giacomazzi Davide , De Agrò Massimo
Published: April 2023

1

By selectively focusing on a specific portion of the environment, animals can solve the problem of information overload, toning down irrelevant inputs and concentrate only on the relevant ones. This may be of particular relevance for animals such as the jumping spider, which possess a wide visual field of almost 360° and thus could benefit from a low-cost system for sharpening attention. Jumping spiders have a modular visual system composed of four pairs of eyes, of which only the two frontal eyes (i.e., AMEs) are motile, whereas the other secondary pairs remain immobile. We hypothesized that jumping spiders can exploit both primary and secondary eyes for stimulus detection and attentional shift, with the two systems working synergistically. In Experiment 1 we investigated AMEs’ attentional responses following a spatial cue presented to the secondary eyes. In Experiment 2, we tested for enhanced attention in the secondary eyes’ visual field congruent with the direction of the AMEs’ focus. In both experiments, we observed that animals were faster and more accurate in detecting a target when it appeared in a direction opposite to that of the initial cue. In contrast with our initial hypothesis, these results would suggest that attention is segregated across eyes, while each system works to compensate the other by attending to different spatial locations, rather than sharing a common attentional focus.
Development, Characterization, and Structural Analysis of a Genetically Encoded Red Fluorescent Peroxynitrite Biosensor
Authors: Pang Yu , Huang Mian , Fan Yichong , Yeh Hsien-Wei , Xiong Ying , Ng Ho Leung , Ai Hui-wang
Published: April 2023

Boronic acid-containing fluorescent molecules have been widely used to sense hydrogen peroxide and peroxynitrite, which are important reactive oxygen and nitrogen species in biological systems. However, it has been challenging to gain specificity. Our previous studies developed genetically encoded, green fluorescent peroxynitrite biosensors by genetically incorporating a boronic acid-containing noncanonical amino acid (ncAA),p-boronophenylalanine (pBoF), into the chromophore of circularly permuted green fluorescent proteins (cpGFPs). In this work, we introducedpBoF to amino acid residues spatially close to the chromophore of an enhanced circularly permuted red fluorescent protein (ecpApple). Our effort has resulted in two responsive ecpApple mutants: one bestows reactivity toward both peroxynitrite and hydrogen peroxide, while the other, namely pnRFP, is a selective red fluorescent peroxynitrite biosensor. We characterized pnRFPin vitroand in live mammalian cells. We further studied the structure and sensing mechanism of pnRFP using X-ray crystallography,11B-NMR, and computational methods. The boron atom in pnRFP adopts an sp2-hybridization geometry in a hydrophobic pocket, and the reaction of pnRFP with peroxynitrite generates a product with a twisted chromophore, corroborating the observed “turn-off” fluorescence response. Thus, this study extends the color palette of genetically encoded peroxynitrite biosensors, provides insight into the response mechanism of the new biosensor, and demonstrates the versatility of using protein scaffolds to modulate chemoreactivity.
Effects of B lymphocytes Invitro Treatment with Rose Extract on Transmembrane Ligands
Authors: Arokiaraj Mark Christopher , Menesson Eric
Published: April 2023

Aims

The study was performed to evaluate the role of red rose extract (Pierre de Ronsard) on B lymphocytes. Specifically the gene expression of CD20, CD30, CD40, and CCR5 in human B cells were studied after treatment with rose extract.

Methods

Red rose extract was prepared at the dilution of 0.0075% (v/v) and stored until use at -20°C. Cell treatment was performed at 37°C on B cells. The cells were plated in 6 well plates at 1.5×106cells per well and stored at -80°C. Total RNA extraction and quality control were performed. RTq-PCR was performed according to Genecopoeia’s instructions. The cycle threshold method (ΔΔCt) was used for data analysis.

Results

The comparative Ct method quantification (2^-ΔCt) and fold change for CD20, CD 30, CD 40 and CCR5 were - 5.65E+01, 4.80E-01, N/A, 2.47E-01; and 0.954,0.377, N/A and 0.577, respectively. The amount of total RNA extracted from about 4.5×106cells was low and did not allow us to measure the RNA profile. The A260/A230 ratios were very low due to the low amount of RNA. The analysis of gene expression by qRT-PCR showed that CD40 was not expressed in untreated cells and cells treated with rose extract with the Ct values over 32. All other genes were expressed and well-measured in both B cell samples.

Conclusion

The treatment with rose extract at 0.0075% (v/v) did not modify the expression of CD20. However, the expression of CD30 and CCR5 decreased with the rose extract treatment. The range of the fold change showed that the result of CD30 expression was more accurate than for CCR5.
Loss of the extracellular matrix protein Perlecan disrupts axonal and synaptic stability duringDrosophiladevelopment
Authors: Guss Ellen J. , Akbergenova Yulia , Cunningham Karen L. , Littleton J. Troy
Published: April 2023

Heparan sulfate proteoglycans (HSPGs) form essential components of the extracellular matrix (ECM) and basement membrane (BM) and have both structural and signaling roles. Perlecan is a secreted ECM-localized HSPG that contributes to tissue integrity and cell-cell communication. Although a core component of the ECM, the role of Perlecan in neuronal structure and function is less understood. Here we identify a role forDrosophilaPerlecan in the maintenance of larval motoneuron axonal and synaptic stability. Loss of Perlecan causes alterations in the axonal cytoskeleton, followed by axonal breakage and synaptic retraction of neuromuscular junctions. These phenotypes are not prevented by blocking Wallerian degeneration and are independent of Perlecan’s role in Wingless signaling. Expression of Perlecan solely in motoneurons cannot rescue synaptic retraction phenotypes. Similarly, removing Perlecan specifically from neurons, glia or muscle does not cause synaptic retraction, indicating the protein is secreted from multiple cell types and functions non-cell autonomously. Within the peripheral nervous system, Perlecan predominantly localizes to the neural lamella, a specialized ECM surrounding nerve bundles. Indeed, the neural lamella is disrupted in the absence of Perlecan, with axons occasionally exiting their usual boundary in the nerve bundle. In addition, entire nerve bundles degenerate in a temporally coordinated manner across individual hemi-segments throughout larval development. These observations indicate disruption of neural lamella ECM function triggers axonal destabilization and synaptic retraction of motoneurons, revealing a role for Perlecan in axonal and synaptic integrity during nervous system development.
Dual functioning by the PhoR sensor is a key determinant toMycobacterium tuberculosisvirulence
Authors: Singh Prabhat Ranjan , Goar Harsh , Paul Partha , Mehta Khushboo , Bamniya Bhanwar , Vijjamarri Anil Kumar , Bansal Roohi , Khan Hina , Karthikeyan Subramanian , Sarkar Dibyendu
Published: April 2023

PhoP-PhoR empowersM. tuberculosisto adapt to diverse environmental conditions, and remains essential for virulence. Although PhoP and PhoR have been structurally characterized, the signal(s) that this TCS responds to remains unknown. In this study, we show that PhoR is a sensor of acidic pH/high salt conditions, which activate PhoP via phosphorylation. Transcriptomic studies uncover that acidic pH-inducible expression of PhoP regulon is significantly inhibited in a PhoR-deletedM. tuberculosis. Using genome-wide screening we further identify a non-canonical mechanism of PhoP phosphorylation by the sensor kinase PrrB. To investigate how phosphorylation of PhoP is regulated, we discovered that PhoR functions as a phosphatase. Our results identify the motif/residues responsible for contrasting kinase/phosphatase dual functioning of PhoP, and collectively determine the homeostatic regulation of intra-mycobacterial P~PhoP which controls the final output of PhoP regulon. Together, these data uncover that PhoR plays a central role in mycobacterial adaptation to low pH conditions within the host macrophage phagosome. Consistent with these results a PhoR-deletedM. tuberculosisremains significantly attenuated in macrophages and animal models.
How reliable are DAO supplements? — A comparison of over-the-counter Diamine oxidase products
Authors: Alemany-Fornes Marc , Bori Jaume , Tintoré Maria , Cuñé Jordi , de Lecea Carlos
Published: April 2023

Diamine oxidase (DAO) supplements have gained increasing attention in recent years due to their potential to support DAO deficiency, histamine intolerance and related symptoms. However, choosing a reliable and trustworthy DAO supplement can be a challenging task for patients and Healthcare practitioners. One of the main concerns is the lack of regulatory oversight on dietary supplements, which may result in misleading or incomplete labelling, incorrect dosages, and inadequate quality control. Such situations may lead to patients consuming supplements with insufficient amounts of active enzyme or with questionable purity, potentially resulting in undesired health outcomes. Thus, we tested the DAO activity of a variety of already in-the-market supplements and compared them with one another and against the specified activity, if any, by the manufacturer. Our results show a great discrepancy in most of the products and significant differences in DAO activity between different manufacturers.
Do NSm virulence factors in the Bunyavirales viral order originate from Gn gene duplication?
Authors: Lefebvre Victor , Foun Lin Ravy Leon , Cole Laura , Cosset François-Loïc , Fogeron Marie-Laure , Böckmann Anja
Published: April 2023

One-third of the nine WHO shortlisted pathogens prioritized for research and development in public health emergencies belong to theBunyaviralesorder. SeveralBunyaviralesspecies carry an NSm protein that acts as a virulence factor. We predicted the structures of these NSm protein and unexpectedly found that in two families, its cytosolic domain is inferred to have a similar fold to the cytosolic domain of the viral envelope-forming glycoprotein N (Gncyto) encoded on the same genome fragment. We show that although the sequence identity between the NSmcytoand Gncytodomains is low, the conservation of the two zinc finger-forming CCHC motifs explains the predicted structural conservation. Importantly, our predictions provide a first glimpse into the long unknown structure of NSm and its link to virulence. Also, these predictions suggest that NSm is the result of a gene duplication event in theBunyavirales NairoviridaeandPeribunyaviridaefamilies, and that such events may be common in the recent evolutionary history of RNA viruses.
A revamped rat reference genome improves the discovery of genetic diversity in laboratory rats
Authors: de Jong Tristan V , Pan Yanchao , Rastas Pasi , Munro Daniel , Tutaj Monika , Akil Huda , Benner Chris , Chen Denghui , Chitre Apurva S , Chow William , Colonna Vincenza , Dalgard Clifton L , Demos Wendy M , Doris Peter A , Garrison Erik , Geurts Aron M , Gunturkun Hakan M , Guryev Victor , Hourlier Thibaut , Howe Kerstin , Huang Jun , Kalbfleisch Ted , Kim Panjun , Li Ling , Mahaffey Spencer , Martin Fergal J , Mohammadi Pejman , Ozel Ayse Bilge , Polesskaya Oksana , Pravenec Michal , Prins Pjotr , Sebat Jonathan , Smith Jennifer R , Woods Leah C Solberg , Tabakoff Boris , Tracey Alan , Uliano-Silva Marcela , Villani Flavia , Wang Hongyang , Sharp Burt M , Telese Francesca , Jiang Zhihua , Saba Laura , Wang Xusheng , Murphy Terence D , Palmer Abraham A , Kwitek Anne E , Dwinell Melinda R , Williams Robert W , Li Jun Z , Chen Hao
Published: April 2023

The seventh iteration of the reference genome assembly forRattus norvegicus— mRatBN7.2—corrects numerous misplaced segments and reduces base-level errors by approximately 9-fold and increases contiguity by 290-fold compared to its predecessor. Gene annotations are now more complete, significantly improving the mapping precision of genomic, transcriptomic, and proteomics data sets. We jointly analyzed 163 short-read whole genome sequencing datasets representing 120 laboratory rat strains and substrains using mRatBN7.2. We defined ∼20.0 million sequence variations, of which 18.7 thousand are predicted to potentially impact the function of 6,677 genes. We also generated a new rat genetic map from 1,893 heterogeneous stock rats and annotated transcription start sites and alternative polyadenylation sites. The mRatBN7.2 assembly, along with the extensive analysis of genomic variations among rat strains, enhances our understanding of the rat genome, providing researchers with an expanded resource for studies involving rats.
On-chip brain slice stimulation: precise control of electric fields and tissue orientation
Authors: Shaner Sebastian , Lu Han , Lenz Maximilian , Garg Shreyash , Vlachos Andreas , Asplund Maria
Published: April 2023

Non-invasive brain stimulation modalities, including transcranial direct current stimulation (tDCS), are widely used in neuroscience and clinical practice to modulate brain function and treat neuropsychiatric diseases. DC stimulation ofex vivobrain tissue slices has been a method used to understand mechanisms imparted by tDCS. However, delivering spatiotemporally uniform direct current electric fields (dcEFs) that have precisely engineered magnitudes and are also exempt from toxic electrochemical by-products are both significant limitations in conventional experimental setups. As a consequence, bioelectronic dose-response interrelations, the role of EF orientation, and the biomechanisms of prolonged or repeated stimulation over several days all remain not well understood. Here we developed a platform with fluidic, electrochemical, and magnetically-induced spatial control. Fluidically, the chamber geometrically confines precise dcEF delivery to the enclosed brain slice and allows for tissue recovery in order to monitor post-stimulation effects. Electrochemically, conducting hydrogel electrodes mitigate stimulation-induced faradaic reactions typical of commonly-used metal electrodes. Magnetically, we applied ferromagnetic substrates beneath the tissue and used an external permanent magnet to enablein siturotational control in relation to the dcEF. By combining the microfluidic chamber with live-cell calcium imaging and electrophysiological recordings, we showcased the potential to study the acute and lasting effects of dcEFs with the potential of providing multi-session stimulation. This on-chip bioelectronic platform presents a modernized yet simple solution to electrically stimulate explanted tissue by offering more environmental control to users, which unlocks new opportunities to conduct thorough brain stimulation mechanistic investigations.

Graphical abstract
Hippocampus and Striatum Showed Distinct Contributions to Longitudinal Changes in Value-Based Learning in Middle Childhood
Authors: Falck Johannes , Zhang Lei , Raffington Laurel , Mohn Johannes J. , Triesch Jochen , Heim Christine , Shing Yee Lee
Published: April 2023

The hippocampal-dependent memory system and striatal-dependent memory system modulate reinforcement learning depending on feedback timing in adults, but their contributions during development remain unclear. In a 2-year longitudinal study, 6-to-7-year-old children performed a reinforcement learning task in which they received feedback immediately or with a short delay following their response. Children’s learning was found to be sensitive to feedback timing modulations in their reaction time and inverse temperature parameter, which quantifies value-guided decision-making. They showed longitudinal improvements towards more optimal value-based learning, and their hippocampal volume showed protracted maturation. Better delayed model-derived learning covaried with larger hippocampal volume longitudinally, in line with the adult literature. In contrast, a larger striatal volume in children was associated with both better immediate and delayed model-derived learning longitudinally. These findings show, for the first time, an early hippocampal contribution to the dynamic development of reinforcement learning in middle childhood, with neurally less differentiated and more cooperative memory systems than in adults.
NAD+metabolism is a key modulator of bacterial respiratory epithelial infections
Authors: Klabunde Björn , Wesener André , Bertrams Wilhelm , Beinborn Isabell , Paczia Nicole , Surmann Kristin , Blankenburg Sascha , Wilhelm Jochen , Serrania Javier , Knoops Kèvin , Elsayed Eslam M. , Laakmann Katrin , Jung Anna Lena , Kirschbaum Andreas , Mraheil Mobarak Abu , Becker Anke , Völker Uwe , Vollmeister Evelyn , Benedikter Birke J. , Schmeck Bernd
Published: April 2023

1.

Lower respiratory tract infections caused byStreptococcusOpneumoniae (Spn)are a leading cause of death globally. Here we investigate the bronchial epithelial response toSpninfection on a transcriptomic, proteomic and metabolic level. We found the NAD+salvage pathway to be dysregulated upon infection in a cell line model, primary human lung tissue andin vivoin rodents, leading to a reduced production of NAD+. Knockdown of NAD+salvage enzymes (NAMPT, NMNAT1) increased bacterial replication. NAD+treatment ofSpninhibited its growth while growth of other respiratory pathogens improved. Boosting NAD+production increased NAD+levels in immortalized and primary cells and decreased bacterial replication upon infection. NAD+treatment ofSpndysregulated the bacterial metabolism and reduced intrabacterial ATP. Enhancing the bacterial ATP metabolism abolished the antibacterial effect of NAD+. Thus, we identified the NAD+salvage pathway as an antibacterial cascade inSpninfections, predicting a novel antibacterial mechanism of NAD+.
Validation of Tau Antibodies for Use in Western Blotting and Immunohistochemistry
Authors: Ellis Michael J. , Lekka Christiana , Tulmin Hanna , O’Brien Darragh P. , Dhayal Shalinee , Zeissler Marie-Louise , Knudsen Jakob G. , Kessler Benedikt M. , Morgan Noel G. , Todd John A. , Richardson Sarah J. , Stefana M. Irina
Published: April 2023

Background

The microtubule-associated protein Tau has attracted diverse and increasing research interest, with Tau being mentioned in the title/abstract of nearly 34,000 PubMed-indexed publications to date. To accelerate studies into Tau biology, the characterisation of its multiple proteoforms, including disease-relevant post-translational modifications (PTMs), and its role in neurodegeneration, a multitude of Tau-targeting antibodies have been developed, with hundreds of distinct antibody clones currently available for purchase. Nonetheless, concerns over antibody specificity and limited understanding of the performance of many of these reagents has hindered research.

Methods

We have employed a range of techniques in combination with samples of murine and human origin to characterise the performance and specificity of 53 commercially-available Tau antibodies by Western blot, and a subset of these, 35 antibodies, in immunohistochemistry.

Results

Continued expression of residual protein was found in presumptive Tau “knockout” human cells and further confirmed through mass-spectrometry proteomics, providing evidence of Tau isoforms generated by exon skipping. Importantly, many total and isoform-specific antibodies failed to detect this residual Tau, as well as Tau expressed at low, endogenous levels, thus highlighting the importance of antibody choice. Our data further reveal that the binding of several “total” Tau antibodies, which are assumed to detect Tau independently of post-translational modifications, was partially inhibited by phosphorylation. Many antibodies also displayed non-specific cross-reactivity, with some total and phospho-Tau antibodies cross-reacting with MAP2 isoforms, while the “oligomer-specific” T22 antibody detected monomeric Tau on Western blot. Regardless of their specificity, with one exception, the phospho-Tau antibodies tested were found to not detect the unphosphorylated protein.

Conclusions

We identify Tau antibodies across all categories (total, PTM-dependent and isoform-specific) that can be employed in Western blot and/or immunohistochemistry applications to reliably detect even low levels of Tau expression with high specificity. This is of particular importance for studying Tau in non-neuronal cells and peripheral tissues, as well as for the confident validation of knockout cells and/or animal models. This work represents an extensive resource that serves as a point of reference for future studies. Our findings may also aid in the re-interpretation of existing data and improve reproducibility of Tau research.
A spatial map of hepatic mitochondria uncovers functional heterogeneity shaped by nutrient-sensing signaling
Authors: Kang Sun Woo Sophie , Cunningham Rory P. , Miller Colin B. , Brown Lauryn A. , Cultraro Constance M. , Harned Adam , Narayan Kedar , Hernandez Jonathan , Jenkins Lisa M. , Lobanov Alexei , Cam Maggie , Porat-Shliom Natalie
Published: April 2023

In the liver, mitochondria are exposed to different concentrations of nutrients due to their spatial positioning across the periportal (PP) and pericentral (PC) axis. How these mitochondria sense and integrate these signals to respond and maintain homeostasis is not known. Here, we combined intravital microscopy, spatial proteomics, and functional assessment to investigate mitochondrial heterogeneity in the context of liver zonation. We found that PP and PC mitochondria are morphologically and functionally distinct; beta-oxidation was elevated in PP regions, while lipid synthesis was predominant in the PC mitochondria. In addition, comparative phosphoproteomics revealed spatially distinct patterns of mitochondrial composition and potential regulation via phosphorylation. Acute pharmacological modulation of nutrient sensing through AMPK and mTOR shifted mitochondrial phenotypes in the PP and PC regions, linking nutrient gradients across the lobule and mitochondrial heterogeneity. This study highlights the role of protein phosphorylation in mitochondrial structure, function, and overall homeostasis in hepatic metabolic zonation. These findings have important implications for liver physiology and disease.
Elevated temperature alters microbial communities, but not decomposition rates, during three years of in-situ peat decomposition
Authors: Roth Spencer , Griffiths Natalie A. , Kolka Randall K. , Oleheiser Keith C. , Carrell Alyssa A. , Klingeman Dawn M. , Seibert Angela , Chanton Jeffrey P. , Hanson Paul J. , Schadt Christopher W.
Published: April 2023

Peatlands store approximately one-third of the global terrestrial carbon and are historically considered carbon sinks due to primary production outpacing microbial decomposition of organic matter. Climate change has the potential to alter the rate at which peatlands store or release carbon, and results from the Spruce and Peatland Responses Under Changing Environments (SPRUCE) experiment have shown net losses of organic matter and increased greenhouse gas production from a boreal peatland in response to whole-ecosystem warming. In this study, we utilized the SPRUCE sites to investigate how warming and elevated CO2impact peat microbial communities and peat soil decomposition. We deployed peat soil decomposition ladders across warming and CO2treatment enclosures for three years, after which we characterized bacterial, archaeal, and fungal communities through amplicon sequencing and measured peat mass and compositional changes across four depth increments. Microbial diversity and community composition were significantly impacted by soil depth, temperature, and CO2treatment. Bacterial/archaeal α-diversity increased significantly with increasing temperature, and fungal α-diversity was significantly lower under elevated CO2treatment. Trans-domain microbial networks showed higher complexity (nodes, edges, degree, betweenness centrality) of microbial communities in decomposition ladders from warmed enclosures, and the number of highly connected, hub taxa within the networks was positively correlated with temperature. Methanogenic hubs were identified in the networks constructed from the warmest enclosures, indicating increased importance of methanogenesis in response to warming. Microbial community responses were not however reflected in measures of peat soil decomposition, as warming and elevated CO2had no significant short-term effects on soil mass loss or composition. Regardless of treatment, on average only 4.5% of the original soil mass was lost after three years and variation between replicates was high, potentially masking treatment effects. Many previous studies from the SPRUCE experiment have shown that warming is accelerating organic-matter decomposition and CO2and CH4production, and our results suggest that these changes may be driven by warming-induced shifts in microbial communities.
Four additional natural 7-deazaguanine derivatives in phages and how to make them
Authors: Cui Liang , Balalkundu Seetharamsingh , Liu Chuanfa , Ye Hong , Hourihan Jacob , Rausch Astrid , Hauß Christopher , Nilsson Emelie , Hoetzinger Matthias , Holmfeldt Karin , Zhang Weijia , Martinez-Alvarez Laura , Peng Xu , Tremblay Denise , Moinau Sylvain , Solonenko Natalie , Sullivan Matthew B. , Lee Yan-Jiun , Mulholland Andrew , Weigele Peter , de Crécy-Lagard Valérie , Dedon Peter C. , Hutinet Geoffrey
Published: April 2023

Bacteriophages and bacteria are engaged in a constant arms race, continually evolving new molecular tools to survive one another. To protect their genomic DNA from restriction enzymes, the most common bacterial defence systems, double-stranded DNA phages have evolved complex modifications that affect all four bases. This study focuses on modifications at position 7 of guanines. Eight derivatives of 7-deazaguanines were identified, including four previously unknown ones: 2’-deoxy-7-(methylamino)methyl-7-deazaguanine (mdPreQ1), 2’-deoxy-7-(formylamino)methyl-7-deazaguanine (fdPreQ1), 2’-deoxy-7-deazaguanine (dDG), and 2’-deoxy-7-carboxy-7-deazaguanine (dCDG). These modifications are inserted in DNA by a guanine transglycosylase named DpdA. Three subfamilies of DpdA had been previously characterized: bDpdA, DpdA1, and DpdA2. Two additional subfamilies were identified in this work: DpdA3, which allows for complete replacement of the guanines, and DpdA4, which is specific to archaeal viruses. Transglycosylases have now been identified in all phages and viruses carrying 7-deazaguanine modifications, indicating that the insertion of these modifications is a post-replication event. Three enzymes were predicted to be involved in the biosynthesis of these newly identified DNA modifications: 7-carboxy-7-deazaguanine decarboxylase (DpdL), dPreQ1formyltransferase (DpdN), and dPreQ1methyltransferase (DpdM), which was experimentally validated and harbors a unique fold not previously observed for nucleic acid methylases.
Disturbance increases functional diversity but decreases phylogenetic diversity of an arboreal tropical ant community
Authors: Hoenle Philipp O. , Plowman Nichola S. , Matos-Maraví Pável , Bello Francesco de , Bishop Tom R. , Libra Martin , Idigel Cliffson , Rimandai Maling , Klimes Petr
Published: April 2023

Tropical rainforest canopies host a highly diverse arthropod fauna, which contribute to ecosystem function through their functional (FD) and phylogenetic diversity (PD). While a lot of previous research has documented the severe negative impacts of disturbance on the FD and PD of ground invertebrate communities, our understanding of arboreal counterparts is limited.

Here, we studied the effects of forest disturbance on an ecologically important invertebrate group, the ants, in a lowland rainforest in New Guinea. We exhaustively sampled 4000 m2area of a primary and a secondary forest for canopy ants. We report > 2800 occurrences of 128 ant species in 852 trees, one of the most comprehensive arboreal collections to date.

To test how ant PD and FD differ between the two forests, we constructed the ant species-level community phylogeny and measured 10 functional traits. Furthermore, we assessed by data exclusion the influence of species which were not nesting in individual trees (visitors) or only nesting (nesters), and of non-native species on FD and PD values. We expected that disturbance would decrease FD and PD in tree dwelling ants. We hypothesized that traits in primary forests would be more overdispersed due to the greater availability of ecological niches, while secondary forests would have stronger trait clustering due to a a stronger habitat filtering caused by more extreme microclimate.

Primary forests had higher species richness and PD than secondary forest. Surprisingly, we found higher FD in secondary forest. This pattern was robust even if we decoupled functional and phylogenetic signals or if non-native ant species were excluded from the data. Visitors did not contribute strongly to FD, but they increased PD. Community trait means further corroborate the functional distinctiveness of arboreal ants among secondary and primary forest, with almost all traits being impacted by disturbance and forest succession.

We conclude that the most plausible explanation is increased competition among closely related ant species in the secondary forest, which drives trait divergence. In the primary forest, abiotic habitat filters leads to more similar morphology and thus lower FD of phylogenetically more diverse ant assemblages.
CRISPR/Cas9 screen identifies KRAS-induced COX-2 as a driver of immunotherapy resistance in lung cancer
Authors: Boumelha Jesse , de Castro Andrea , Bah Nourdine , Cha Hongui , de Carné Trécesson Sophie , Rana Sareena , Anastasiou Panayiotis , Mugarza Edurne , Moore Christopher , Goldstone Robert , East Phil , Litchfield Kevin , Lee Se-Hoon , Molina-Arcas Miriam , Downward Julian
Published: April 2023

Oncogenic KRAS impairs anti-tumour immune responses, but effective strategies to combine KRAS inhibitors and immunotherapies have so far proven elusive. In vivo CRISPR-Cas9 screening in an immunogenic murine lung cancer model identifies mechanisms by which oncogenic KRAS promotes immune evasion, most notably expression of immunosuppressive cyclooxygenase-2 (COX-2) in cancer cells. Oncogenic KRAS was a potent inducer of COX-2 in both mouse and human lung cancer which was suppressed using KRAS inhibitors. COX-2 acting via prostaglandin E2 (PGE2) promotes resistance to immune checkpoint blockade (ICB) in both mouse and human lung adenocarcinoma. Targeting COX-2/PGE2 remodelled the tumour microenvironment by inducing pro-inflammatory polarisation of myeloid cells and influx of activated cytotoxic CD8+ T cells, which increased the efficacy of ICB. Restoration of COX-2 expression contributed to tumour relapse after prolonged KRAS inhibition. We propose testing COX-2/PGE2 pathway inhibitors in combination with KRAS G12C inhibition or ICB in patients with KRAS-mutant lung cancer.
Colorectal cancer as a model for biological evolution
Authors: Ledda Marco , Pluchino Alessandro , Ragusa Marco
Published: April 2023

Complexity in cancer research has led to the creation of powerful analytical tools for helping experimentalin vivoandin vitromethods. These tools range from systems of differential equations resolved in computer simulations, to lattice models and agent-based models (ABMs). These analytical methods are focused on studying cell behavior and dynamic cell populations. Among these, those that are increasingly used are ABMs because they can incorporate multi-scale features ranging from the individual up to the population level, mixing rules based on mathematical and conceptual parameters, and combining statistical/population assumptions with individual heterogeneity. In this work, we present an ABM that simulates tumor progression in a colonic crypt, with the aim of providing an experimentalin silicoenvironment for testing results achieved in traditional lab research, and developing alternative scenarios of tumor development. As the first part of an ongoing project, the long-term goal is to reproduce and study, the general evolutionary mechanisms of a biological system.
Different behavioural profiles between invasive and native nudibranchs: means for invasion success?
Authors: Macali Armando , Ferretti Sara , Scozzafava Serena , Carere Claudio
Published: April 2023

Behaviour is predicted to be a primary determinant of the success of the invasion process during the early phases of colonization. Comparing sympatric invaders and native species may provide a good approach to unravel behavioural traits involved in an invasion process. In this study, we carried out an experimental simulation of the introduction and the establishment phase into a new environment and assessed the expression of activity, alertness and habituation in a non-indigenous Mediterranean population of the South African nudibranchGodiva quadricolorcomparing its profiles with those of the sympatric nativeCratena peregrinaandCaloria quatrefagesi. Individuals of these three species were subjected to three behavioural tests: spontaneous activity, carried out in the introduction phase (immediately after sampling) and after a week of acclimatization; alert test, in which a potential threat was simulated by means of a tactile stimulus; habituation test, where the same alert test stimulus was repeated five times at thirty- minute intervals. Native nudibranch had repeatable traits, although with species differences perhaps related to their different ecological niches. The comparison with the invasive species highlighted its low repeatability in activity levels, suggesting higher plasticity, a strong tendency to locomotor activity, and a marked sensitization in the habituation test. Such traits could play an important and active role in the ongoing invasion process.