bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of Gaze, Vision, and Memory on Hand Kinematics in a Placement Task
Authors: Luabeya Gaelle N. , Yan Xiaogang , Freud Erez , Crawford J. Douglas
Published: September 2023

People usually reach for objects toplacethem in some position and orientation, but the placement component of this sequence is often ignored. For example, reaches are influenced by gaze position, visual feedback, and memory delays, but their influence on object placement is unclear. Here, we tested these factors in a task where participants placed and oriented a trapezoidal block against 2D visual templates displayed on a frontally located computer screen. In Experiment 1, participants matched the block to three possible orientations: 0° (horizontal), +45° and −45°, with gaze fixated 10° to the left/right. The hand and template either remained illuminated (closed-loop), or visual feedback was removed (open-loop). In Experiment 2, a memory delay was added, and participants sometimes performed saccades (centripetal, centrifugal, or opposite side). In Experiment 1, the hand consistently overshot the template relative to gaze (similar to reaching), especially in the open-loop task. After a memory delay, location errors were influenced by both template orientation and gaze position. Based on previous reach experiments, we expected these errors to be independent of the previous eye position, but placement overshoot also depended on previous saccade metrics. Hand orientation over-rotated relative to template orientation (all conditions). Orientation was influenced by gaze position in Experiment 1, but this vanished in the presence of a memory delay. These results demonstrate interactions between gaze, location, and orientation signals in the planning and execution of hand placement and suggest different neural mechanisms for closed-loop, open-loop, and memory delay placement.

NEW & NOTEWORTHY

Eye-hand coordination studies usually focus on object acquisition, but placement is equally important. Here, we investigated how gaze position influences object placement toward a 2D template, with different levels of visual feedback. Like reach, placement overestimated goal location relative to gaze, but was also influenced by previous saccade metrics. Gaze also modulated hand orientation, which generally overestimated template orientation. Gaze influence was feedback-dependent, with location errors increasing but orientation errors decreasing after a memory delay.
Multifactorial approach is needed to unravel the maturation phases of human neurons derived from induced pluripotent stem cells
Authors: Ben Mahmoud Maissa , Rátkai Anikó , Bauer Krisztina , Bencsik Norbert , Szücs Attila , Schlett Katalin , Tárnok Krisztián
Published: September 2023

Neurons derived from induced pluripotent stem cells (h-iPSC-Ns) provide an invaluable model for studying the physiological aspects of neuronal development and diseases. However, multiple studies have also demonstrated that h-iPSC-Ns exhibit a high degree of functional and epigenetic diversity. Due to the imprecise characterization and significant variation among the currently available maturation protocols, it is essential to establish a set of criteria to standardize models and accurately characterize and define the developmental properties of neurons derived from iPSCs.

In this study, we conducted a comprehensive analysis of the h-iPSC-Ns via electrophysiological and microscopic techniques to follow their functional development at the cellular and network levels. This enabled us to provide a thorough description of the maturation process of h-iPSC-Ns over a 10-week periodin vitro. Specifically, we have used conventional whole-cell patch-clamp and dynamic clamp techniques, alongside morphometry, to assess the characteristics of maturing h-iPSC-Ns. Additionally, we utilized calcium imaging to monitor the progression of synaptic activity and network communication. At the single cell level, human neurons exhibited gradually decreasing membrane resistance in parallel with improved excitability by 5 weeks of maturation. Their firing profiles were consistent with those of mature regular firing type of neurons. At the network level we observed the development of abundant fast glutamatergic and depolarizing GABAergic synaptic connections together with synchronized network activity. The identified sequence of differentiation events are consistent and offers a robust framework for developing targeted experiments at varying stages of neuronal maturation. This framework allows for the use of different, age-related methodologies or a singular set of experiments for a culture’s maturation.
Mosquitoes do not like bitter: new perspectives for biorational repellents
Authors: Lazzari Claudio R. , Ortega-Insaurralde Isabel , Esnault Jérémy , Costa Eloïse , Crespo José E. , Barrozo Romina B.
Published: September 2023

Chemical repellents play a crucial role in personal protection, serving as essential elements in reducing the transmission of vector-borne diseases. A biorational perspective that extends beyond the olfactory system as the classical target may be a promising direction to move. The taste system provides reliable information regarding food quality, helping animals discriminate between nutritious and potentially harmful food sources, often associated with a bitter taste. Understanding how bitter compounds affect feeding in blood-sucking insects could unveil novel molecules with the potential to reduce biting and feeding. Here, we investigated the impact of two naturally occurring bitter compounds, caffeine and quinine, on the feeding decisions in femaleAedes aegyptimosquitoes at two distinctive phases: (1) when the mosquito explores the biting substrate using external taste sensors and (2) when the mosquito takes a sip of food and tastes it using internal taste receptors. We assessed the aversiveness of bitter compounds through both an artificial feeding condition (artificial feeder test) and a real host (arm-in-cage test). Our findings revealed different sensitivities in the external and internal sensory pathways responsible for detecting bitter taste inAe. aegypti. Internal detectors exhibited responsiveness to lower doses compared to the external sensors. Quinine exerted a more pronounced negative impact on biting and feeding activity than caffeine. The implications of our findings are discussed in the context of mosquito food recognition and the potential practical implications for personal protection.
KSHV 3.0: A State-of-the-Art Annotation of the Kaposi’s Sarcoma-Associated Herpesvirus Transcriptome Using Cross-Platform Sequencing
Authors: Prazsák István , Tombácz Dóra , Fülöp Ádám , Torma Gábor , Gulyás Gábor , Dörmő Ákos , Kakuk Balázs , Spires Lauren McKenzie , Toth Zsolt , Boldogkői Zsolt
Published: September 2023

Kaposi’s sarcoma-associated herpesvirus (KSHV) is a large, oncogenic DNA virus belonging to the gammaherpesvirus subfamily. KSHV has been extensively studied with various high-throughput RNA-sequencing approaches to map the transcription start and end sites, the splice junctions, and the translation initiation sites. Despite these efforts, the comprehensive annotation of the viral transcriptome remains incomplete. In the present study, we generated a long-read sequencing dataset of the lytic and latent KSHV transcriptome using native RNA and direct cDNA sequencing methods. This was supplemented with CAGE sequencing based on a short-read platform. We also utilized datasets from previous publications for our analysis. As a result of this combined approach, we have identified a number of novel viral transcripts and RNA isoforms and have either corroborated or improved the annotation of previously identified viral RNA molecules, thereby notably enhancing our comprehension of the transcriptomic architecture of the KSHV genome. We also evaluated the coding capability of transcripts previously thought to be non-coding, by integrating our data on the viral transcripts with translatomic information from other publications.

IMPORTANCE

Deciphering the viral transcriptome of KSHV is of great importance because we can gain insight into the molecular mechanism of viral replication and pathogenesis, which can help develop potential targets for antiviral interventions. Specifically, the identification of substantial transcriptional overlaps by this work suggests the existence of a genome-wide interference between transcriptional machineries. This finding indicates the presence of a novel regulatory layer, potentially controlling the expression of viral genes.
Chitinase 3-like-1 (CHI3L1) in the Pathogenesis of Epidermal Growth Factor Receptor Mutant Non-Small Cell Lung Cancer
Authors: Kamle Suchitra , Ma Bing , Schor Gail , Bailey Madison , Pham Brianna , Cho Inyoung , Khan Hina , Azzoli Christopher , Hofstetter Mara , Lee Chang-Min , Herbst Roy , Politi Katerina , Lee Chun Geun , Elias Jack A.
Published: September 2023

Non-small cell lung cancer (NSCLC) accounts for 85% of all lung cancers. In NSCLC, 10-20% of Caucasian patients and 30-50% of Asian patients have tumors with activating mutations in the Epidermal Growth Factor Receptor (EGFR). A high percentage of these patients exhibit favorable responses to treatment with tyrosine kinase inhibitors (TKI). Unfortunately, a majority of these patients develop therapeutic resistance with progression free survival lasting 9-18 months. The mechanisms that underlie the tumorigenic effects ofEGFRand the ability of NSCLC to develop resistance to TKI therapies, however, are poorly understood. Here we demonstrate that CHI3L1 is produced by EGFR activation of normal epithelial cells, transformed epithelial cells with wild typeEGFRand cells with cancer-associated, activatingEGFRmutations. We also demonstrate that CHI3L1 auto-induces itself and feeds back to stimulate EGFR and its ligands. Highly specific antibodies against CHI3L1 (anti-CHI3L1/FRG) and TKI, individually and in combination, abrogated the effects of EGFR activation on CHI3L1 and the ability of CHI3L1 to stimulate the EGFR axis. Anti-CHI3L1 also interacted with osimertinib to reverse TKI therapeutic resistance and induce tumor cell death and inhibit pulmonary metastasis while stimulating tumor suppressor genes includingKEAP1. CHI3L1 is a downstream target of EGFR that feeds back to stimulate and activate the EGFR axis. Anti-CHI3L1 is an exciting potential therapeutic forEGFRmutant NSCLC, alone and in combination with osimertinib or other TKIs.
Asymmetric Contribution of a Selectivity Filter Gate in Triggering Inactivation of CaV1.3 Channels
Authors: del Rivero Morfin Pedro J. , Kochiss Audrey L. , Liedl Klaus R. , Flucher Bernhard E. , Fernández-Quintero Monica L.I. , Ben-Johny Manu
Published: September 2023

Voltage-dependent and Ca2+-dependent inactivation (VDI and CDI, respectively) of CaVchannels are two biologically consequential feedback mechanisms that fine-tune Ca2+entry into neurons and cardiomyocytes. Although known to be initiated by distinct molecular events, how these processes obstruct conduction through the channel pore remains poorly defined. Here, focusing on ultra-highly conserved tryptophan residues in the inter-domain interfaces near the selectivity filter of CaV1.3, we demonstrate a critical role for asymmetric conformational changes in mediating VDI and CDI. Specifically, mutagenesis of the domain III-IV interface, but not others, enhanced VDI. Molecular dynamics simulations demonstrate that mutations in distinct selectivity filter interfaces differentially impact conformational flexibility. Furthermore, mutations in distinct domains preferentially disrupt CDI mediated by the N- versus C-lobes of CaM, thus uncovering a scheme of structural bifurcation of CaM signaling. These findings highlight the fundamental importance of the asymmetric arrangement of the pseudo-tetrameric CaVpore domain for feedback inhibition.
Neuroinflammation induces nerve growth factor dependent nociceptor sensitisation in a neonatal rodent model of platinum-based chemotherapy induced neuropathic pain
Authors: Da Vitoria Lobo Marlene , Hardowar Lydia , Valentine Tameille , Tomblin Lucy , Guest Charlotte , Sharma Dhyana , Paul-Clark Mark , Hulse Richard Philip
Published: September 2023

Chemotherapy-induced neuropathic pain (CINP) is a common adverse health related comorbidity that manifests later in life in paediatric patients treated for cancer. CIPN pathology progressively develops over time resulting in a delayed but long-lasting neuropathic pain. Current analgesic strategies are ineffective, aligning closely with our lack of understanding of CINP. Recent studies have indicated alterations in sensory neuronal maturation as component of CINP. The aim of this study was to investigate how cisplatin induces nerve growth factor mediated neuroinflammation and nociceptor sensitisation. In a rodent model of cisplatin induced survivorship pain, there was a significant infiltration of nerve growth factor positive macrophages into the dorsal root ganglia (DRG), demonstrating a robust neuroinflammatory response. Additionally, it was observed that CD11b/F480 positive monocyte/macrophages challenged with cisplatin expressed more NGF. Additionally, DRG derived primary sensory neuron cultures from neonatal mice demonstrated enhanced NGF-dependent TRPV1 mediated nociceptor activity after cisplatin treatment. Increased nociceptor activity was also observed when cultured neurons were treated with conditioned media from cisplatin activated monocyte/macrophages. This elevated nociceptor activity was dose-dependently inhibited by a neutralising monoclonal antibody to NGF. Intraperitoneal administration of NGF neutralising antibody significant reduction in mechanical hypersensitivity was given to mice with cisplatin-induced juvenile survivorship pain there was a as well as suppression of cisplatin induced aberrant nociceptor intraepidermal nerve fibre density. These findings identify the NGF/TrkA signalling pathway as a potential novel therapeutic target for analgesia in adult survivors of childhood cancer.
S100A8/A9 predicts triple-negative breast cancer response to PIM kinase and PD-1/PD-L1 inhibition
Authors: Begg Lauren R. , Orriols Adrienne M. , Zannikou Markella , Yeh Chen , Vadlamani Pranathi , Kanojia Deepak , Bolin Rosemary , Dunne Sara F. , Balakrishnan Sanjeev , Camarda Roman , Roth Diane , Zielinski-Mozny Nicolette A. , Yau Christina , Vassilopoulos Athanassios , Huang Tzu-Hsuan , Kim Kwang-Youn A. , Horiuchi Dai
Published: September 2023

It remains elusive why some triple-negative breast cancer (TNBC) patients respond poorly to existing therapies while others respond well. Our retrospective analysis of historical gene expression datasets reveals that increased expression of immunosuppressive cytokine S100A8/A9 in early-stage tumors is robustly associated with subsequent disease progression in TNBC. Although it has recently gained recognition as a potential anticancer target, S100A8/A9 has not been integrated into clinical study designs evaluating molecularly targeted therapies. Our small molecule screen has identified PIM kinase inhibitors as capable of decreasing S100A8/A9 expression in multiple cell types, including TNBC and immunosuppressive myeloid cells. Furthermore, combining PIM inhibition and immune checkpoint blockade induces significant antitumor responses, especially in otherwise resistant S100A8/A9-high PD-1/PD-L1-positive tumors. Importantly, serum S100A8/A9 levels mirror those of tumor S100A8/A9 in a syngeneic mouse model of TNBC. Thus, our data suggest that S100A8/A9 could be a predictive and pharmacodynamic biomarker in clinical trials evaluating combination therapy targeting PIM and immune checkpoints in TNBC and encourage the development of S100A8/A9-based liquid biopsy tests.
Selection by differential survival among marine animals in the Phanerozoic
Authors: Tamre Erik , Parsons Christopher
Published: September 2023

The Gaia hypothesis posits that Earth’s biosphere functions as a single self-stabilizing system, but a key challenge is explaining how this could have arisen through Darwinian evolution. One theory is that of “selection by differential survival”, in which a clade’s extinction probability decreases with age as it accumulates adaptations resisting environmental disturbances. While this is hard to assess during early Earth history, we can assess whether this process operated among marine animal genera throughout the Phanerozoic. To that end, we analyzed time ranges of 36,117 extinct animal genera using fossil occurrence data from the Paleobiology Database in order to calculate marine metazoan extinction age selectivity, extinction rates, and speciation rates over the Phanerozoic. We identify four signatures of selection by differential survival: lower extinction rates among older lineages, heritability and taxonomically nested propagation of extinction resistance, reduced age selectivity during rare environmental perturbations, and differential extinction rather than speciation as the primary driver of the phenomenon. Evidence for this process at lower taxonomic levels also implies its possibility for life as a whole – indeed, the possibility of Gaia.
A haplotype-resolved chromosome-level assembly and annotation of European hazelnut (C. avellanacv. Jefferson) provides insight into mechanisms of eastern filbert blight resistance
Authors: Talbot S.C. , Vining K.J. , Snelling J.W. , Clevenger J. , Mehlenbacher S.A.
Published: September 2023

European hazelnut (Corylus avellanaL.) is an important tree nut crop. Hazelnut production in North America is currently limited in scalability due toAnisogramma anomala,a fungal pathogen that causes Eastern Filbert Blight (EFB) disease in hazelnut. Successful deployment of EFB resistant cultivars has been limited to the state of Oregon, where the breeding program at Oregon State University (OSU) has released cultivars with a dominant allele at a single resistance locus identified by classical breeding, linkage mapping, and molecular markers. ‘Jefferson’ is resistant to the predominant EFB biotype in Oregon and has been selected by the OSU breeding program as a model for hazelnut genetic and genomic research. Here, we present a near complete, haplotype-resolved chromosome-level hazelnut genome assembly forC. avellana‘Jefferson’. This new assembly is a significant improvement over a previously published genome draft. Analysis of genomic regions linked to EFB resistance and self-incompatibility confirmed haplotype splitting and identified new gene candidates that are essential for downstream molecular marker development, thereby facilitating breeding efforts.
Myocardial B cells have specific gene expression and predicted interactions in Dilated Cardiomyopathy and Arrhythmogenic Right Ventricular Cardiomyopathy
Authors: Bermea Kevin C. , Duque Carolina , Cohen Charles D. , Bhalodia Aashik , Rousseau Sylvie , Lovell Jana , Zita Marcelle Dina , Mugnier Monica R. , Adamo Luigi
Published: September 2023

Introduction

Growing evidence from animal models indicates that the myocardium hosts a population of B cells that play a role in the development of cardiomyopathy. However, there is minimal data on human myocardial B cells in the context of cardiomyopathy.

Methods

We integrated single-cell and single-nuclei datasets from 45 healthy human hearts, 70 hearts with dilated cardiomyopathy (DCM), and 8 hearts with Arrhythmogenic Right Ventricular Cardiomyopathy (ARVC). Interactions between B cells and other cell types were investigated using the CellChat Package. Differential gene expression analysis comparing B cells across conditions was performed using DESeq2. Pathway analysis was performed using Ingenuity, KEGG, and GO pathways analysis.

Results

We identified 1,100 B cells, including naive B cells and plasma cells. B cells showed an extensive network of interactions within the healthy myocardium that included outgoing signaling to macrophages, T cells, endothelial cells, and pericytes, and incoming signaling from endothelial cells, pericytes, and fibroblasts. This niche relied on ECM-receptor, contact, and paracrine interaction; and changed significantly in the context of cardiomyopathy, displaying disease-specific features. Differential gene expression analysis showed that in the context of DCM both naive and plasma B cells upregulated several pathways related to immune activation, including upregulation of oxidative phosphorylation, upregulation of leukocyte extravasation, and, in naive B cells, antigen presentation.

Discussion

The human myocardium contains naive B cells and plasma cells, integrated into a diverse and dynamic niche that has distinctive features in healthy myocardium, DCM, and ARVC. Naive myocardial-associated B cells likely contribute to the pathogenesis of human DCM.
Precise Alterations in Hippocampal Neural Deterioration and Neural Correlates of Visual Short-Term Memory in Individuals with Amnestic Mild Cognitive Impairment
Authors: Xie Ye , Zhao Tinghao , Li Yunxia , Ku Yixuan
Published: September 2023

Purpose

Atrophy of the hippocampus is an early biomarker of Alzheimer’s disease (AD) and a main contributor to the patients’ mnemonic degeneration. While previous research has mostly focused on how hippocampal atrophy impaired long-term memory performance, its relation to short-term memory impairment, which was also found among patients with AD, remains largely uninvestigated. Filling this gap, the current study examined how atrophy in the hippocampus and its subfields may have influenced visual short-term memory (VSTM) in patients with amnestic mild cognitive impairment (aMCI), a common precursor of AD.

Methods

Fifty-eight aMCI patients and 69 healthy controls (HC) matched in age were included in the current study. VSTM was assessed using an adapted change detection task with a memory load of 2 or 4 items. Hippocampal subfields were automatically segmented in T1-weighted image using FreeSurfer and were manually inspected for errors. Volumes of the subfields were extracted and compared between aMCI and HC subjects using ANCOVA with age, gender and education as covariates. Furthermore, we also examined the partial correlation between VSTM performances and hippocampal subfield volumes with age, gender and education as covariates.

Results

Compared to HC subjects, aMCI subjects had lower response accuracy (ACC) and lower memory capacity under both load conditions and had longer reaction time (RT) in the 2-load condition. Left hippocampus volume was significantly smaller in aMCI and was positively correlated with ACC and capacity in HC but not in aMCI. Among the hippocampal subfields, left hippocampal tail, left molecular layer, left dentate gyrus (DG), left CA4, bilateral subiculum, bilateral presubiculum, bilateral fimbria, were significantly smaller while right hippocampal fissure was significantly widened in aMCI compared to HC. Volumes of the left subiculum, left molecular layer, left DG and left CA4 were positively correlated with ACC and capacity in HC but not in aMCI. Bilateral fimbria volume was negatively correlated with RT under the 2-load condition in HC but not in aMCI.

Conclusion

The results of this study suggested that hippocampal deterioration, especially in subfields related to information input and output (e.g. molecular layer, DG, subiculum), may have contributed to VSTM impairment in aMCI by disrupting hippocampal-cortical communications. This finding adds to increasing evidence of hippocampal engagement in short-term memory processes and points to VSTM impairment as a potential neuropsychological indicator for MCI and AD.
Upregulation of ATP Citrate Lyase Phosphorylation and Neutral Lipid Synthesis through Viral Growth Factor Signaling during Vaccinia Virus Infection
Authors: Pant Anil , Brahim Belhaouari Djamal , Dsouza Lara , Yang Zhilong
Published: September 2023

Like all other viruses, poxviruses rely on host cells to provide metabolites and energy. Vaccinia virus (VACV), the prototype poxvirus, induces profound metabolic alterations in host cells. We previously showed that VACV infection increases the tricarboxylic acid (TCA) cycle intermediates, including citrate, that can be transported to the cytosol to be converted to acetyl-CoA forde novofatty acid biosynthesis. ATP citrate lyase (ACLY) is a pivotal enzyme converting citrate to acetyl-CoA. Here, we report that VACV infection stimulates the S455 phosphorylation of ACLY, a post-translational modification that stimulates ACLY activity. We demonstrate that the chemical and genetic inhibition of ACLY severely suppresses VACV replication. Remarkably, we found that virus growth factor (VGF)-induced signaling is essential for the VACV-mediated upregulation of ACLY phosphorylation. Furthermore, the upregulation of ACLY phosphorylation during VACV infection is dependent on the activation of the cellular Akt kinase that phosphorylates ACLY. Finally, we report that VGF-induced ACLY phosphorylation via the EGFR-Akt pathway is important for VACV stimulations of neutral lipid droplet synthesis. These findings identified a previously unknown way of rewiring cell metabolism by a virus and a novel function for VGF in the governance of virus-host interactions through the induction of a key enzyme at the crossroads of the TCA cycle and fatty acidde novobiosynthesis. Our study also provides a mechanism for the role played by VGF and its downstream signaling cascades in the modulation of lipid metabolism in VACV-infected cells.

Importance

ATP citrate lyase is a key metabolic enzyme that sits at the crossroads of glucose, glutamine, and lipid metabolism. However, how virus infection affects this protein is unclear. Using chemical, genetic, and metabolic approaches we show that VACV, the prototype poxvirus, increases the phosphorylation of ACLY in primary human fibroblasts in a VGF-dependent manner. We further show that the VGF-EGFR-Akt signaling pathway is vital for VACV-induced lipid droplet synthesis. Our findings identified ACLY as a potential target for novel antiviral development against pathogenic poxviruses. Our study also expands the role of growth factor signaling in boosting VACV replication by targeting multiple metabolic pathways.
Application of CRISPR-Cas12a technology in zebrafish to simulate ß-globin gene (HBB) mutations
Authors: Shafique Farheen , Ali Shaukat , Balckburn Edward , ul Hassan Mahreen
Published: September 2023

Background

The human ß-globin (HBB) gene translated into the hemoglobin subunit beta (ß) of the hemoglobin protein. When mutated, it can lead to the blood disorder ß-thalassemia. Recently, the CRISPR-Cas12 technology has shown promising potential in treating different genetic abnormalities.

Objectives

This study aimed to evaluate the feasibility of zebrafish hbbe1.1 gene editing via CRISPR-Cas12a gene-technology.

Methods

In current study, thalassemic mutations were replicated in zebrafish to improve gene editing. the embryonic hemoglobin genehbbe1.1was preferred over adult hemoglobinhbß-a1due to its early detection at larval stages. The CRISPR-Cas12 technology was utilised in combination with phosphorothioated “rescue template” (ssDNA L33P) to introduce base edits to the DNA sequence of zebrafish.

Results

These experiments indeed resulted in the alteration of a single amino acid at the protein level. With the help of this genetic editing method, we were able to generate a novel zebrafish strain that carried specific amino acid alterations resembling the pathogenic mutations found in HBB for ß-thalassemia disease. However, in several cases additional indels or base alterations were observed.

Conclusion

Our findings suggests that MMEJ double stranded break repair mechanism causes more knock-in events and germline inheritance than HR-mediated events. Modifying the technique could improve results by reducing MMEJ frequency.
The atypical cadherin FAT1 is a novel regulator of STAT1, driving its pro-tumorigenic effect via the STAT1/PDCD4 axis in glioblastoma
Authors: Khan Md. Tipu , Malik Nargis , Jalota Akansha , Sharma Shaifali , Almas Mariyam , Luthra Kalpana , Suri Vaishali , Suri Ashish , Seth Pankaj , Chosdol Kunzang , Sinha Subrata
Published: September 2023

FAT1 is an atypical cadherin that has been shown to act both as an oncogene and a tumour suppressor gene (TSG) in different tumor types. We have earlier shown that upregulated FAT1 acts as an oncogene in glial tumors by promoting pro-tumorigenic inflammation and EMT in primary human glioblastoma and in cell lines. One effect was through the suppression of the Tumor Suppressor Gene (TSG), Programmed Cell Death 4 (PDCD4). Here, we have studied how, in glioblastoma, upregulated FAT1 affects downstream events that control PDCD4 expression.In silicoanalysis of the PDCD4 promoter revealed multiple STAT1 binding sites. We also found a positive correlation in mRNA levels of STAT1 and FAT1 in the Glioblastoma databases, as well as in resected patient derived tumor samples by qPCR. Increased FAT1 as well as STAT1 were associated with poor prognosis in these data bases. In the glioblastoma cell lines LN229 and U87MG, FAT1 knockdown resulted in decreased STAT1 expression. Also, STAT1 knockdown resulted in increased PDCD4 expression, implying that STAT1 may mediate FAT1’s role in suppressing PDCD4. Further, ChIP experiments showed that STAT1 protein binds to the PDCD4 promoter and upon FAT1 knockdown, STAT1 binding to the PDCD4 promoter reduces. As for FAT1, STAT1 knockdown also reduces the expression of pro-inflammatory cytokines and EMT markers, also migration and invasion of glioma derived cell lines. This work identifies STAT1 as a novel downstream mediator of FAT1 which mediates its pro-tumorigenic action in suppressing the TSG, PDCD4.
Microfluidic interfaces for chronic bidirectional access to the brain
Authors: Marcigaglia Simone , De Plus Robin , Vandendriessche Charysse , Cuypers Marie-Lynn , Cools Jordi , Hoffman Luis Diego , Vandenbroucke Roosmarijn , Dewilde Maarten , Haesler Sebastian
Published: September 2023

Abstract

Here, we used micron-scale 3D printing to develop microfluidic interfaces which provide chronic fluidic access to the brain of preclinical research models. In mice, we show the delivery interface enables faster, more precise and physiologically less disruptive fluid injection. Moreover, we demonstrate the blood brain barrier (BBB) is intact after chronic implantation of the sampling interface and establish frequent, longitudinal sampling of CSF and biomarkers from the ventricle over long time periods of up to 200 days.
Microfibril-associated glycoprotein 4 forms octamers that mediate interactions with elastogenic proteins and cells
Authors: Wozny Michael R. , Nelea Valentin , Siddiqui Iram Fatima S. , Wanga Shaynah , de Waard Vivian , Strauss Mike , Reinhardt Dieter P.
Published: September 2023

Microfibrillar-associated protein 4 (MFAP4) is a 36-kDa extracellular glycoprotein with critical roles in human pathologies, including fibrosis in several organ systems, chronic obstructive pulmonary disease, and cardiovascular disorders. In elastic tissues such as arteries, lungs, and skin, MFAP4 associates with microfibrils and elastic fibres, which are the central extracellular fibres affected in thoracic aneurysms. MFAP4 directly interacts with elastogenic proteins, including fibrillin-1 and tropoelastin, and with cells via integrins. MFAP4 multimerisation represents a critical hallmark required for its physiological and pathological properties. However, molecular details and functional consequences of MFAP4 multimerisation are lacking.

Here we present a cryo-electron microscopy structure of human MFAP4. In the presence of calcium, MFAP4 assembles as an octamer with D2 point group symmetry, where two sets of homodimers constitute the top and bottom halves of each octamer. Each of the homodimers is linked together by an inter-molecular disulfide bond. An engineered C34S missense mutation in MFAP4 prevented disulfide-bond formation between monomers, but the mutant formed octamers similar to wild type MFAP4. The atomic model, built into the 3.55 Å cryo-EM map, suggests that several salt-bridges are important for interactions within and between homodimers, while non-polar interactions are important for octamer halves to assemble. In the absence of calcium, MFAP4 dissociates into tetramers, representing the top/bottom halves of the octamers. Binding studies with elastogenic proteins, including fibrillin-1, tropoelastin, LTBP4, and small fibulins showed that MFAP4 has multiple surfaces for protein-protein interactions, which depend upon the higher-order assembly of MFAP4. While the disulfide-bond mediated by C34S contributes little to those protein interactions, it modulated cell interaction. When MFAP4 forms assemblies with fibrillin-1, it abrogates MFAP4 interactions with cells. Overall, the study provides detailed molecular structure-function relationships of MFAP4 interactions with elastogenic proteins and cells.
Multi-omics resources for the Australian southern stuttering frog (Mixophyes australis) reveal assorted antimicrobial peptides
Authors: Tang Simon , Peel Emma , Belov Katherine , Hogg Carolyn J. , Farquharson Katherine A.
Published: September 2023

The number of genome-level resources for non-model species continues to rapidly expand. However, frog species remain underrepresented, with up to 90% of frog genera having no genomic or transcriptomic data. Here, we assemble the first genomic and transcriptomic resources for the recently described southern stuttering frog (Mixophyes australis). The southern stuttering frog is ground-dwelling, inhabiting naturally vegetated riverbanks in south-eastern Australia. Using PacBio HiFi long-read sequencing and Hi-C scaffolding, we generated a high-quality genome assembly, with a scaffold N50 of 369.3 Mb and 95.1% of the genome contained in twelve scaffolds. Using this assembly, we identified the mitochondrial genome, and assembled six tissue-specific transcriptomes. We also bioinformatically characterised novel sequences of two families of antimicrobial peptides (AMPs) in the southern stuttering frog, the cathelicidins and β-defensins. While traditional peptidomic approaches to peptide discovery have typically identified one or two AMPs in a frog species from skin secretions, our bioinformatic approach discovered 12 cathelicidins and two β-defensins that were expressed in a range of tissues. We investigated the novelty of the peptides and found diverse predicted activities. Our bioinformatic approach highlights the benefits of multi-omics resources in peptide discovery and contributes valuable genomic resources in an under-represented taxon.
Chemical proteomic approach for in-depth glycosylation profiling of plasma carcinoembryonic antigen in cancer patients
Authors: Chen Jin , Yang Lijun , Li Chang , Zhang Luobin , Gao Weina , Xu Ruilian , Tian Ruijun
Published: September 2023

Carcinoembryonic antigen (CEA) of human plasma is a biomarker of many cancer diseases, and its N-glycosylation accounts for 60% of molecular mass. It is highly desirable to characterize its glycoforms for providing additional dimension of features to increase its performance in prognosis and diagnosis of cancers. However, to systematically characterize its site-specific glycosylation is challenging due to its low abundance. Here, we developed a highly sensitive strategy for in-depth glycosylation profiling of plasma CEA through chemical proteomics combined with multi-enzymatic digestion. A trifunctional probe was utilized to generate covalent bond of plasma CEA and its antibody upon UV irradiation. As low as 1 ng/mL CEA in plasma could be captured and digested with trypsin and chymotrypsin for intact glycopeptide characterization. Twenty six out of 28 potential N-glycosylation sites were well identified, which were the most comprehensive N-glycosylation site characterization of CEA on intact glycopeptide level as far as we known. Importantly, this strategy was applied to the glycosylation analysis of plasma CEA in cancer patients. Differential site-specific glycoforms of plasma CEA were observed in patients with colorectal carcinomas (CRC) and lung cancer. The distributions of site-specific glycoforms were different as the progression of CRC, and most site-specific glycoforms were overexpressed in stage II of CRC. Overall, we established a highly sensitive chemical proteomic method to profile site-specific glycosylation of plasma CEA, which should generally applicable to other well-established cancer glycoprotein biomarkers for improving their cancer diagnosis and monitoring performance.

In Brief

A chemical proteomic approach for glycosylation profiling of proteins was established for glycosylation characterization of plasma CEA with low abundance. Although CEA has been widely used in diagnosis and prognosis of many cancers, it lacks specificity and sensitivity. We found that the glycosylation of CEA on intact glycopeptide level provided additional dimension of molecular features to improve the performance of CEA in cancer diagnosis and progression.

Highlights

A chemical proteomic approach for glycosylation profiling of proteins with low abundance

Glycosylation identification of plasma CEA on intact glycopeptide level with high sensitivity and reproducibility

Glycosylation features of plasma CEA in cancer patients with CRC and lung cancer and in CRC patients at different progression stages

Graphical Abstract
Modeling the mosaic structure of bacterial genomes to infer their evolutionary history
Authors: Sheinman Michael , Arndt Peter F. , Massip Florian
Published: September 2023

The timing and phylogeny of bacterial evolution is difficult to reconstruct because of a scarce fossil record, deep genomic divergences and complexities associated with molecular clocks. Studying bacterial evolutionary history using rich and rapidly accumulating genomic data requires accurate modeling of genome evolution, taking into account that different parts of bacterial genomes have different history. In particular, along the genome, different loci are subject to different selective pressure. In addition, some are horizontally transferred from one bacterium to another, resulting in a mosaic-like genome structure. An important technical aspect is that loci with high effective mutation rates can diverge beyond the aligner detection limit, biasing the genome-wide divergence estimate towards more conserved loci. Therefore, the genome-wide molecular clock cannot be directly applied to study bacterial evolutionary history. In this article, we propose a novel method to gain insight into bacterial evolution based on statistical properties of genomic sequences comparisons. The length distribution of the sequence matches is shaped by the effective mutation rates of different loci, by the horizontal transfers and by the aligner sensitivity. Based on these inputs we build a model and demonstrate that it accounts for the empirically observed distributions, taking theEnterobacteriaceaefamily as an example. Using the model and the empirical data we fit the evolutionary parameters: time divergences and horizontal transfer rates. Based on the estimated time divergences we build a time-calibrated phylogenetic tree, demonstrating the accuracy of the method and its ability to unravel vertical and horizontal transfers in bacterial genomes.
A new multiplexed magnetic-capture - droplet digital PCR (ddPCR) tool for monitoring wildlife population health and pathogen surveillance
Authors: Tschritter Christina , de Groot Peter V. C. , Branigan Marsha , Dyck Markus , Sun Zhengxin , Lougheed Stephen C.
Published: September 2023

Anthropogenic stressors are exacerbating the emergence and spread of pathogens worldwide. In regions like the Arctic where ecosystems are particularly susceptible, marked changes are predicted in regional diversity, intensity, and patterns of infectious diseases. To understand such rapidly changing host-pathogen dynamics and mitigate the impacts of novel pathogens, we need sensitive disease surveillance tools. We developed and validated a novel multiplexed, magnetic-capture and ddPCR tool for the surveillance of multiple pathogens in polar bears, a sentinel species that is considered susceptible to climate change and other stressors with a pan-Arctic distribution. Through sequence-specific magnetic capture, we concentrated five target template sequences from three zoonotic bacteria (Erysipelothrix rhusiopathiae,Francisella tularensis, andMycobacterium tuberculosiscomplex) and two parasitic (Toxoplasma gondiiandTrichinellaspp.) pathogens from large quantities (< 100 g) of host tissue. We then designed and validated two multiplexed probe-based ddPCR assays for the amplification and detection of the low concentration target DNA. Validations used 48 polar bear tissues (muscle and liver). We detected 14, 1, 3, 4, and 22 tissue positives forE. rhusiopathiae,F. tularensis,Mycobacterium tuberculosiscomplex,T. gondii, andTrichinellaspp., respectively. These multiplexed assays offer a rapid, specific tool for quantifying and monitoring the changing geographical and host distributions of pathogens relevant to human and animal health.
Spinal cord pathology in a Dravet Syndrome mouse model
Authors: Antinao Diaz Juan , Chilcott Ellie , Keegan Anna , Schorge Stephanie , Waddington Simon N , Karda Rajvinder
Published: September 2023

Objectives

Dravet syndrome is a severe epileptic encephalopathy that begins in early childhood. More than 80% of patients with Dravet syndrome exhibit a haploinsufficiency inSCN1A, which encodes the voltage-gated sodium ion channel NaV1.1. The epilepsy is believed be caused by specific deficit ofSCN1Ain inhibitory interneurons of the hippocampus. However, the aetiology of other symptoms including gait disturbances, ataxia, cardiac issues and dysautonomia is less clear.

Methods

In anScn1aknock-out (Scn1a-/-) mouse model which recapitulates clinical phenotypes, we assessed NaV1.1 and neuroinflammation throughout the central nervous system.

Results

Consistent with current understanding, wild-type expression of NaV1.1 transcript and protein were absent in knock-out mice in the prefrontal cortex, striatum, hippocampus, thalamus, and cerebellum. Increased GFAP was detected in the brain only in the hippocampus. Transcript and protein were detected in wild-type cervical, thoracic and lumbar spinal cord but not in knock-out mice. Unexpectedly, GFAP was increased in all three spinal regions. Therefore, we proceeded to perform transcriptomic analysis of cortex, hippocampus and spinal cord. Pathways associated with monooxygenase activity, fatty acid ligases and lactate transporters were highly dysregulated in the spinal cord.

Conclusion

The existence and relevance of pathology of the spinal cord in Dravet syndrome has received scant attention. Our findings are consistent with some systemic symptoms of Dravet syndrome, with the benefits of treatments which may modulate the astrocyte-neuron lactate shuttle such as Stiripentol and ketogenic dietary regimes, and with the efficacy of intrathecal delivery of therapeutics.

Key Points

Decrease of endogenousScn1aand NaV1.1 expression inScn1a-/-mice has a widespread impact on the gene expression profile in the spinal cord.

Increased GFAP expression observed in the spinal cord ofScn1a-/-mice.

Differentially expressed genes related to monooxygenase activity, fatty acid ligases and lactate transporters in cervical spinal cord ofScn1a-/-mice.
Sensory experience steers representational drift in mouse visual cortex
Authors: Bauer Joel , Lewin Uwe , Herbert Elizabeth , Gjorgjieva Julijana , Schoonover Carl , Fink Andrew , Rose Tobias , Bonhoeffer Tobias , Hübener Mark
Published: September 2023

Representational drift – the gradual continuous change of neuronal representations – has been observed across many brain areas. It is unclear whether drift is caused by synaptic plasticity elicited by sensory experience, or by the intrinsic volatility of synapses. Here, using chronic two-photon calcium imaging in mouse primary visual cortex, we find that the preferred stimulus orientation of individual neurons slowly drifts over the course of weeks. By using cylinder lens goggles to limit visual experience to a narrow range of orientations, we show that the direction of drift, but not its magnitude, is biased by the statistics of visual input. A network model suggests that drift of preferred orientation largely results from synaptic volatility, which under normal visual conditions is counteracted by experience-driven Hebbian mechanisms, stabilizing preferred orientation. Under deprivation conditions these Hebbian mechanisms enable adaptation. Thus, Hebbian synaptic plasticity steers drift to match the statistics of the environment.
Regeneration of actin filament branches from the same Arp2/3 complex
Authors: Ghasemi Foad , Cao LuYan , Mladenov Miroslav , Guichard Bérengère , Way Michael , Jégou Antoine , Romet-Lemonne Guillaume
Published: September 2023

Branched actin filaments are found in many key cellular structures. Branches are nucleated by the Arp2/3 complex activated by nucleation-promoting factor (NPF) proteins and bound to the side of pre-existing ‘mother’ filaments. Over time, branches dissociate from their mother filament, leading to network reorganization and turnover, but this mechanism is less understood. Here, using microfluidics and purified proteins, we examined the dissociation of individual branches under controlled biochemical and mechanical conditions. We observe that Arp2/3 remains bound to the mother filament after most debranching events, even when accelerated by force. Unexpectedly, this mother-remaining Arp2/3 readily nucleates a new actin filament branch, without being activated anew by an NPF: it simply needs to exchange its nucleotide and bind an actin monomer. The protein GMF, which accelerates debranching, prevents branch re-nucleation. Our results suggest that actin filament re-nucleation can provide a self-repair mechanism, helping branched networks to sustain mechanical stress in cells over extended periods of time.
eQTLs identify regulatory networks and drivers of variation in the individual response to sepsis
Authors: Burnham Katie L. , Milind Nikhil , Lee Wanseon , Kwok Andrew J. , Cano-Gamez Eddie , Mi Yuxin , Geoghegan Cyndi G. ,   GAinS Investigators, McKechnie Stuart , Soranzo Nicole , Hinds Charles J. , Knight Julian C. , Davenport Emma E.
Published: September 2023

Sepsis is a clinical syndrome of life-threatening organ dysfunction caused by a dysregulated response to infection, for which disease heterogeneity is a major obstacle to developing targeted treatments. We have previously identified gene expression-based patient subgroups (Sepsis Response Signatures: SRS) informative for outcome and underlying pathophysiology. Here we aimed to investigate the role of genetic variation in determining the host transcriptomic response and to delineate regulatory networks underlying SRS. Using genotyping and RNA-seq data on 638 adult sepsis patients, we report 16,049 independent expression (eQTLs) and 32 co-expression module (modQTLs) quantitative trait loci in this disease context. We identified significant interactions between SRS and genotype for 1,578 SNP-gene pairs, and combined transcription factor (TF) binding site information (SNP2TFBS) and predicted regulon activity (DoRothEA) to identify candidate upstream regulators. These included HIF1A and CEBPB, which were associated with progenitor and immature neutrophil subsets respectively, further implicating glycolysis and emergency granulopoiesis in SRS1. Overall, these approaches identified putative mechanistic links between host genetic variation, cell subtypes, and the individual transcriptomic response to infection. Understanding the regulatory networks underlying patient heterogeneity provides additional information for developing immunomodulatory treatments and a personalised medicine approach to treating sepsis.