bluuebunny/crossref_metadata_2025 huggingface dataset:
Loss of Insulin Signaling in Microglia Impairs Cellular Uptake of Aβ and Neuroinflammatory Response Exacerbating Alzheimer-like Neuropathology
Authors: Chen Wenqiang , Liu Xiangyu , Munoz Vitor Rosetto , Kahn C. Ronald
Published: August 2024

Insulin receptors are present on cells throughout the body, including the brain. Dysregulation of insulin signaling in neurons and astrocytes has been implicated in altered mood, cognition, and the pathogenesis of Alzheimers disease (AD). To define the role of insulin signaling in microglia, the primary phagocytes in brain critical for maintenance and damage repair, we created mice with an inducible microglia-specific insulin receptor knockout (MG-IRKO). RiboTag profiling of microglial mRNAs revealed that loss of insulin signaling results in alterations of gene expression in pathways related to innate immunity and cellular metabolism. In vitro, loss of insulin signaling in microglia results in metabolic reprograming with an increase in glycolysis and impaired uptake of Aβ. In vivo, MG-IRKO mice exhibit alterations in mood and social behavior, and when crossed with the 5xFAD mouse model of AD, the resultant mice exhibit increased levels of Aβ; plaque and elevated neuroinflammation. Thus, insulin signaling in microglia plays a key role in microglial cellular metabolism, neuroinflammation and the ability of the cells to take up Aβ; such that reduced insulin signaling in microglia alters mood and social behavior and accelerates AD pathogenesis. Together these data indicate key roles of insulin action in microglia and the potential of targeting insulin signaling in microglia in treatment of AD.
Zfp263 is a transcriptional checkpoint of neutrophil development
Authors: Devaprasad Abhinandan , Ai Zhichao , Mukherjee Ananda Kishore , Panczel Aron , Khoyratty Tariq , Ballesteros Ivan , Rubio-Ponce Andrea , Grinsven Erinke van , Wang Lihui , Gomez Julia Salafranca , Eames Hayley L , Hidalgo Andres , Mocsai Attila , Udalova Irina A
Published: August 2024

During development neutrophils undergo extensive changes in nuclear morphology, gene expression programmes, and acquire specific effector functions. However, molecular mechanisms controlling neutrophil development remain largely unknown. Here we systematically analysed changes in gene expression and chromatin landscape across stages of neutrophil differentiation, to construct a network of transcription factors, predicted to control neutrophil development and acquisition of functions. Zinc finger protein 263 (Zfp263) emerged at the apex of this network. Neutrophil maturation ex vivo and in vivo was blocked in cells lacking Zfp263 and accelerated if its levels were induced. Mechanistically, Zfp263 acts as a direct transcriptional repressor of the ERK1/2 pathway, that supports the proliferative capacity of neutrophil progenitors and limits their differentiation into mature cells. Blocking ERK1/2 signalling in the Zfp263 deficient cells using chemical inhibitors, rescues neutrophil differentiation. Other targets of Zfp263 include genes involved in neutrophil migration, cytokine and chemokine expression, reactive oxygen species and formation of neutrophil extracellular traps. Thus, our study outlines a framework for modulation of neutrophil development and/or function, aimed at neutrophil-mediated diseases.
Confinement and Catalysis WithinDe NovoDesigned Peptide Barrels
Authors: Petrenas Rokas , Hawkins Olivia A. , Jones Jacob F. , Scott D. Arne , Fletcher Jordan M. , Obst Ulrike , Lombardi Lucia , Pirro Fabio , Leggett Graham J. , Oliver Thomas A.A. , Woolfson Derek N.
Published: August 2024

De novoprotein design has advanced such that many peptide assemblies and protein structures can be generated predictably and quickly. The drive now is to bring functions to these structures, for example, small-molecule binding and catalysis. The formidable challenge of binding and orienting multiple small molecules to direct chemistry is particularly important for paving the way to new functionalities. To address this, here we describe the design, characterization, and application of small-molecule:peptide ternary complexes in aqueous solution. This uses α-helical barrel (αHB) peptide assemblies, which comprise 5 or more α-helices arranged around central channels. These channels are solvent accessible, and their internal dimensions and chemistries can be altered predictably. Thus, αHBs are analogous to ‘molecular flasks’ made in supramolecular, polymer, and materials chemistry. Using Förster resonance energy transfer as a readout, we demonstrate that specific αHBs can accept two different organic dyes, 1,6-diphenyl-1,3,5-hexatriene and Nile Red in close proximity. In addition, two anthracene molecules can be accommodated within an αHB to promote photocatalytic anthracene-dimer formation. However, not all ternary complexes are productive, either in energy transfer or photocatalysis, illustrating the control that can be exerted by judicious choice and design of the αHB.
Pharmacological CDK4/6 inhibition promotes vulnerability to lysosomotropic agents in breast cancer
Authors: Nehme Jamil , Maassen Sjors , Bravaccini Sara , Zanoni Michele , Gianni Caterina , Giorgi Ugo De , Soto-Gamez Abel , Altulea Abdullah , Gheorghe Teodora , Wang Boshi , Demaria Marco
Published: August 2024

Breast cancer is a leading cause of mortality worldwide. Pharmacological inhibitors of Cyclin- Dependent Kinases (CDK) 4 and 6 (CDK4/6i) inhibit breast cancer growth by inducing a senescent-like state. However, the long-term treatment efficacy remains hindered by the development of drug resistance. Clearance of senescent-like cancer cells may extend the durability of treatment. In this study, we showed that CDK4/6i-treated breast cancer cells exhibit various senescence-associated phenotypes, but remain insensitive to common senolytic compounds. By searching for novel vulnerabilities, we identified a significantly increased lysosomal mass and altered lysosomal structure across various breast cancer cell types upon exposure to CDK4/6i in preclinical systems and clinical specimens. We demonstrated that these lysosomal alterations render breast cancer cells sensitive to lysosomotropic agents, such as L- leucyl-L-leucine methyl ester (LLOMe) and salinomycin. Importantly, sequential treatment with CDK4/6i/lysosomotropic agents effectively reduced the growth of both Hormone Receptor- positive (HR+) and triple-negative breast cancer (TNBC) cells in vivo. This sequential therapeutic strategy offers a promising approach to eliminate CDK4/6i-induced senescent(-like) cells, potentially reducing tumor recurrence and enhancing the overall efficacy of breast cancer therapy.
Virtual Clinical Trials of BMP4 Differentiation Therapy: Digital Twins to Aid Glioblastoma Trial Design
Authors: Harbour Nicholas , Curtin Lee , Hubbard Matthew E , Jackson Pamela R , Rani Vinitha , Carrano Anna , Farias Virginea de Araujo , Kenchappa Rajappa , Quinones-Hinojosa Alfredo , Owen Markus , Swanson Kristin R
Published: August 2024

We leverage an integrative mathematical modeling framework to translate the impact of preclinical findings in virtual clinical trials. We develop a virtual clinical trial pipeline to face the real-world problem of numerous of actual early phase clinical trials that have failed for glioma/glioblastoma, the most common primary brain tumor. Even with the most promising preclinical data, designing clinical trials is fraught with challenges, including controlling for the many parameters used to inform patient selection criteria. Here, we introduce a virtual trial pipeline that allows us to consider the variability from some of these criteria that can be used for future trials of novel therapies. As an example, we apply this to the proposed delivery of BMP4 to stem cell niches present in glioblastoma, the most aggressive glioma, known for its inter- and intra-patient heterogeneity. The proposed approach of BMP4 treatment, delivered through adipose-derived mesenchymal stem cells, aims to promote cellular differentiation away from the treatment-resistance stem cell niches towards a more treatment-vulnerable state. This pipeline will help us narrow down strategies for future trials, optimize timing of treatments relative to key standard-of-care treatments, and predict synergy amongst the developed treatments.
Selection can favor a recombination landscape that limits polygenic adaptation
Authors: Parée Tom , Noble Luke , Roze Denis , Teotónio Henrique
Published: August 2024

Meiotic crossover positions are uneven along eukaryotic chromosomes, giving rise to heterogeneous recombination rate landscapes. Genetic modifiers of local and genome-wide crossover positions have been described, but the selective pressures acting on them and their potential effect on adaptation in already-recombining populations remain unclear. We performed experimental evolution using a mutant that modifies the position of crossovers along chromosomes in the nematodeCaenorhabditis elegans, without any detectable direct fitness effect. Our results show that when the recombination landscape is fixed, adaptation is facilitated by the modifier allele that, on average, increases recombination rates in genomic regions containing heritable fitness variation. However, in polymorphic populations containing both the wild-type and mutant modifier alleles, the allele that facilitates adaptation tends to decrease in frequency. This is likely because the allele that reduces recombination between selected loci at the genome-wide scale increases recombination in its chromosomal vicinity, and may thus benefit from local associations it establishes with beneficial genotype combinations. These results demonstrate that indirect selection acting on a recombination modifier mainly depends on its local effect, which may be decoupled from its consequences on genome-wide polygenic adaptation.
Latent Diffusion for Conditional Generation of Molecules
Authors: Kaufman Benjamin , Williams Edward C. , Pederson Ryan , Underkoffler Carl , Panjwani Zahid , Wang-Henderson Miles , Mardirossian Narbe , Katcher Matthew H. , Strater Zack , Grandjean Jean-Marc , Lee Bryan , Parkhill John
Published: August 2024

Abstract

Designing a small molecule therapeutic is a challenging multi-parameter optimization problem. Key properties, such as potency, selectivity, bioavailability, and safety must be jointly optimized to deliver an effective clinical candidate. We present COATI-LDM, a novel application of latent diffusion models to the conditional generation of property-optimized, drug-like small molecules. Diffusive generation of latent molecular encodings, rather than direct diffusive generation of molecular structures, offers an appealing way to handle the small and mismatched datasets that are common for molecular properties. We benchmark various diffusion guidance schemes and sampling methods against a pre-trained autoregressive transformer and genetic algorithms to evaluate control over potency, expert preference, and various physicochemical properties. We show that conditional diffusion allows control over the properties of generated molecules, with practical and performance advantages over competing methods. We also apply the recently introduced idea of particle guidance to enhance sample diversity. We prospectively survey a panel of medicinal chemists and determine that we can conditionally generate molecules that align with their preferences via a learned preference score. Finally, we present a partial diffusion method for the local optimization of molecular properties starting from a seed molecule. Conditional generation of small molecules using latent diffusion models on molecular encodings provides a highly practical and flexible alternative to prior molecular generation schemes.
Large-scale manipulation of radial positioning does not affect most aspects of genome organization
Authors: Guetta Mor , Kariti Hagai , Sherman Liora , Israitel Lena , Tel-Paz Moran , Avidan Nili , Shemer Revital , Kaplan Noam
Published: August 2024

Although the spatial organization of the genome is closely linked to biological function, genome structure is highly stochastic. Within this heterogeneity, specific function-associated structural elements must be maintained, even when the nucleus is deformed due to physiological physical constraints. The radial positioning of genomic loci - their distance from the nuclear periphery - has long been considered an important feature of genome organization which is correlated with both structural elements and genomic activity.

In the current study, we developed an experimental system for manipulating the radial positioning of the genome, by expressing the sperm-specific protein Prm1 in somatic cells. By microscopy, we observe that initial Prm1 nuclear foci develop within 72 hours into a large Prm1 focus occupying most of the nuclear interior, while the entire genome is driven towards the nuclear periphery, resulting in a 3-5 fold reduction in the volume that the genome occupies. Noting that this system enables isolation of a pure population of cells with reorganized nuclei, we then used Hi-C to study the effects of this perturbation. Remarkably, we find that interaction patterns are largely robust to this major nuclear reorganization, with minor changes which mostly reflect a strengthening of heterochromatin self-interactions.

Our experimental system provides means for manipulating nuclear organization in a reproducible manner, potentially allowing to examine radial positioning decoupled from other features of genome organization. Highlighting the complementary nature of microscopic and genomic methods, our work further suggests a remarkable resilience of genome structure such that large-scale nuclear changes, including chromosome compression and changes in radial positioning, can occur without extensive alteration of functional genome organization.
Conformational dynamics of the HIV-1 envelope glycoprotein from CRF01_AE is associated with susceptibility to antibody-dependent cellular cytotoxicity
Authors: Diaz-Salinas Marco , Chatterjee Debashree , Nayrac Manon , Medjahed Halima , Prevost Jeremie , Pazgier Marzena , Finzi Andres , Munro James
Published: August 2024

The HIV-1 envelope glycoprotein (Env) is expressed at the surface of infected cells and as such can be targeted by non-neutralizing antibodies (nnAbs) that mediate antibody-dependent cellular cytotoxicity (ADCC). Previous single-molecule Forster resonance energy transfer (smFRET) studies demonstrated that Env from clinical isolates predominantly adopt a 'closed' conformation (State 1), which is resistant to nnAbs. After interacting with the cellular receptor CD4, the conformational equilibrium of Env shifts toward States 2 and 3, exposing the coreceptor binding site (CoRBS) and permitting binding of antibodies targeting this site. We showed that the binding of anti-CoRBS Abs enables the engagement of other nnAbs that target the cluster A epitopes on Env. Anti-cluster A nnAbs stabilize an asymmetric Env conformation, State 2A, and have potent ADCC activity. CRF01_AE strains were suggested to be intrinsically susceptible to ADCC mediated by nnAbs. This may be due to the presence of a histidine at position 375, known to shift Env towards more 'open' conformations. In this work, through adaptation of an established smFRET imaging approach, we report that the conformational dynamics of native, unliganded HIV-1CRF01_AE Env indicates frequent sampling of the State 2A conformation. This is in striking contrast with Envs from clades A and B, for example HIV-1JR-FL, which do not transition to State 2A in the absence of ligands. These findings inform on the conformational dynamics of HIV-1CRF01_AE Env, which are relevant for structure-based design of both synthetic inhibitors of receptor binding, and enhancers of ADCC as therapeutic alternatives.
An NADH-controlled gatekeeper of ATP synthase
Authors: Schildhauer Fabian , Ryl Petra SJ , Lauer Simon M , Lenz Swantje , Barlas Ayse Bercin , Ouzounidis Vasileios R , Jeffrey Kate , Marcu Daniel-Cosmin , O'Reilly Francis J , Graziadei Andrea , Stuiver Marchel , Schmidt Kita , Ewers Helge , Spahn Christian MT , Karaca Ezgi , Busch Karl Emanuel , Cheerambathur Dhanya , Schwefel David , Rappsilber Juri
Published: August 2024

ATP fuels crucial cellular processes and is obtained mostly by oxidative phosphorylation (OXPHOS) at the inner mitochondrial membrane. While significant progress has been made in mechanistic understanding of ATP production, critical aspects surrounding its substrate supply logistics are poorly understood. We identify an interaction between mitochondrial apoptosis-inducing factor 1 (AIFM1) and adenylate kinase 2 (AK2) as gatekeeper of ATP synthase. This interaction is NADH-dependent and influenced by glycolysis, linking it to the cell's metabolic state. Genetic interference with AIFM1/AK2 association impedes the ability of Caenorhabditis elegans animals to handle altered metabolic rates and nutrient availability. Together, the results imply AIFM1 as a cellular NADH sensor, placing AK2 next to the OXPHOS complexes for local ADP regeneration as the substrate for ATP synthesis. This metabolic signal relay balances ATP synthase substrate supply against ATP conservation, enabling cells to adapt to fluctuating energy availability, with possible implications for AIFM1-related mitochondrial diseases.
Potassium induces conformational changes in the Sabia virus spike complex
Authors: Cohen-Dvashi Hadas , Katz Michael , Diskin Ron
Published: August 2024

Hemorrhagic fever viruses from the Arenaviridae are a source of concern due to their potential to cause lethal outbreaks and the lack of effective therapeutics. Several Clade-B viruses, which are endemic to the Americas, are pathogens that sporadically infect humans following zoonotic transmission from small rodents. Brazilian hemorrhagic fever, caused by the Sabia virus (SBAV), is one of several such diseases from Clade-B arenaviruses. Despite their importance and the risks they impose, many fundamental questions remain regarding their biology and function. Here, we present the structure of the spike complex from the Sabia virus, which mediates viral attachment and entry to the host cells. Our study reveals two distinct conformational states of the spike, representing its native closed state and an open state that it assumes during cell entry. We show that potassium, in combination with acidic pH, promotes the opening of the spike, which is required for achieving efficient cell entry. This structure further informs us about the architecture of Clade-B arenaviral spikes and how they vary from spikes of other members of the Arenaviridae.
Flexibility and hydration of the Qosite determine multiple pathways for proton transfer in cytochromebc1
Authors: Camilo Sofia R. G. , Arantes Guilherme M.
Published: August 2024

The detailed catalytic activity of cytochromebc1(or respiratory complex III) and the molecular mechanism of the Q cycle remain elusive. At the Qosite, the cycle begins with oxidation of the coenzyme-Q substrate (quinol form) in a bifurcated two-electron transfer to the iron-sulfur (FeS) cluster and the hemebLcenter. The uptake of the two protons released during quinol oxidation is less understood, with one proton likely delivered to the histidine side chain attached to the FeS cluster. Here, we present extensive molecular dynamics simulations with enhanced sampling of side-chain torsions at the Qosite and analyze available sequences and structures of severalbc1homologues to probe the interactions of quinol with potential proton acceptors and identify viable pathways for proton transfer. Our findings reveal that side chains at the Qosite are highly flexible and can adopt multiple conformations. Consequently, the quinol head is also flexible, adopting three distinct binding modes. Two of these modes are proximal to the hemebLand represent reactive conformations capable of electron and proton transfer, while the third, more distal mode likely represents a pre-reactive state, consistent with recent cryo-EM structures ofbc1with bound coenzyme-Q. The Qosite is highly hydrated, with several water molecules bridging interactions between the quinol head and the conserved side chains Tyr147, Glu295, and Tyr297 in cytochromeb(numbering according toR. sphaeroides), facilitating proton transfer. A hydrogen bond network and at least five distinct proton wires are established and possibly transport protons via a Grotthuss mechanism. Asp287 and propionate-A of hemebLin cytochromebare in direct contact with external water and are proposed as the final proton acceptors. The intervening water molecules in these proton wires exhibit low mobility, and some have been resolved in recent experimental structures. These results help to elucidate the intricate molecular mechanism of the Q-cycle and pave the way to a detailed understanding of chemical proton transport in several bioenergetic enzymes that catalyze coenzyme-Q redox reactions.
Proton pump inhibitors modulate esophageal epithelial barrier function and crosstalk with eosinophils
Authors: Gautam Ravi , Lal Megha , Carroll Margaret C , Mrozek Zoe , Trachsel Tina , Beers Jarad , Ruffner Melanie A
Published: August 2024

Background: Eosinophilic esophagitis (EoE) is a chronic allergic disease characterized by esophageal dysfunction, type-2 inflammation, and esophageal eosinophilic infiltrate. While proton pump inhibitor (PPI) therapy is commonly used for EoE management, the underlying mechanism of action remains unclear. Methods: Air-liquid interface culture of esophageal epithelial cells was employed to investigate the impact of the PPI omeprazole on barrier integrity in IL-13-treated cultures. Epithelial chemokine secretion was assessed following stimulation with IL-13 and omeprazole, and the migration of eosinophils from healthy human donors was evaluated using 3 μm pore-sized transwells. A co-culture system of epithelial cells and eosinophils was employed to study chemokine secretion and eosinophil adhesion and activation markers. Results: Omeprazole treatment in the IL-13-treated air-liquid interface (ALI) model resulted in 186 differentially expressed genes and restored barrier integrity compared to ALI treated with IL-13 alone. Omeprazole treatment reduced STAT6 phosphorylation, downregulated calpain 14, and upregulated desmoglein-1 in the IL-13-treated air-liquid interface samples. IL-13-induced upregulation of Eotaxin-3, CXCL10, and periostin, but this was downregulated by omeprazole. Further, the expression of CD11b, CD18, and CD69 was lower on eosinophils from omeprazole-treated epithelial-eosinophil co-cultures, which also had lower levels of eotaxin-3, CXCL10, CCL2, and CCL4. Conclusion: Omeprazole reduced the effects of IL-13 in both the epithelial air-liquid interface model and eosinophil-epithelial co-cultures, reducing barrier dysfunction, chemokine expression, and upregulation of eosinophil adhesion markers.
Inhibiting EZH2 complements steroid effects in Duchenne muscular dystrophy
Authors: Jeon Eun Young , Kwak Yejin , Kang Hyeji , Jin Se Young , Park Soojin , Kim Ryeo Gyeong , Ko Dayoung , Won Jae-Kyung , Cho Anna , Jung Inkyung , Lee Chul-Hwan , Park Jeongbin , Kim Hyun-Young , Chae Jong-Hee , Choi Murim
Published: August 2024

Duchenne muscular dystrophy (DMD) is a devastating X-linked disorder caused by mutations in the dystrophin gene. Despite recent advances in understanding the disease etiology and applying emerging treatment methodologies, glucocorticoid derivatives remain the only general therapeutic option that can slow disease development. However, the precise molecular mechanism of glucocorticoid action remains unclear, and there is still need for additional remedies to complement the treatment. Here, using single-nucleus RNA-sequencing and spatial transcriptome analyses of human and mouse muscles, we investigated pathogenic features in DMD patients and palliative effects of glucocorticoids. Our approach further illuminated the importance of proliferating satellite cells, and revealed increased activity of a signal transduction pathway involving EZH2 in the patient cells. Subsequent administration of EZH2 inhibitors toDmdmutant mice resulted in improved muscle phenotype through maintaining the immune-suppressing effect but overriding the muscle weakness and fibrogenic effects exerted by glucocorticoids. Our analysis reveals pathogenic mechanisms that can be readily targeted by extant therapeutic options for DMD.

Teaser

A survey of DMD tissues in human and mouse suggests EZH2 as a critical factor in DMD satellite cells; its inhibition resulted in better prognosis.
A convenient single-cell newly synthesized transcriptome assay reveals gene expression dynamics during early-stage T-cell activation
Authors: Lyu Jun , Xu Xiaoyan , Chen Chongyi
Published: August 2024

Sequencing newly synthesized transcriptome alongside regular transcriptome in single cells enables the study of gene expression temporal dynamics during rapid chromatin and gene regulation processes. However, existing assays to profile single-cell newly synthesized transcriptome require in-house technical expertise to achieve high cellular throughput, limiting their widespread application. Here, we developed NOTE-seq, a method that simultaneously profiles regular and newly synthesized transcriptomes in single cells. NOTE-seq integrates 4-thiouridine labeling of newly synthesized RNA, thiol-alkylation-based chemical conversion, and a streamlined workflow on the 10X Genomics platform, offering high cellular throughput that is accessible and convenient for regular biology laboratories without specialized single-cell expertise. Using NOTE-seq, we characterized the temporal dynamics of gene expression during early-stage T-cell activation in Jurkat and naïve T cells, identified transcription factors and regulons, and discoveredFli-1as a master transcription factor for gene regulation upon T-cell stimulation. Interestingly, chemotherapeutic topoisomerase inhibitor affectsFli-1level in T cells, indicating potential complications for the immune system.
VRS5 (HvTB1) binds to the promoter of tillering and floral homeotic genes to regulate their expression
Authors: Winkelmolen T. , Colleoni P. , Moscou M. , Hoseinzadeh P. , Oldach K. , Schmidt R.C. , Immink R. , van Esse G.W.
Published: August 2024

Variation in shoot architecture, or tillering, is an important adaptive trait targeted during the domestication of crops. A well-known regulatory factor in shoot architecture is TEOSINTE BRANCHED 1 (TB1). TB1 and its orthologs have a conserved function in integrating environmental signals to regulate axillary branching, or tillering in cereals. The barley ortholog of TB1,VULGARE ROW-TYPE SIX 5(VRS5) does not only regulate tillering, but is also involved in regulating row-type by inhibiting lateral spikelet development. These discoveries predominantly come from genetic studies, but how VRS5 regulates these processes on a molecular level remains largely unknown. By combining transcriptome analysis betweenvrs5and wild type at different developmental stages and DAP-sequencing to locate the genome-wide DNA binding sites of VRS5, we identifiedbona fidetargets of VRS5. We found that VRS5 targets abscisic acid related genes, potentially to inhibit tillering in a conserved way. While later in inflorescence development, row-type geneVRS1and several known floral development genes, like MIKCc type MADS-box genes, are targeted. This study identifies several significant genes for mutational analysis, representing a selection ofbona fidetargets that will contribute to a deeper understanding of the VRS5 network and its role in shaping barley development.
Variant-to-function mapping of late-onset Alzheimer’s disease GWAS signals in human microglial cell models implicatesRTFDC1at theCASS4locus
Authors: Burton Elizabeth A. , Argenziano Mariana , Cook Kieona , Ridler Molly , Lu Sumei , Su Chun , Manduchi Elisabetta , Littleton Sheridan H. , Leonard Michelle E. , Hodge Kenyaita M. , Wang Li-San , Schellenberg Gerard D. , Johnson Matthew E. , Pahl Matthew C. , Pippin James A. , Wells Andrew D. , Anderson Stewart A. , Brown Christopher D. , Grant Struan F.A. , Chesi Alessandra
Published: August 2024

Late-onset Alzheimer’s disease (LOAD) research has principally focused on neurons over the years due to their known role in the production of amyloid beta plaques and neurofibrillary tangles. In contrast, recent genomic studies of LOAD have implicated microglia as culprits of the prolonged inflammation exacerbating the neurodegeneration observed in patient brains. Indeed, recent LOAD genome-wide association studies (GWAS) have reported multiple loci near genes related to microglial function, includingTREM2,ABI3, andCR1. However, GWAS alone cannot pinpoint underlying causal variants or effector genes at such loci, as most signals reside in non-coding regions of the genome and could presumably confer their influence frequently via long-range regulatory interactions. We elected to carry out a combination of ATAC-seq and high-resolution promoter-focused Capture-C in two human microglial cell models (iPSC-derived microglia and HMC3) in order to physically map interactions between LOAD GWAS-implicated candidate causal variants and their corresponding putative effector genes. Notably, we observed consistent evidence that rs6024870 at the GWASCASS4locus contacted the promoter of nearby gene,RTFDC1. We subsequently observed a directionallly consistent decrease inRTFDC1expression with the the protective minor A allele of rs6024870 via both luciferase assays in HMC3 cells and expression studies in primary human microglia. Through CRISPR-Cas9-mediated deletion of the putative regulatory region harboring rs6024870 in HMC3 cells, we observed increased pro-inflammatory cytokine secretion and decreased DNA double strand break repair related, at least in part, toRTFDC1expression levels. Our variant-to-function approach therefore reveals that the rs6024870-harboring regulatory element at the LOAD ‘CASS4’GWAS locus influences both microglial inflammatory capacity and DNA damage resolution, along with cumulative evidence implicatingRTFDC1as a novel candidate effector gene.
3D scanning measurement method for seed size research
Authors: Málik-Roffa Hajnalka , Málik Tamás , Tőzsér Dávid , Tóthmérész Béla
Published: August 2024

Premise

Traditionally used seed size and surface area calculation techniques are often inaccurate or destructive. This is especially relevant for organisms with irregular shapes. It is believed that three-dimensional (3D) printing and scanning could also be disruptive technologies. However, these methods hold immense potential for ecology and evolution sciences, which have a longstanding tradition of inventing and creating objects for research, education, and outreach.

Methods and Results

We describe how to 3D scan seeds reproducible on an inexpensive structured light based commodity 3D scanner. We then compared novel 3D and traditional 2D approaches by reproducibility and timing.

Conclusions

This study demonstrates how 3D scanning methods are easily applicable to the quantification of seed shape and providing an easy entry into it.
Automatic crystal identification for crystallography:a comparison between direct methods and artificial intelligence strategies
Authors: Lages Camila F A , Nicoleti Vitoria Y , Correa Maicon R , Mausbach Paulo , Ramos Felipe C , de Araujo Evandro A , Nascimento Andrey Z , Miqueles Eduardo X
Published: August 2024

Crystallography is an important, and well-established technique for assessing crystalline atomic structure of a wide variety of materials. An auxiliary optical microscope instrument is used to perform the data acquisition at the manaca beamline, a crystallography experimental station from sirius (the 4th generation Brazilian synchrotron). The importance of detecting these crystals in the auxiliary image is to define the best order in which each crystal will be measured by the incident beam since each one of them must be measured only once; improving beamtime and maximizing user experience at the facility. Detecting rough crystal positions from these auxiliary images is a scientific computing task, which could benefit from some open artificial intelligence tools. This manuscript compares a real-time object detection interface (Yolo) with a new, simple and effective strategy obtained from a particular partial differential equation.
Generation of humanized mouse models to support therapeutic development for SYNGAP1 and STXBP1 disorders
Authors: Felix Alex J. , Wilson Taryn , Randell Rani , Marotta Nicolas , Uchida Keita , Boland Michael J. , Davidson Beverly L. , Prosser Benjamin L.
Published: August 2024

Heterozygous variants inSYNGAP1andSTXBP1lead to distinct neurodevelopmental disorders caused by haploinsufficient levels of post-synaptic SYNGAP1 and pre-synaptic STXBP1, which are critical for normal synaptic function. While several gene-targeted therapeutic approaches have proven efficaciousin vitro, these often target regions of the human gene that are not conserved in rodents, hindering the pre-clinical development of these compounds and their transition to the clinic. To overcome this limitation, here we generate and characterizeSyngap1andStxbp1humanized mouse models in which we replaced the mouseSyngap1andStxbp1gene, respectively, with the human counterpart, including regulatory and non-coding regions. Fully humanizedSyngap1mice present normal viability and can be successfully crossed with currently availableSyngap1haploinsufficiency mouse models to generateSyngap1humanized haploinsufficient mice.Stxbp1mice were successfully humanized, yet exhibit impaired viability (particularly males) and reduced STXBP1 protein abundance. Mouse viability could be improved by outcrossing this model to other mouse strains, whileStxbp1humanized females and hybrid mice can be used to evaluate target engagement of human-specific therapeutics. Overall, these humanized mouse models represent a broadly available tool to further pre-clinical therapeutic development for SYNGAP1 and STXBP1 disorders.
Correlated evolutionary rates reveal novel components and cross-compartment connectivity in plant proteostasis systems
Authors: Gatts Tony C , Rehmann Elizabeth A , Sloan Daniel B , Forsythe Evan S
Published: August 2024

Plant cells rely on an interconnected network of proteins interacting at many levels (e.g. physical enzyme complexes, gene regulatory modules, and biosynthetic pathways). Pairs of proteins that interact at any of these levels have been shown to exhibit phylogenetic signatures of evolutionary rate covariation (ERC), providing a basis for detecting functional interactions among proteins. Here, we apply ERCnet, a bioinformatic tool for performing genome-scale ERC analyses, to predict a plant protein-protein interactome network. We find a clustered set of proteins that exhibit strong signatures of ERC with the plastid caseinolytic protease (Clp) and other plastid proteostasis components, thus forming a functional module within the network. In addition to including proteins with known or predicted functions in protein import, transcription, translation, and degradation in plastids, the module also includes proteins with previously unknown molecular function, thus providing evidence that these proteins may contribute to plastid proteostasis in novel ways. Perhaps the most surprising members of this module are a set of proteins that are not thought to localize to the plastid at all. These proteins include a mitochondrial-localized pentatricopeptide repeat (PPR) protein with genetic evidence of interaction with the mitochondrial Clp system and two nuclear-localized actin-related proteins involved in chromatin remodeling and epigenetic regulation of nuclear genes. We speculate that these non-plastid-localized proteins act as mediators of organellar crosstalk and retrograde signaling of cellular proteostasis status in plants. In summary, our results highlight the connected nature of plant proteostasis systems and point to a promising set of novel proteostasis protein candidates.
Regulation of angiogenesis by signal sequence-derived peptides
Authors: Ghim Mean , Wei Linyan , Jung Jae-Joon , The Erlinda , Kukreja Gunjan , Neishabouri Afarin , Ahmad Azmi A. , Raza M. Zawwad , Golbazi Arvene , Hedayatyanfard Keshvad , Nie Lei , Zhang Jiasheng , Sadeghi Mehran M.
Published: August 2024

Background

The neuropilin-like, Discoidin, CUB and LCCL domain containing 2 (DCBLD2) is a transmembrane protein with an unusually long signal sequence (SS) composed of N-terminal (N) and C-terminal (C) subdomains, separated by a transition (tra) subdomain. DCBLD2 interacts with VEGFR-2 and regulates VEGF-induced endothelial cell signaling, proliferation and migration, as well as angiogenesis. The exact mechanisms by which DCBLD2 interacts with VEGFR2 to modulate VEGF signaling remain unclear.

Methods

Searching for VEGFR2 interacting DCBLD2 domains, we generated various constructs containing different DCBLD2 domain combinations and conducted co-immunoprecipitation and signaling studies in HEK 293T and endothelial cells. Several peptides were synthesized based on the identified domain, and their effect on VEGF signaling was assessed in vitro in cell culture and in vivo using matrigel plug and corneal micropocket assays. The effect of the lead peptide was further evaluated using a murine hindlimb ischemia model.

Results

DCBLD2 SS interacted with VEGFR2 and promoted VEGF signaling. SS was not cleaved in the mature DCBLD2 and its hydrophobic transmembrane ‘traC’ segment, but not the ‘N’ subdomain, was involved in DCBLD2-VEGFR2 interaction. The smallest unit in DCBLD2 SS that interacts with VEGFR2 was the L5VL5 sequence. Even after the central valine was removed, the L10 sequence mimicked the DCBLD2 SS traC’s effect on VEGF-signaling, while shorter or longer poly-leucine sequences were less effective. Finally, a synthetic traC peptide enhanced VEGF signaling in vitro, promoted VEGF-induced angiogenesis in vivo, and improved blood flow recovery following hindlimb ischemia.

Conclusion

DCBLD2 SS along with its derivative peptides can promote VEGFR2 signaling and angiogenesis. Synthetic peptides based on DCBLD2 SS hold promise as therapeutic agents for regulating angiogenesis. Importantly these findings refine the traditional view of signal sequences as mere targeting elements, revealing a role in cellular signaling. This opens new avenues for research and therapeutic strategies.
Detecting behavioural oscillations with increased sensitivity: A modification of Brookshire’s (2022) AR-surrogate method
Authors: Harris Anthony M. , Beale Henry A.
Published: August 2024

A core challenge of cognitive neuroscience is to understand how cognition changes over time within the same individual. For example, the tendency for behavioural responses in a range of cognitive domains to oscillate over time has been studied extensively. Recently, however, the phenomenon of behavioural oscillations has been called into question by indications that past findings might reflect aperiodic temporal structure rather than true oscillations. Brookshire (2022) proposed methods to control for aperiodic temporal structure while examining oscillations in behavioural time-courses and found no evidence of behavioural oscillations in reanalyses of four published datasets. However, Brookshire’s (2022) method has been criticised for having low sensitivity to detect effects of realistic magnitude, so it is currently unclear whether these findings suggest that behavioural oscillations are not present in these and perhaps many other datasets, or whether they are false negatives. Here, we present a modification of Brookshire’s (2022) AR-surrogate method with increased sensitivity to detect effects of realistic magnitude, adequate control of false positives, and other desirable properties such as the ability to increase statistical power by adding more participants. Using this method, we reanalyse the same publicly available datasets and show significant behavioural oscillations in each of them, suggesting oscillations in behaviour are a robust phenomenon upon which to draw theoretical inferences. The participant-level AR-surrogate method is currently the most sensitive method available for analysing behavioural oscillations while controlling for the contribution of aperiodic data fluctuations.
Boundaries in the eyes: measure event segmentation during naturalistic video watching using eye tracking
Authors: Li Jiashen , Cheng Zhengyue , Hao Xin , Liu Wei
Published: August 2024

During naturalistic information processing, individuals spontaneously segment their continuous experiences into discrete events, a phenomenon known as event segmentation. Traditional methods for assessing this process, which include subjective reports and neuroimaging techniques, often disrupt real-time segmentation or are costly and time intensive. Our study investigated the potential of measuring event segmentation by recording and analyzing eye movements while participants view naturalistic videos. We collected eye movement data from healthy young adults as they watched commercial films (N=104), or online Science, Technology, Engineering, and Mathematics (STEM) educational courses (N=44). We analyzed changes in pupil size and eye movement speed near event boundaries and employed inter-subject correlation analysis (ISC) and hidden Markov models (HMM) to identify patterns indicative of event segmentation. We observed that both the speed of eye movements and pupil size dynamically responded to event boundaries, exhibiting heightened sensitivity to high-strength boundaries. Our analyses further revealed that event boundaries synchronized eye movements across participants. These boundaries, can be effectively identified by HMM, yielded higher within-event similarity values and aligned with human-annotated boundaries. Importantly, HMM-based event segmentation metrics responded to experimental manipulations and predicted learning outcomes. This study provided a comprehensive computational framework for measuring event segmentation using eye-tracking. With the widespread accessibility of low-cost eye-tracking devices, the ability to measure event segmentation from eye movement data promises to deepen our understanding of this process in diverse real-world settings.
RNA quality control by CCR4 safeguards chromatin integrity and centromere function inArabidopsis
Authors: Shimada Atsushi , Saze Hidetoshi
Published: August 2024

The centromere is a pivotal chromatin domain that ensures accurate chromosome segregation during cell division. However, the epigenome regulation of the centromere and its impact on centromere function remain largely elusive. Here in the model plantArabidopsis, we show that CCR4, the catalytic subunit of the RNA deadenylation complex CCR4-NOT, is essential for maintenance of the centromere epigenome and chromosome integrity. We demonstrate that CCR4 is involved in shortening of the poly(A) tails of transcripts originated from centromeric transposons and repeats, thereby promoting the production of small interfering RNAs (siRNAs). The CCR4-dependent siRNAs guide non-CG DNA methylation at centromere repeats, and CCR4 cooperates with canonical DNA methylation pathways to enhance centromeric H3K9 methylation and ensure mitotic chromosome stability. Our study illustrates the crucial role of RNA quality control in RNA interference and reveals the elaborate mechanism that safeguards plant centromeres through epigenomic regulation.
Morphological criteria for staging near-hatching embryos of the domesticated Mallard (Anas platyrhynchos) and Swan Goose (Anser cygnoides)
Authors: Arnaout Bassel , Brzezinski Kaylen , Steventon Benjamin , Field Daniel J.
Published: August 2024

Studying waterfowl embryology necessitates reliable and precise staging tables. These are descriptions of embryonic features throughout development from fertilisation to hatching stage, which are used to approximate the extent of embryonic development. Waterfowl staging tables have previously been established based on morphological features from fertilisation to approximately ten days before hatching. Embryonic changes over the final ten days of pre-hatching development have also been documented and proposed as useful staging criteria. However, the reliability of these changes as useful staging criteria across different waterfowl breeds has not been fully examined. To examine the reliability of these criteria, which focus on size changes of the bill and middle toes, for staging near-hatching embryos, we examined twenty-seven Mallard and Swan Geese embryos and measured their beak and toe-length sizes. We compared our results with previously published data, and found that size variation across breeds within the same developmental stage is equivalent to within-breed variation across different stages, suggesting limited reliability of beak and middle toe length sizes for staging waterfowl embryos. Consequently, we devised novel staging criteria for waterfowl based on four easily measurable morphological traits and show that these criteria allow correct stage identification with over 70% accuracy. Our results highlight the importance of quantifying accuracy for developing reliable embryonic staging tables.
Identifying novel chemical matter against the Chikungunya virus nsP3 macrodomain through crystallographic fragment screening
Authors: Aschenbrenner Jasmin C. , de Godoy Andre Schutzer , Fairhead Michael , Tomlinson Charles W.E. , Winokan Max , Balcomb Blake H. , Capkin Eda , Chandran Anu V. , Golding Mathew , Koekemoer Lizbe , Lithgo Ryan M. , Marples Peter G. , Ni Xiaomin , Thompson Warren , Wild Conor , Xavier Mary-Ann E. , Fearon Daren , von Delft Frank
Published: August 2024

Chikungunya virus (CHIKV) causes severe fever, rash and debilitating joint pain that can last for months1,2or even years. Millions of people have been infected with CHIKV, mostly in low and middle-income countries, and the virus continues to spread into new areas due to the geographical expansion of its mosquito hosts. Its genome encodes a macrodomain, which functions as an ADP-ribosyl hydrolase, removing ADPr from viral and host-cell proteins interfering with the innate immune response. Mutational studies have shown that the CHIKV nsP3 macrodomain is necessary for viral replication, making it a potential target for the development of antiviral therapeutics. We, therefore, performed a high-throughput crystallographic fragment screen against the CHIKV nsP3 macrodomain, yielding 109 fragment hits covering the ADPr-binding site and two adjacent subsites that are absent in the homologous macrodomain of SARS-CoV-2 but may be present in other alphaviruses, such as Venezuelan equine encephalitis virus (VEEV) and eastern equine encephalitis virus (EEEV). Finally, a subset of overlapping fragments was used to manually design three fragment merges covering the adenine and oxyanion subsites. The rich dataset of chemical matter and structural information discovered from this fragment screen is publicly available and can be used as a starting point for developing a CHIKV nsP3 macrodomain inhibitor.