bioRxiv Science⌕ Search

Biology subjects

Dai, W.

Publications and source records attributed to Dai, W..

At least 37 records · Page 2Linked to original sources

Deep learning guided discovery of antibacterial polymeric nanoparticles

Drug-resistant bacterial infections pose a serious threat to global health, driving the development of novel antibacterial strategies beyond classic antibiotics. Self-assembled polymeric nanoparticles (SANPs) have emerged as promising candidates due to their unique structural properties, which enhance membrane interaction, drug loading, and therapeutic accumulation. The structural diversity and tunable properties of SANPs provide a flexible design platform for antibacterial applications. However, navigating the vast chemical space of SANPs remains a significant challenge due to the complex structure-activity relationships. To address this, we developed PolyCLOVER, a deep learning-guided framework for the efficient discovery of antibacterial SANPs. The framework integrates multi-stage self-supervised learning, active learning, and high-throughput experimentation to iteratively guide the discovery of SANPs with potent antibacterial activity and low toxicity. Through this framework, we identified three potent SANPs within a combinatorial library of [~]100,000 poly({beta}-amino ester)s, with minimum inhibitory concentrations of 4 g/mL and 8 g/mL against clinically multidrug-resistant S. aureus and A. baumannii, respectively. Moreover, they can also act as antibiotic carriers, restoring the sensitivity of pathogens to penicillin G. In vivo studies demonstrated their therapeutic efficacy both as monotherapies and in combination therapies with antibiotics. These results demonstrate PolyCLOVER as a powerful framework for discovering multi-functional SANPs to combat antibiotic resistance.

microbiology↗

Modification and validation of a GAD-GFP mouse line without accelerated aging-related hearing loss.

GABAergic neurons in the inferior colliculus (IC) play a crucial role in auditory processing by extracting specific features of sounds (Ono et al., 2005). The Gad67-GFP mouse model developed by Tamamaki et al. in 2003 on a Swiss background facilitates studying these neurons by using a green fluorescent protein that is expressed endogenously via the GAD67 promoter. Unfortunately, this mouse suffers from accelerated aging-related hearing loss, limiting its utility in studying the auditory system. Here, we report the results of an 8-generation backcross of this line onto CBA/CaJ mice, which produces mice with stable low-threshold hearing while retaining GFP expression in GAD+ neurons. Additionally, this study investigates mechanisms that underlie hearing loss in the Gad67-GFP mouse model by focusing specifically on cochlear hair cells (HCs) and ribbon synapses, which may contribute to both model-specific hearing loss and clinical disorders like presbycusis. Findings revealed the newly generated F1 mouse model that resulted from the Gad67-GFP x CBA/CaJ backcross maintained better hearing thresholds when compared to ABR data for Gad67 and Swiss mice and very closely resembled those of the CBA/CaJ mice, mirroring progression of presbycusis in humans. Additionally, all morphological changes observed in cochlear structure correlated to ABR thresholds. F1 mice continued maintained expression of the GAD67 promoter in the IC via immunostaining.

neuroscience↗

In Situ Detection of α-Synuclein Seeding Activity Using Quiescent Seed Amplification Assay

The prion-like propagation of proteopathic -synuclein (Syn) "seeds" is a central pathogenic mechanism in synucleinopathies. However, methods to directly visualize this "seeding activity" within intact tissue are lacking. Conventional antibody-based techniques provide only static snapshots of pathological deposits, while sensitive seed amplification assays lose spatial information via mechanical fragmentation. Here, we introduce the Syn Quiescent Seed Amplification Assay (QSAA), a method for specific, robust, in situ amplification of Syn seeds directly within tissue sections. QSAA demonstrates superior sensitivity and earlier detection than pS129 and ThS staining, revealing widespread seeding in a PD mouse model, and successfully identifies endogenous seeds in archival human brains. By integrating with immunofluorescence, IF-QSAA uncovered microglial seeding activity and established that pS129 phosphorylation and seeding activity represent distinct pathological states. This offers unprecedented spatial insights into synucleinopathy pathogenesis, holding significant promises for early diagnosis and therapeutic evaluation.

neuroscience↗

Mechanics of the Spatiotemporal Evolution of Sulcal Pits in the Folding Brain

Understanding the development of complex brain surface morphologies during the fetal stage is essential for uncovering mechanisms behind brain disorders linked to abnormal cortical folding. However, knowledge of the spatiotemporal evolution of fetal brain landmarks is limited due to the lack of longitudinal data capturing multiple timepoints for individual brains. In this study, we develop and validate a true-scale, image-based mechanical model to explore the spatiotemporal evolution of brain sulcal pits in individual fetal brains. Our model, constructed using magnetic resonance imaging (MRI) scans from the first timepoint of longitudinal data, predicts the brains surface morphology by comparing the distribution of sulcal pits between predicted models and MRI scans from a later timepoint. This dynamic model elucidates how a smooth fetal brain with primary folds evolves to form secondary and tertiary folds. Our results align with imaging data, showing that sulcal pits are stable during brain development and can serve as key markers linking prenatal and postnatal brain characteristics. The model provides a robust platform to study the evolution of sulcal pits in both healthy and disordered brains, which is crucial as altered sulcal pits patterns are seen in disorders such as autism spectrum disorder (ASD), polymicrogyria, down syndrome, and agenesis of the corpus callosum. This research represents a significant advancement in understanding fetal brain development and its connection to disorders that manifest as abnormal sulcal pit patterns later in life.

bioengineering↗

A Novel Optimized Silver Nitrate Staining Method for Visualizing the Osteocyte Lacuno-Canalicular System

A 50% silver nitrate solution is commonly used to stain and characterize the osteocyte lacuno-canalicular system (LCS) in bone biology research. However, variability in reagent concentrations and types, along with inconsistent staining procedures, have limited the broader application of this method in osteocyte research. In this study, we present a novel optimized silver nitrate staining technique aimed at addressing these limitations. This new method utilizes a 1 mol/L silver nitrate solution in combination with a type-B gelatin-formic acid solution at various concentrations (0.05%-0.5% gelatin and 0.05%-5% formic acid, or 1%-2% gelatin and 0.1%-2% formic acid) in volume ratios of 4:1, 2:1, or 1:1, or a 0.5 mol/L silver nitrate solution at a 4:1 ratio. The staining process is carried out for 1 hour under ultraviolet light or 90 minutes under regular room light (or dark), followed by washing with Milli-Q water to terminate the reaction. We applied this new method to stain the osteocyte LCS in bone samples from different species and pathological bone models. The technique consistently produced clear, distinct staining patterns across all samples. Moreover, our novel method revealed a greater number of LCS compared to the traditional 50% silver nitrate solution. This suggests that the commonly used 50% silver nitrate method may disrupt or inadequately reveal the LCS in certain species, potentially leading to an underestimation of LCS density and number. In conclusion, our novel silver nitrate staining method provides a simpler and more cost-effective alternative to the traditional technique. By offering a more accurate and comprehensive analysis of the LCS across species, this approach has the potential to advance research on osteocyte morphogenesis, as well as the functional and evolutionary adaptations of the osteocyte LCS across different taxa.

pathology↗

Local ionic conditions modulate the aggregation propensity and influence the structural polymorphism of alpha-synuclein

Parkinsons Disease (PD) is characterized by the aggregation of alpha-synuclein (aSyn), a presynaptic protein that transitions from a disordered monomer into beta-sheet rich amyloid fibrils. The precise triggers and mechanisms underlying aSyn misfolding and aggregation remain unclear, hindering the development of effective therapeutics. Monomeric aSyn is an intrinsically disordered protein (IDP) with high conformational flexibility. Local environmental factors, such as ion concentrations, can influence the conformational ensemble of aSyn, impacting its aggregation propensity and resulting in fibril polymorphism. In this study, we explore the impact of physiologically relevant ions, mainly Ca2+ and Na+, on the aggregation kinetics, monomer structural dynamics, and fibril polymorphism of aSyn. Using ThT fluorescence assays, we demonstrate that all ions accelerate aSyn aggregation, with Ca2+ having the most significant effect. Using Heteronuclear Single Quantum Correlation Nuclear Magnetic Resonance (1H-15NHSQC NMR) spectroscopy, we validate the specific binding of Ca2+ ions at the C-terminus, whereas Na+ ions display non-specific interactions along the sequence of aSyn. Small-angle neutron scattering (SANS) and hydrogen-deuterium exchange mass spectrometry (HDX-MS) further reveal that Na+ and Ca2+ induce distinct conformational changes in the aSyn monomer, with Na+ leading to more extended structures and Ca2+ promoting a moderate extension of the protein. Molecular dynamics simulations (MD) corroborate these findings, showing that Na+ ions increase the proteins extension, particularly between the non-amyloid beta component (NAC) region and the C-terminus, whereas Ca2+ ions bias the ensemble towards a more moderately elongated structure. Using MD, we further investigate the local environment and in particular the solvent effect and show the water persistence times in the hydration shell are also increased in the presence of Ca2+ ions, indicating that the aggregation propensity of the monomer is due to a combination of conformational bias of the monomer and solvent mobility. Atomic force microscopy (AFM) of aSyn fibrils formed under these different ionic conditions reveal distinct fibril polymorphs, suggesting that ion-induced conformational biases in the monomer contribute to the diversity of fibril structures. Collectively, these findings underscore the pivotal influence of the local ionic milieu in shaping the structure and aggregation propensity of aSyn, thus offering valuable insights into the molecular underpinnings of PD and potential therapeutic avenues aimed at manipulating aSyn conformational dynamics.

biophysics↗

Monitoring of Cell-free Human Papillomavirus DNA in Metastatic or Recurrent Cervical Cancer: Clinical Significance and Treatment Implications

PurposeMonitoring of circulating human papillomavirus (HPV) cell-free DNA (cfDNA) is a minimally invasive approach for surveillance in HPV-associated cancers, particularly cervical cancer. The aim of this study was to monitor circulating HPV cfDNA levels in patients with recurrent or metastatic cervical cancer during treatment and follow-up, assessing the utility of HPV cfDNA as a tumor marker for disease surveillance and in guiding clinical treatment decisions. Experimental DesignIn this prospective pilot observational study, levels of HPV cfDNA in serum samples from 28 patients with recurrent or metastatic HPV-positive cervical cancer were measured via digital droplet polymerase chain reaction. Results for HPV cfDNA levels were matched to clinical outcomes and to serum levels of squamous cell carcinoma antigen (SCC-Ag) to assess the clinical potential of HPV cfDNA as a tumor marker. ResultsHPV cfDNA was detected in all 28 patients (100% detection rate). Notably, median baseline HPV cfDNA levels varied according to the metastatic pattern observed in individual patients (P=0.019). Patients with a combined multi-metastatic pattern (lymph node + hematogenous {+/-} diffuse serosal metastasis) exhibited a higher median baseline HPV cfDNA level compared to those with a single-metastasis pattern (local recurrence, lymph node metastasis, or hematogenous metastasis) (P = 0.003). All participants exhibited changes in HPV cfDNA levels over a median monitoring period of 2 months (range 0.3 to 16.9 months) prior to evaluations for treatment response or disease progression. Among 26 patients initially diagnosed with squamous cell cervical cancer, the positivity rate was 100% for HPV cfDNA and 69.2% for SCC-Ag (P = 0.004, 95% confidence interval, 0-0.391). Among 20 patients longitudinally monitored for squamous cell cervical cancer, the concordance with changes in disease status was 90% for HPV cfDNA and 50% for SCC-Ag (P = 0.014, 95% confidence interval, 0.022-0.621). ConclusionsHPV cfDNA is a promising tumor marker for HPV-positive cervical cancer. In the context of precision medicine, HPV cfDNA is poised to play an increasingly pivotal role in monitoring treatment efficacy, providing valuable insights into disease progression, and guiding clinical decisions.

cancer biology↗

Variation in moment-to-moment brain state engagement changes across development and contributes to individual differences in executive function

Neural variability, or variation in brain signals, facilitates dynamic brain responses to ongoing demands. This flexibility is important during development from childhood to young adulthood, a period characterized by rapid changes in experience. However, little is known about how variability in the engagement of recurring brain states changes during development. Such investigations would require the continuous assessment of multiple brain states concurrently. Here, we leverage a new computational framework to study state engagement variability (SEV) during development. A consistent pattern of SEV changing with age was identified across cross-sectional and longitudinal datasets (N>3000). SEV developmental trajectories stabilize around mid-adolescence, with timing varying by sex and brain state. SEV successfully predicts executive function (EF) in youths from an independent dataset. Worse EF is further linked to alterations in SEV development. These converging findings suggest SEV changes over development, allowing individuals to flexibly recruit various brain states to meet evolving needs.

neuroscience↗

A near-complete genome assembly of the bearded dragon Pogona vitticeps provides insights into the origin of Pogona sex chromosomes

BackgroundThe agamid dragon lizard Pogona vitticeps is one of the most popular domesticated reptiles to be kept as pets worldwide. The capacity of breeding in captivity also makes it emerging as a model species for a range of scientific research, especially for the studies of sex chromosome origin and sex determination mechanisms. ResultsBy leveraging the CycloneSEQ and DNBSEQ sequencing technologies, we conducted whole genome and long-range sequencing for a captive-bred ZZ male to construct a chromosome-scale reference genome for P. vitticeps. The new reference genome is [~]1.8 Gb in length, with a contig N50 of 202.5 Mb and all contigs anchored onto 16 chromosomes. Genome annotation assisted by long-read RNA sequencing greatly expanded the P. vitticeps lncRNA catalog. With the chromosome-scale genome, we were able to characterize the whole Z sex chromosome for the first time. We found that over 80% of the Z chromosome remains as pseudo-autosomal region (PAR) where recombination is not suppressed. The sexually differentiated region (SDR) is small and occupied mostly by transposons, yet it aggregates genes involved in male development, such as AMH, AMHR2 and BMPR1A. Finally, by tracking the evolutionary origin and developmental expression of the SDR genes, we proposed a model for the origin of P. vitticeps sex chromosomes which considered the Z-linked AMH as the master sex-determining gene. ConclusionsOur study provides novel insights into the sex chromosome origin and sex determination of this model lizard. The near-complete P. vitticeps reference genome will also benefit future study of amniote evolution and may facilitate genome-assisted breeding.

genomics↗

Increased burden of rare risk variants across gene expression networks predisposes to sporadic Parkinson's disease

Alpha-synuclein (Syn) is an intrinsically disordered protein that accumulates in the brains of patients with Parkinsons disease and forms intraneuronal inclusions called Lewy Bodies. While the mechanism underlying the dysregulation of Syn in Parkinsons disease is unclear, it is thought that prionoid cell-to-cell propagation of Syn has an important role. Through a high throughput screen, we recently identified 38 genes whose knock down modulates Syn propagation. Follow up experiments were undertaken for two of those genes, TAX1BP1 and ADAMTS19, to study the mechanism with which they regulate Syn homeostasis. We used a recently developed M17D neuroblastoma cell line expressing triple mutant (E35K+E46K+E61K) "3K" Syn under doxycycline induction. 3K Syn spontaneously forms inclusions that show ultrastructural similarities to Lewy Bodies. Experiments using that cell line showed that TAX1BP1 and ADAMTS19 regulate how Syn interacts with lipids and phase separates into inclusions, respectively, adding to the growing body of evidence implicating those processes in Parkinsons disease. Through RNA sequencing, we identified several genes that are differentially expressed after knock-down of TAX1BP1 or ADAMTS19. Burden analysis revealed that those differentially expressed genes (DEGs) carry an increased frequency of rare risk variants in Parkinsons disease patients versus healthy controls, an effect that was independently replicated across two separate cohorts (GP2 and AMP-PD). Weighted gene co-expression network analysis (WGCNA) showed that the DEGs cluster within modules in regions of the brain that develop high degrees of Syn pathology (basal ganglia, cortex). We propose a novel model for the genetic architecture of sporadic Parkinsons disease: increased burden of risk variants across genetic networks dysregulates pathways underlying Syn homeostasis, thereby leading to pathology and neurodegeneration.

cell biology↗

Parallel Analyses by Mass Spectrometry (MS) and Reverse Phase Protein Array (RPPA) Reveal Complementary Proteomic Profiles in Triple-Negative Breast Cancer (TNBC) Patient Tissues and Cell Cultures

High-plex proteomic technologies have made substantial contributions to mechanism studies and biomarker discovery in complex diseases, particularly cancer. Despite technological advancements, inherent limitations in individual proteomic approaches persist, impeding the achievement of comprehensive quantitative insights into the proteome. In this study, we employed two widely used proteomic technologies, Mass Spectrometry (MS) and Reverse Phase Protein Array (RPPA) to analyze identical samples, aiming to systematically assess the outcomes and performance of the different technologies. Additionally, we sought to establish an integrated workflow by combining these two proteomic approaches to augment the coverage of protein targets for discovery purposes. We used fresh frozen tissue samples from triple-negative breast cancer (TNBC) and cell line samples to evaluate both technologies and implement this dual-proteomic strategy. Using a single-step protein denaturation and extraction protocol, protein samples were subjected to reverse phase chromatography (LC) followed by electrospray ionization (ESI)-mediated MS/MS for proteomic profiling. Concurrently, identical sample aliquots were analyzed by RPPA for profiling of over 300 proteins and phosphoproteins that are in key signaling pathways or druggable targets in cancer. Both proteomic methods demonstrated the expected ability to differentiate samples by groups, revealing distinct proteomic patterns under various experimental conditions, albeit with minimal overlap in identified targets. Mechanism-based analysis uncovered divergent biological processes identified with the two proteomic technologies, capitalizing on their complementary exploratory potential.

cancer biology↗

Structural and Biophysical Dynamics of Fungal Plasma Membrane Proteins and Implications for Echinocandin Action in Candida glabrata

Fungal plasma membrane proteins represent key therapeutic targets for antifungal agents, yet their structure and spatial distribution in the native context remain poorly characterized. Herein, we employ an integrative multimodal approach to elucidate the structural and functional organization of plasma membrane protein complexes in Candida glabrata, focusing on prominent and essential membrane proteins, the polysaccharide synthase {beta}-(1,3)-glucan synthase (GS) and the proton pump Pma1. Cryo-electron tomography (cryo-ET) and live cell imaging reveal that GS and Pma1 are heterogeneously distributed into distinct plasma membrane microdomains. Treatment with caspofungin, an echinocandin antifungal that targets GS, alters the plasma membrane and disrupts the native distribution of GS and Pma1. Based on these findings, we propose a model for echinocandin action that considers how drug interactions with the plasma membrane environment lead to inhibition of GS. Our work underscores the importance of interrogating the structural and dynamic characteristics of fungal plasma membrane proteins in situ to understand function and facilitate precisely targeted development of novel antifungal therapies.

cell biology↗

Targeting oncogenic K-Ras mutants with a small-molecule degrader through Nedd4-1

K-Ras mutations represent a most prevalent oncogenic alteration in human cancers. Despite of tremendous efforts, it remains a big challenge to develop inhibitors that can target the oncogenic K-Ras mutants, especially mutants without specific active or charged side chains such as K-RasG12V. Here, taking advantage of our previous finding that Nedd4-1 is a bona fide E3 ubiquitin ligase for wild-type Ras proteins, we developed a compound XMU-MP-9 that can promote ubiquitination and degradation of various K-Ras mutants including K-RasG12V, and significantly inhibit proliferation and tumor development of K-Ras mutant harboring cells. Mechanistically, XMU-MP-9 acts as a bifunctional compound to bind the C2 domain of Nedd4-1 and an allosteric site of K-Ras to enhance Nedd4-1 and K-Ras interaction, and to induce a conformational change of Nedd4-1/K-Ras complex to allow Nedd4-1 targeting K128 of K-Ras for ubiquitination. Hence, our study presents a robust strategy to develop small-molecule degrader of K-Ras mutants, and also sheds light on the development of small-molecule degraders for H-Ras and N-Ras mutants.

cancer biology↗

ΔNp63-restricted viral mimicry response impedes cancer cell viability and remodels tumor microenvironment in esophageal squamous cell carcinoma

Tumor protein p63 isoform {Delta}Np63 plays roles in the squamous epithelium and squamous cell carcinomas (SCCs), including esophageal SCC (ESCC). By integrating data from cell lines and our latest patient-derived organoid cultures, derived xenograft models, and clinical sample transcriptomic analyses, we identified a novel and robust oncogenic role of {Delta}Np63 in ESCC. We showed that {Delta}Np63 maintains the repression of cancer cell endogenous retrotransposon expression and cellular double-stranded RNA sensing. These subsequently lead to a restricted cancer cell viral mimicry response and suppressed induction of tumor-suppressive type I interferon (IFN-I) signaling through the regulations of Signal transducer and activator of transcription 1, Interferon regulatory factor 1, and cGAS-STING pathway. The cancer cell {Delta}Np63-IFN-I signaling axis affects both the cancer cell and tumor-infiltrating immune cell (TIIC) compartments. In cancer cells, depletion of {Delta}Np63 resulted in reduced cell viability. {Delta}Np63 expression is negatively associated with the anticancer responses to viral mimicry booster treatments targeting cancer cells. In the tumor microenvironment, cancer cell TP63 expression negatively correlates with multiple TIIC signatures in ESCC clinical samples. {Delta}Np63 depletion leads to increased cancer cell antigen presentation molecule expression and enhanced recruitment and reprogramming of tumor-infiltrating myeloid cells. Similar IFN-I signaling and TIIC signature association with {Delta}Np63 were also observed in lung SCC. These results support the potential application of {Delta}Np63 as a therapeutic target and a biomarker to guide candidate anticancer treatments exploring viral mimicry responses.

cancer biology↗

Machine learning-based spike sorting reveals how subneuronal concentrations of monomeric Tau cause a loss in excitatory postsynaptic currents in hippocampal neurons

Extracellular recordings of neuronal activity constitute a powerful tool for investigating the intricate dynamics of neural networks and the activity of individual neurons. Microelectrode arrays (MEAs) allow for recordings with a high electrode count, ranging from 10s to 1000s, generating extensive datasets of neuronal information. Furthermore, MEAs capture extracellular field potentials from cultured cells, resulting in highly complex neuronal signals that necessitate precise spike sorting for meaningful data extraction. Nevertheless, conventional spike sorting methods face limitations in recognising diverse spike shapes, thereby constraining the full utilisation of the rich dataset acquired from MEA recordings. To overcome these limitations, we have developed a machine learning algorithm, named PseudoSort, which employs advanced self-supervised learning techniques, a distinctive density-based pseudo-labelling strategy, and an iterative fine-tuning process to enhance spike sorting accuracy. Through extensive benchmarking on large-scale simulated datasets, we demonstrate the superior performance of PseudoSort compared to recently developed machine learning-based (ML) spike sorting algorithms. We showcase the practical application of PseudoSort by utilising MEA recordings from hippocampal neurons exposed to subneuronal concentrations of monomeric Tau, a protein associated with Alzheimers disease (AD). Our results, validated against patch clamp experiments, unveil that monomeric Tau at subneuronal concentrations induces stimulation-dependent disruptions in both local and global activity of hippocampal neurons. Remarkably, patch clamp electrophysiology highlights the effect of combined Tau and neuronal stimulation treatment on excitatory postsynaptic currents, whereas PseudoSort excels in identifying neuronal clusters that exhibit diminished firing capacity following Tau treatment alone, i.e., in the absence of stimulation. This comprehensive approach validates the prowess of PseudoSort and unravels the intricate effects of Tau on neuronal activity, particularly in the context of AD.

neuroscience↗

Plk3 Regulates Bacteremia and Supports Sepsis

ObjectiveSepsis, which is the bodys response to overwhelming infection, can lead to septic shock, characterized by thrombocytopenia, hypotension, and organ damage. Polo-like kinase 3 (Plk3) is a ubiquitously expressed serine/threonine kinase, but its exact role in immune function is unknown. Approach and ResultsWe used Plk3-/- and WT mice to evaluate the function of Plk3 in several models of severe sepsis. We found that WT mice die within 48 hours of 100% cecal ligation and puncture (CLP), while Plk3-/- mice survive. Survival following cecal slurry (CS) injection mirrored that of CLP as recipient WT mice succumbed, while recipient Plk3-/- mice survived. Analysis of bacterial load 24 hours after CLP revealed that WT blood and peritonea were loaded with bacteria, but bacteria were virtually undetectable in the peritonea or blood of Plk3-/- mice. To determine if bacteria infiltrate the blood of Plk3-/- mice shortly after infection, we measured bacteria 1 and 3 hours after CS injection. We found a time-dependent increase in bacteria in the blood of WT mice that was not observed in Plk3-/- mice. To determine if the lack of bacteria in the blood of Plk3-/- mice is due to enhanced clearance, we injected E. coli IV into WT and Plk3-/- mice. We found 75% mortality for both WT and Plk3-/- mice within 72 hours following IV injection suggesting that survival of Plk3-/- mice following enteric infection is likely due to reduced bacteremia. ConclusionCollectively our data suggest that Plk3 supports the systemic dissemination of bacteria and subsequent sepsis following enteric infection.

immunology↗

Revealing nanoscale structure and interfaces of protein and polymer condensates via cryo-electron microscopy

Liquid-liquid phase separation (LLPS) is a ubiquitous demixing phenomenon observed in various molecular solutions, including in polymer and protein solutions. Demixing of solutions results in condensed, phase separated droplets which exhibit a range of liquid-like properties driven by transient intermolecular interactions. Understanding the organization within these condensates is crucial for deciphering their material properties and functions. This study explores the distinct nanoscale networks and interfaces in the condensate samples using a modified cryo-electron microscopy (cryo-EM) method. The method involves initiating condensate formation on electron microscopy grids to control droplet size and stage in the phase separation process. The versatility of this method is demonstrated by imaging three different classes of condensates. We further investigate the condensate structures using cryo-electron tomography which provides 3D reconstructions, uncovering porous internal structures, unique core-shell morphologies, and inhomogeneities within the nanoscale organization of protein condensates. Comparison with dry-state transmission electron microscopy emphasizes the importance of preserving the hydrated structure of condensates for accurate structural analysis. We correlate the internal structure of protein condensates with their amino acid sequences and material properties by performing viscosity measurements that support that more viscous condensates exhibit denser internal assemblies. Our findings contribute to a comprehensive understanding of nanoscale condensate structure and its material properties. Our approach here provides a versatile tool for exploring various phase-separated systems and their nanoscale structures for future studies.

biophysics↗

Genomic evidence for hybridization and introgression between blue peafowl and green peafowl and selection for white plumage

The blue peafowl (Pavo cristatus) and the green peafowl (Pavo muticus) have significant public affection due to their stunning appearance, although the green peafowl is currently endangered. Some studies have suggested introgression between these the two species, although evidence is mixed. In this study, we successfully assembled a high-quality chromosome-level reference genome of the blue peafowl, including the autosomes, Z and W sex chromosomes as well as a complete mitochondria DNA sequence. Data from 77 peafowl whole genomes, 76 peafowl mitochondrial genomes and 33 peahen W chromosomes genomes provide the first substantial genetic evidence for recent hybridization between green and blue peafowl. We found three hybrid green peafowls in zoo samples rather than in the wild samples, with blue peafowl genomic content of 16-34%. Maternal genetic analysis showed that two of the hybrid female green peafowls contained complete blue peafowl mitochondrial genomes and W chromosomes. Hybridization of endangered species with its relatives is extremely detrimental to conservation. Some animal protection agencies release captive green peafowls in order to maintain the wild population of green peafowls. Therefore, in order to better protect the endangered green peafowl, we suggest that purebred identification must be carried out before releasing green peafowls from zoos into the wild in order to preventing the hybrid green peafowl from contaminating the wild green peafowl. In addition, we also found that there were historical introgression events of green peafowl to blue peafowl in four Zoo blue peafowl individuals. The introgressed genomic regions contain IGFBP1 and IGFBP2 genes that could affect blue peafowl body size. Finally, we identified that the nonsense mutation (g.4:12583552G>A) in the EDNRB2 gene is the genetic causative mutation for white feather color of blue peafowl (also called white peafowl), which prevents melanocytes from being transported into feathers, such that melanin cannot be deposited.

genomics↗