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Wang, J.

Publications and source records attributed to Wang, J..

At least 415 records · Page 23Linked to original sources

CCycDB: an integrative knowledgebase to fingerprint microbially mediated carbon cycling processes

Microorganisms play essential roles in mediating biogeochemical cycling of carbon across Earths ecosystems. Understanding the processes and underlying mechanisms for microbially mediated carbon cycling is therefore critical for advancing global ecology and climate change research. To comprehensively depict these complex biogeochemical processes, we developed CCycDB, a knowledge-based functional gene database, to accurately fingerprint microbially-mediated carbon cycling pathways and gene families, particularly from shotgun metagenomes. The CCycDB database comprises 4,676 gene families classified into six major functional categories, further structured into 45 level-1 and 188 level-2 sub-categories, encompassing a total of 10,991,724 high-quality reference sequences. Validation using both synthetic and real-world datasets demonstrated that CCycDB outperforms existing orthology databases in terms of accuracy, coverage and specificity. By directly targeting carbon-cycling functional gene families, CCycDB provided promising routines to reconstruct both functional gene and taxonomic profiles associated with microbially mediated carbon cycling. Application of CCycDB to shotgun metagenomes from diverse and complex ecosystems revealed pronounced habitat-specific differences in carbon cycling processes and their associated microbial taxa. Collectively, CCycDB provides a powerful and reliable tool for profiling carbon cycling processes from both functional and taxonomic perspectives in complex ecosystems. CCycDB is accessible at https://ccycdb.github.io/. Impact StatementThe microbially mediated carbon cycling processes are the most complex biogeochemical processes in the Earths biosphere, playing profound regulatory roles on global climate changes. A key bottleneck in linking microbial communities to global change is the lack of integrated tools for comprehensive carbon cycle profiling. Here, we present CCycDB, a tool that serves a dual purpose--first being a reference database that obtains functional gene and taxonomic profiles and functioning as a customized routine for efficiently aligning sequences and querying associated functional information. CCycDB enables researchers to accurately link microbial community dynamics to carbon cycling and transforming pathways, thereby advancing integrated global change studies with microbes and ecological research via complex metagenomic datasets.

microbiology↗

Ca2+-mediated protein citrullination regulates proliferation in the regenerating and malignant CNS

Unlike most adult mammals, regenerative vertebrates can awaken dormant neural stem cells in response to injury. Understanding how this process is regulated could guide strategies to activate stem cells for tissue repair and limit aberrant proliferation in cancers of the CNS. Here, using zebrafish injury models and high-speed live imaging, we identify hydrodynamically-activated Ca{superscript 2} signalling as a key driver of neural stem cell activation. Local injury-associated changes in CSF flow activate mechanoreceptors at the site of spinal cord lesions, triggering pulsatile Ca{superscript 2} activity and progenitor proliferation. We identify Ca{superscript 2}-regulated peptidylarginine deiminase enzymes (PADs) as key downstream effectors that citrullinate intracellular targets in a Ca{superscript 2}-dependent manner to drive progenitor activation. Finally, we show that PAD inhibitors suppress the growth of aggressive medulloblastoma cells in preclinical laboratory models. Together, these findings uncover a novel mechanism of proliferation control in the vertebrate CNS and highlight the value of regenerative studies for identifying therapeutic targets. Graphical Summary O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=198 SRC="FIGDIR/small/701650v1_ufig1.gif" ALT="Figure 1"> View larger version (67K): org.highwire.dtl.DTLVardef@3b7a56org.highwire.dtl.DTLVardef@dcc81forg.highwire.dtl.DTLVardef@d6a208org.highwire.dtl.DTLVardef@127de86_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIDamage signals trigger changes in cilia activity and CSF flow to create a transient hydrodynamic niche at the site of spinal cord injuries. C_LIO_LISpecialised CSF-contacting neurons detect altered CSF circulation and initiate a signalling relay that culminates in elevated Ca{superscript 2} activity within dormant neural progenitors. C_LIO_LICa{superscript 2}-activated PAD enzymes link Ca{superscript 2} signalling to cell cycle progression by citrullinating intracellular targets in a Ca{superscript 2}-dependent manner. C_LIO_LIPAD inhibition suppresses medulloblastoma growth in preclinical laboratory models, revealing a conserved regulatory mechanism with translational potential. C_LI

neuroscience↗

OSTM1 is a ubiquitin E3 ligase that suppresses B-cell malignancy by activating the cAMP/PKA/CREB pathway

Osteoclastogenesis-associated transmembrane protein 1 (OSTM1) is a glycosylated, membrane-integral protein that regulates lysosomal homeostasis, with loss-of-function mutations causing autosomal recessive osteopetrosis. Through a whole-genome CRISPR/Cas9 screen, we identified OSTM1 as a previously unrecognized tumor suppressor in B-cell malignancies. Consistent with this role, OSTM1 is frequently deleted or downregulated across a broad spectrum of human B-cell cancers. In mice, B-cell-specific monoallelic or biallelic ablation of Ostm1 cooperates with Cdkn2a loss to drive lymphomagenesis with near-complete penetrance. Mechanistically, we uncover a cytosolic, non-glycosylated fraction of OSTM1 that functions as a ubiquitin E3 ligase to promote proteasomal degradation of phosphodiesterase 3B (PDE3B). As PDE3B hydrolyzes cAMP and suppresses the tumor-protective PKA/CREB/CREBBP signaling axis, loss of OSTM1 leads to PDE3B stabilization, attenuation of cAMP signaling, and enhanced oncogenic transformation. Together, our findings establish OSTM1 as a critical suppressor of B-cell lymphomagenesis through regulation of the cAMP/PKA/CREB pathway.

cancer biology↗

Entomoscope 2.0 and ENIMAS 2.0: An Open-Source, AI-Integrated Platform for Rapid and Affordable Insect Digitization

The rapid decline of global biodiversity necessitates scalable and accessible tools for monitoring insect populations, yet the high cost and slow pace of specimen digitization remain significant bottlenecks. To address this challenge, we present the Entomoscope 2.0, an open-source platform that integrates a low-cost photomicroscope with an AI-integrated software suite, ENIMAS 2.0. The system combines optimized hardware with an end-to-end software module for digital specimen curation. This module comprises automated specimen cropping, background standardization, morphometric analysis using an Oriented Bounding Box (OBB) with a human-in-the-loop (HITL) supervision, and a flexible interface for rapid taxonomic screening using custom AI identification models. With a material cost of only 400 {euro}, the system offers a cost-effective alternative to expensive commercial solutions. We compare results from the Entomoscope 2.0 with a high-end commercial system (Keyence) using 54 insect specimens and demonstrate the efficiency of the proposed AI workflow. Entomoscope 2.0 completed the whole digitization process in an average of 54.6 seconds per specimen, representing a 2.28-fold increase in speed over the Keyence systems multi-step workflow (124.3 seconds). Crucially, all hardware specifications and construction manuals are freely available to support widespread adoption. By lowering financial barriers and accelerating research workflows, the Entomoscope 2.0 platform offers a practical solution to enable high-throughput digitization for researchers, educators, and citizen scientists.

bioengineering↗

Small brown planthopper infestation enhances it reproduction and insecticide tolerance by manipulating glucose distribution and levels in rice

The evolutionary arms race between plants and insects involves not only direct defense and counter-defense but also sophisticated resource manipulation. However, how herbivorous insects respond to host nutritional signals to modulate their fitness traits remains unclear. This study investigates how the small brown planthopper (SBPH, Laodelphax striatellus) manipulates host plant carbohydrate allocation, and to elucidate the molecular mechanisms by which the acquired glucose enhances SBPH fecundity and insecticide tolerance. Using molecular, pharmacological, and biochemical approaches, we found that SBPH infestation induced systemic carbohydrate reallocation in rice, elevating whole-plant glucose levels by promoting aerial accumulation while depleting root reserves. Host-derived glucose enhanced SBPH fecundity by activating the target of rapamycin (TOR) pathway, upregulating juvenile hormone (JH) signaling, and increasing vitellogenin production. For imidacloprid tolerance, glucose boosted glutathione S-transferase (GST) activity via two synergistic mechanisms: by upregulating glutamate cysteine ligase (GCL) to increase glutathione synthesis, and transcriptionally via the glucose-TOR-JH axis to induce LsGSTe1 (SBPH epsilon class GST) and LsGSTo1 (SBPH omega class GST) expression. Our findings establish host-derived glucose as a central signaling molecule that SBPH utilizes to modulate conserved pathways for simultaneous optimization of reproduction and insecticide resistance. This reveals a multifaceted nutrient-responsive mechanism in insect pests and identifies the glucose-TOR-JH axis as critical molecular targets for developing nutrient-based pest control strategies, such as disrupting insect nutrient-sensing pathways or modulating host carbohydrate metabolism. HighlightsO_LISBPH infestation elevates whole-plant glucose levels by promoting aerial accumulation while suppressing root abundance. C_LIO_LISBPH-induced glucose in the rice aerial tissues boosts reproduction of SBPH via TOR-JH-Vg pathway. C_LIO_LISBPH-induced glucose in the rice aerial tissues enhances imidacloprid tolerance of SBPH through GCL-GSH-GST and TOR-JH-GST axis. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=152 SRC="FIGDIR/small/701313v2_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@39e084org.highwire.dtl.DTLVardef@1c310cborg.highwire.dtl.DTLVardef@1838575org.highwire.dtl.DTLVardef@1aedebe_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

Progerin-Induced Nuclear Envelope Remodeling is Shaped by Cell Division and NUP153

The nuclear envelope (NE) undergoes dynamic remodeling during both physiological and pathological processes. Nuclei in cells from patients with accelerated aging diseases and from elder individuals are often lobular and convoluted. Despite extensive study, the steps of phenotype acquisition and the co-factors required are not yet fully understood. Here, we focused on progerin, a mutant form of lamin A that causes Hutchinson Gilford Progeria Syndrome (HGPS). Using an inducible cell-based system, we characterized two distinct stages of NE remodeling. Correlative light and electron microscopy of interphase-arrested cells showed that prior to cell division, progerin primarily affects the inner nuclear membrane (INM), inducing focal expansion, invagination, and the formation of multi-membranous structures, while the outer nuclear membrane remains largely unaffected. These focal regions of progerin accumulation are enriched for specific INM proteins and the nucleoporin NUP153 but largely exclude nuclear pore complexes. Live and fixed image analysis demonstrated that, upon cell division and NE reassembly, progerin-expressing cells develop pronounced nuclear lobulations characteristic of HGPS. Appreciation of this stepwise development of phenotype lends insight into cellular manifestations of aging in mitotic versus post-mitotic cell types and provides a system in which to study factors that contribute to these distinct stages in the disruption of nuclear morphology. Depletion of NUP153 reduced NE foci formation in interphase-arrested cells expressing GFP-progerin, suggesting NUP153 promotes or stabilizes INM invagination. Aberrant nuclear architecture is just one cellular feature that changes during normal and accelerated aging but its etiology provides a critical framework for understanding accompanying consequences on chromatin packaging, DNA damage, and the ER stress response.

cell biology↗

Antigen presentation requirements for effective cDC1-based cancer immunotherapy

Type 1 conventional dendritic cells (cDC1s) are important for generating and sustaining antitumor immunity. Accordingly, the abundance of cDC1s in human tumors correlates with improved outcomes in cancer. Capitalizing on this role, we previously demonstrated that vaccination with in vitro-derived murine cDC1s elicits durable tumor control in multiple preclinical models; however, the immunological mechanisms underlying the efficacy of cDC1 vaccination remain unclear. Here, we examined whether in vitro-derived cDC1s resemble tumor-infiltrating DC populations and whether MHC-I and MHC-II antigen presentation contribute to cDC1-mediated tumor control following vaccination in melanoma. As expected, MHC-I- or MHC-II-deficiency had minimal impact on the transcriptional state of cDC1s in homeostasis or following stimulation with the adjuvant poly dI:dC. Moreover, in vitro-derived cDC1s cultured under steady-state conditions closely resembled tumor-infiltrating cDC1s, whereas their poly dI:dC-stimulated counterparts resembled CCR7+ tumor-infiltrating DC populations, also referred to as mregDCs or LAMP3+ DCs. Our data further show that both MHC-I and MHC-II contribute to tumor control upon cDC1 vaccination, and coexpression of MHC-I and MHC-II on the same cDC1 is necessary for a robust vaccine response. We also identified an important function for host cDC1s in supporting the efficacy of vaccination with in vitro-derived cDC1s, as judged by impaired tumor control in Irf8+32-/- mice, which lack endogenous cDC1s. Overall, these results indicate that effective antitumor responses depend on MHC-I and MHC-II antigen presentation by vaccine-delivered cDC1s, with additional contributions from host cDC1s. Key pointsO_LIIn vitro-generated cDC1s resemble intratumoral DC populations found in mice and humans. C_LIO_LIMHC-I and MHC-II antigen presentation by vaccine-delivered cDC1s contribute to antitumor efficacy. C_LIO_LICoexpression of MHC-I and MHC-II on the same cDC1 enhances vaccine responses. C_LIO_LIAntitumor responses reflect the activity of vaccine and endogenous cDC1s. C_LI

immunology↗

BRD8 Disruption Unlocks Retrotransposon-Driven Tumor-Intrinsic Immunogenicity via Redistribution of Histone Acetylation in Liver Cancer

Overall response rate for immune checkpoint blockade (ICB) therapy remains limited in advanced hepatocellular carcinoma (HCC). Epigenetic perturbation has been shown to enhance tumor-intrinsic immunogenicity and thus reverse resistance to immunotherapy in multiple cancer types. Here, through domain-focused CRISPR/Cas9 genetic screens targeting chromatin regulators, we identified the bromodomain-containing protein 8 (BRD8) as a novel epigenetic factor that functions to suppress tumor-intrinsic immunity through BRD8/EP400 chromatin remodeling complex in HCC. Depletion of BRD8 potently induces anti-tumor immune responses and sensitizes tumors to ICB therapy in mouse tumor models. Mechanistically, BRD8 knockout leads to partial redistribution of H3K27ac to repetitive elements, activating double-stranded RNA (dsRNA) accumulation and subsequent type I interferon (IFN) response, leading to tumor rejection via remodeling the tumor microenvironment. The first-in-class selective chemical probe DN02 targeting the bromodomain of BRD8 also exhibits immunomodulatory and anti-tumor effects. In line with these findings, low BRD8 expression correlated with enriched interferon signatures, elevated immune cell infiltration, and prolonged survival in HCC patients. Together, this study reveals BRD8 as a selective epigenetic vulnerability for promoting tumor immunogenicity, and its bromodomain as a druggable target to overcome immunotherapy resistance in liver cancer patients. SignificanceThe induction of viral mimicry by epigenetic interference could trigger tumor-intrinsic immune response and enhance immunotherapy efficacy. Here, through domain-focused CRISPR/Cas9 screening, we identify BRD8 complex as a regulator involved in the suppression of repetitive elements by controlling H3K27ac distribution and propose BRD8 as a novel target to enhance anti-tumor immunity.

cancer biology↗

A developmental timer coordinates organism-wide microRNA transcription

The development of distinct tissues must be precisely coordinated to ensure that growth and cell fate transitions occur in the correct temporal order across the organism, yet the mechanisms that coordinate these timing events remain unclear. In Caenorhabditis elegans, stage-specific cell fate transitions are driven by pulsatile transcription of heterochronic microRNAs, but the source of these rhythms has been unknown. Here, we identify a developmental timer composed of the transcription factor MYRF-1 and the PERIOD-like repressor LIN-42 that operates in all somatic cells. MYRF-1 binds conserved regulatory elements upstream of heterochronic microRNA genes and drives synchronized, once-per-stage transcriptional pulses across tissues, while concurrently activating lin-42 expression. Newly synthesized LIN-42 directly associates with MYRF-1, limiting its nuclear residence and transcriptional activity to constrain the amplitude and duration of each transcriptional burst. This reciprocal transcriptional/translational feedback loop generates organism-wide, phase-locked microRNA expression, coupling tissue-specific development to organismal growth through a shared timing mechanism.

developmental biology↗

Telomere-to-telomere gap-free and phased genome assembly reveals post-allopolyploidization subgenomic diversification of tobacco centromeres

Common tobacco (Nicotiana tabacum) is an economically important crop worldwide whose allotetraploid genome sequence remains incompletely assembled. We report a 4.2Gb telomere-to-telomere gap-free N. tabacum reference genome phased into two subgenomes (subS/subT) , revealing differential evolutionary trajectories between two subgenomes marked by a rapid divergence in paternal T-genome, especially for heterochromatic centromeric regions. To understand the evolutionary dynamics of tobacco centromeres following polyploidization, we characterize the centromere architecture in tetraploid N. tabacum and its diploid progenitors, demonstrating clear subgenomic diversification and repositioning of centromeres during speciation and allopolyploidization. These findings enrich our knowledge on centromere paradox in polyploid genomes.

genomics↗

Medication and Developmental Stage Shape the Brain-Age Gap in ADHD

Identifying robust biomarkers for Attention-Deficit/Hyperactivity Disorder (ADHD) remains challenging. The brain-age gap (BAG), which is the difference between predicted biological brain age and chronological age, has been proposed as a marker of atypical neurodevelopment, yet findings have been inconsistent across studies, limiting its clinical translation. We hypothesized that two major sources of heterogeneity, medication status and developmental stage, contribute to these discrepancies. To test this, we analyzed structural MRI data from a large multi-site cohort (N = 947; ages 7-26 years). Individuals with ADHD were stratified by medication history (medicated vs. unmedicated) and developmental stage (childhood vs. adolescence) and compared with matched typically developing controls. We estimated global- and network-level BAG using publicly available pretrained models. Our results revealed that BAG differences were contingent on medication status, with abnormalities predominantly observed in unmedicated individuals. Developmental stage further moderated the presence of this effect: BAG abnormalities were present significantly in children with ADHD but not in adolescents. Network-level analyses demonstrated spatial specificity, with significant effects in visual, ventral attention, and dorsal attention networks. Together, these findings provide compelling evidence that BAG in ADHD is both medication- and development-dependent, highlighting the need to account for treatment exposure and developmental stage when evaluating BAG as a biomarker and further underscoring the importance of early intervention during childhood.

neuroscience↗

Sleep dynamics and epileptogenesis following Kainic acid in epilepsy-susceptible (DBA/2J) and epilepsy-resistant (C57BL/6) mice

Susceptibility to epileptogenesis varies in humans and outbred mouse strains. We hypothesized that baseline sleep abnormalities increase susceptibility to epileptogenesis following Kainic acid (KA) and abnormal circadian rhythm or sleep homeostasis (SH) contribute to worse seizures in epilepsy-susceptible DBAs. Following EEG electrode implantation, C57 and DBA mice underwent repeated low-dose KA or saline treatment. Seizures, interictal spikes and sleep were examined over 8-weeks with continuous electroencephalography (EEG). Seizures were manually scored, and interictal spikes and sleep were analyzed using machine learning algorithms. Slow wave activity (SWA) was derived from non-rapid eye movement (NREM) sleep following Fourier transform, and SH was measured by the decay of SWA during sleep and its rise with preceding wakefulness. The impact of post-KA seizures on circadian rhythm was determined using Cosinor analysis. Seizures were longer and more frequent in KA-treated DBAs than C57s. Interictal spike were much greater in saline-treated DBAs than C57s. SWA across the 24-hours was lower in DBAs at baseline. KA treatment decreased REM and increased SWA activity in DBAs but not in C57s. Cosinor analysis revealed circadian rhythm abnormalities in DBAs but not in C57s. Seizures impaired SH in DBAs, with no increase in SWA with preceding wakefulness and a progressive loss of SWA decay during lights-on over the 8-week recording. These findings suggest that baseline sleep abnormalities, poorly adaptable circadian rhythms and impaired SH are associated with increased vulnerability to epileptogenesis. Therapies enhancing circadian rhythm and SH after an insult may be avenues to mitigate epileptogenesis in vulnerable populations. Significance StatementSusceptibility to spontaneous seizures after a central nervous system insult varies in humans as well as mouse models. The reasons behind the differential susceptibility are not entirely known. Here we show in a Kainic acid model, epilepsy susceptible DBA/2J mice have baseline sleep abnormalities and following Kainic acid treatment, these mice develop more frequent and longer seizures than the epilepsy-resistant C57BL/6 mice. We also show that the epilepsy-susceptible DBAs have abnormalities in circadian rhythm as well as sleep homeostasis measures compared to epilepsy-resistant C57s, which may be contributing to progression of epileptogenesis and worse seizures. Therapies targeted at enhancing sleep, circadian rhythms or sleep homeostasis may be avenues to mitigate development of spontaneous seizures after a central nervous system insult.

neuroscience↗

Integration of early-stage cryopreservation and cell cycle modulation into a flexible kidney organoid differentiation system

Kidney organoids derived from human pluripotent stem cells (hPSCs) represent a promising platform for modeling nephrogenesis and renal diseases. However, conventional differentiation protocols are continuous and time-sensitive, limiting their scalability and reproducibility. Here, we developed a method to pause and resume organoid formation through cryopreservation at an early differentiation stage using a chemically defined formulation that maintains high post-thaw viability and differentiation potential. We further found that synchronizing hPSCs in the G1 phase with PD-0332991 enhanced post-thaw organoid formation and transcriptional fidelity, while G2 phase enrichment with Ro-3306 promoted the development of SLC12A3-positive distal convoluted tubules. The post-thaw organoids exhibited well-organized nephron architecture and function comparable to uninterrupted cultured controls. This platform proved effective for modeling BK polyomavirus (BKV) infection, drug-induced nephrotoxicity, and renal fibrosis. Together, our cryopreservation and cell cycle synchronization strategy provides a flexible, practical framework to advance organoid-based research and translation.

developmental biology↗

Biomimetic model for the identification of distinctive microenvironmental factors in glioblastoma radiosensitivity

Radiation therapy (RT) has long been included in the treatment of glioblastoma (GBM). However, radioresistance in cancer cells as well as toxicity in normal tissues are major obstacles to clinical efficacy. Improved understanding of the mechanisms of tumor microenvironment-induced radioresistance during and after radiation therapy can provide fundamental insights to improve clinical outcomes in GBM. Here, using three-dimensional engineered hydrogel models in vitro, we report the influence of extracellular matrix, hypoxia, and adjacent neuronal cells in radiotherapeutic sensitivity. We find that mechanical cues and oxygen availability regulate cellular response to radiation, with softer matrices allowing for more DNA damage. Hyaluronan fragments from the extracellular matrix also modulate rapid metabolic response to radiation, especially in hypoxic environments. We show that neuronal networks influence tumor metabolic activity and the inflammatory response. Overall, we demonstrate here that alternative radiation strategies, such as low dose rate radiation therapy and microenvironmental regulation, have the potential to be more effective in a specific subset of radiosensitive GBM tumors.

bioengineering↗

ThermiQuantTM AquaStream: A Portable Instrument for Quantitative Colorimetric Isothermal Nucleic Acid Amplification Reactions in Paper and Tube Formats

Microfluidic paper-based analytical devices ({micro}PADs) are an attractive format for colorimetric nucleic acid amplification tests (NAATs) because they enable low-cost, portable diagnostics in resource-limited settings. However, researchers often optimize colorimetric assays in liquid reactions in tubes before translating them to {micro}PADs. Since both formats require separate instruments for incubation and real-time sensing, direct comparison of reactions between the two formats is difficult. To address these cross-platform limitations, we developed ThermiQuant AquaStream, a portable benchtop device (15 x 20 x 16 cm, [~]5 kg; cost: USD 327) that supports seamless colorimetric loop-mediated isothermal amplification (LAMP) reactions in both {micro}PADs and tubes under a common workflow. The system enables real-time reaction tracking (every 30 seconds) through onboard image processing, precise isothermal control ({+/-}0.5 {degrees}C) using a repurposed consumer-grade sous-vide heater, and medium-throughput (24 tubes or 42 {micro}PADs). Testing with synthetic SARS-CoV-2 orf7ab DNA fragments demonstrated a limit of detection corresponding to a 95% probability of detection (LOD95) of 110 copies per reaction in tube (22 copies/{micro}L) and 39 copies per reaction in {micro}PADs (5 copies/{micro}L), estimated using probit regression. In both formats the limit of quantification (LOQ), defined as the lowest concentration yielding a coefficient of variation (CV) of Tq [≤] 10%, was 250 copies/reaction resulting in a strong linear (R2 = 0.98 & R2 = 0.96 respectively for tube and {micro}PADs) standard calibration curves. ThermiQuant AquaStream provides an affordable and versatile benchtop platform capable of supporting both tube- and {micro}PAD-based colorimetric LAMP assays, serving as a proof-of-concept research tool for assay development and molecular diagnostics in One Health settings such as clinics, farms, and field environments.

bioengineering↗

Engineered Allosteric Control Enhances Specificity and Potency of CRISPR-CasX-based Epigenetic Repressors

Epigenetic silencers are targeted therapeutics capable of persistently silencing genes without altering DNA sequence. However, many current silencers rely on direct fusion of a constitutively active DNA methyltransferase catalytic domain, that may cause off-target DNA methylation and cellular toxicity. In contrast, endogenous DNA methyltransferases contain a regulatory domain (ADD) that requires recognition of a permissive histone state before DNA methylation can occur. We reinstate this sequential proofreading logic in a compact CasX-based epigenetic repressor, generating Epigenetic Long-term CasX Repressors (ELXRs) that read local chromatin state before writing DNA methylation. In ELXRs, a repressor domain converts active chromatin into a state that unlocks the allosteric DNA methyltransferase. This creates a sequential, multi-gated process that preferentially restricts DNA methylation to the target site. Surprisingly, implementing this proofreading mechanism improved not only specificity, but also activity. ELXRs decreased off-target methylation up to 10-fold and rescued DNMT3A-dependent growth defects, with transcriptome profiling showing up to 150-fold fewer dysregulated genes, while also increasing on-target repression up to 4-fold across multiple loci. In mouse models, lipid nanoparticle delivery produced potent, durable PCSK9 silencing with precise promoter methylation. Together, these results demonstrate that coupling molecular recognition to effector activity through sequential proofreading can simultaneously improve specificity and potency.

molecular biology↗

Striatal Acetylcholine Dip-Rebound Is Induced by Direct Pathway Neurons and Encodes Action-Outcome Contingency

Learning contingencies between actions and outcomes is pivotal in adaptive behavior and requires ongoing flexible sensory, motor, and reward information integration. The striatum is central to the integration. However, it remains unclear how the striatal neurons interact with each other to facilitate learning. Here we show that in the dorsomedial striatum, direct-pathway medium spiny neurons (dMSNs), but not dopamine (DA), inhibit cholinergic interneurons, inducing a characteristic acetylcholine (ACh) dip-rebound, encoding action-outcome (A-O) contingency. Using genetically encoded sensors and in vivo fiber photometry, we find that dMSN activation, ACh dip-rebound, and DA transients emerge only after mice acquire the contingency. dMSN activity and ACh dynamics persist even when rewards are probabilistic, adapt in time as the learned relationship between action and reward evolves, and vanish when the contingency is degraded. Ex vivo recordings and in vivo optogenetics further show that dMSN activity is both sufficient and required to generate the ACh dip-rebound through GABAergic inhibition of cholinergic interneurons. Disrupting the dip-rebound slows acquisition and accelerates extinction. Together, these findings reveal a previously unrecognized dMSN-ACh circuit mechanism that encodes contingency during instrumental learning, operating alongside DA signals that track reward outcomes.

neuroscience↗

Dual Engram Architecture within a Single Striatal Cell Type Distinctly Controls Alcohol Relapse and Extinction

Relapse is a major obstacle in the treatment of alcohol and drug addiction and is thought to be driven by persistent drug-associated memories formed during their use. Behavioral therapies such as extinction training can reduce relapse and are proposed to work by creating a competing memory trace. However, where and how these opposing memories are stored in the brain is unknown. Here, we show that two anatomically and functionally distinct engram ensembles within the same genetically defined striatal cell type, direct-pathway medium spiny neurons (dMSNs), encode these opposing memories. Using engram-tagging tools in mice, we found that the acquisition of operant alcohol learning recruits a dMSN ensemble enriched in the striatal matrix compartment that stores alcohol-associated memories and whose activation selectively promotes relapse. Conversely, extinction of alcohol seeking recruits a dMSN ensemble enriched in the striosome compartment that stores extinction-related memories and whose activation suppresses relapse. Furthermore, we reveal that the physical memory trace storing the relapse-promoting memory is embedded within persistently strengthened corticostriatal synapses engaged during learning, and that artificially reproducing this plasticity is sufficient to trigger relapse-like behavior. These findings uncover a dual-engram architecture within dMSNs that governs relapse and extinction, providing a mechanistic framework for understanding how competing memories regulate drug-seeking behavior. HighlightsO_LIAcquisition and extinction of alcohol learning recruits distinct dMSN ensembles. C_LIO_LIAcquisition recruits matrix-enriched dMSN ensembles to promote relapse. C_LIO_LIExtinction recruits striosome-enriched dMSN ensembles to suppress relapse. C_LIO_LIEngram dMSNs show lasting synaptic potentiation and mimicking this potentiation triggers relapse. C_LI

neuroscience↗