Biology subjects

Zheng, W.

Publications and source records attributed to Zheng, W..

At least 91 records · Page 5Linked to original sources

bioRxiv2025

The Role of N-acetylcysteine Amide in Acute Graft-versus-host Disease Mouse Model

Graft-versus-host disease (GvHD) remains one of the major complications following allogeneic hematopoietic cell transplantation (allo-HCT), resulting in reduced quality of life, morbidity, and mortality in transplanted patients. Clinical strategies to prevent GvHD are frequently associated with off-target effects and dose-related toxicity. Given that oxidative stress is elevated in allo-HCT recipients and contributes to the pathogenesis of GvHD, the current study aims to characterize the role of N-acetylcysteine amide (NACA), a novel antioxidant, as the prophylactic treatment for acute GvHD. Using a murine GvHD model, we found that oral administration of NACA significantly reduced GvHD severity, prolonged survival, and improved the clinical manifestations and integrity of target organs compared to saline or N-acetylcysteine (NAC) treatment. NACA modulated splenic T cells differentiation with an increase in the regulatory (Treg) subset and a decrease in the cytotoxic (CD8+) subset. Moreover, inflammatory mediators, such as ROS and pro-inflammatory cytokines were downregulated by NACA treatment. In addition, NACA hindered donor T-cell proliferation in the recipients, and restrained Th1 and Th17, but not Th2 polarization. Importantly, NACA did not influence full donor engraftment in bone marrow and spleen. Taken together, our findings provide a new candidate for GvHD prophylactic treatment by targeting oxidative stress that can be easily translated to clinical use. Key PointsO_LINACA provides superior prophylactic effect against aGvHD compared to NAC in an allogeneic transplantation mouse model. C_LIO_LINACA treatment neither showed systemic toxicity nor altered the engraftment of the donor cells. C_LI

pharmacology and toxicology↗

bioRxiv2025

A novel antioxidant N-acetylcysteine Amide Alleviates Cyclophosphamide-induced Endothelial Damage

The alkylating agent cyclophosphamide (Cy) is one of the important corner stones in cancer treatment. Cy is used also as a part of conditioning regimens prior to hematopoietic cell transplantation and as a prophylactic treatment post transplantation in graft-versus-host disease. Existing evidence showed that high doses of Cy are associated with a number of side effects, including damage on arterial endothelium, which might contribute to late cardiovascular disorders. Oxidative stress has been characterized in such pathogenesis and is an exploitable target for treatment. Herein, the study aimed to investigate the protective role of the novel antioxidant N-acetylcysteine amide (NACA) in Cy-induced endothelial injury and explore the underlying mechanism. Our in vivo results showed that NACA partially reduced the endothelial injury and recovered the integrity of arterial endothelium in the mice treated with Cy. In addition, we found that NACA decreased the cytotoxicity of Cy on endothelial cells through alleviating caspase-dependent apoptosis, DNA damage and oxidative stress. Meanwhile, NACA pre-treatment rebalanced endothelial nitric oxide synthase (eNOS) and arginase I and preserved the angiogenic capability of endothelial cells which was compromised by Cy through blockage of Notch signaling pathway. Interestingly, in comparison to N-acetylcysteine (NAC), its amide derivative NACA showed superior ability to alleviate Cy-induced endothelial damage. In conclusion, the current study proved the robust endothelial protective potential of NACA, facilitating clinical use of the novel antioxidant.

pharmacology and toxicology↗

bioRxiv2025

ERK and PKA signaling drive the evolutionary expansion of the cortex

The human cerebral cortex, essential for intelligence, cognition, and language, evolved its uniqueness through mechanisms that drive neuronal expansion. Here, we demonstrate that ERK and PKA signaling cooperatively preserve the neurogenic capacity of cortical radial glia (RGs) by inhibiting gliogenic YAP/TAZ and SHH signaling. YAP/TAZ signaling drives cortical RGs toward ependymal fate, while SHH signaling facilitates the generation of cortical tripotential intermediate progenitor cells that produce cortical astrocytes, oligodendrocytes, and olfactory bulb interneurons. During evolution, cortical outer radial glia (oRGs) acquire dominant ERK/PKA signaling through a self-reinforcing feedback mechanism that suppresses both YAP and SHH pathways, thereby significantly enhancing oRG self-renewal capacity and extending the neurogenic period. These findings reveal that cortical neurogenesis, gliogenesis, and evolutionary expansion are coordinately regulated by an integrated ERK, PKA, YAP/TAZ, and SHH signaling, identifying a unifying principle of mammalian cortical development and evolution.

developmental biology↗

bioRxiv2025

PERADIGM: Phenotype Embedding Similarity-based Rare Disease Gene Mapping

Identifying genes associated with rare diseases remains challenging due to the scarcity of patients and the limited statistical power of traditional association methods. Here, we introduce PERADIGM (Phenotype Embedding similarity-based RAre DIsease Gene Mapping), a novel framework that leverages natural language processing techniques to integrate comprehensive phenotype information from electronic health records for rare disease gene discovery. PERADIGM employs an embedding model to capture relationships between ICD-10 codes, providing a nuanced representation of individual phenotypes. By utilizing patient similarity scores, it enhances the identification of candidate genes associated with disease-specific phenotypes, surpassing conventional methods that rely on binary disease status. We applied PERADIGM to the UK Biobank dataset for three rare diseases: autosomal dominant polycystic kidney disease (ADPKD), Marfan syndrome, and neurofibromatosis type 1 (NF1). PERADIGM identified additional candidate genes associated with ADPKD-related and Marfan syndrome-related phenotypes, some of which are supported by existing literature, and demonstrated enhanced signal detection for NF1-specific phenotypes beyond traditional methods. Our findings demonstrate the potential of PERADIGM to identify genes associated with rare diseases and related phenotypes by incorporating phenotype embeddings and patient similarity, providing a powerful tool for precision medicine and a deeper understanding of rare disease genetics and clinical manifestations.

genetics↗

bioRxiv2025

National Scale Genomic Engine for Precision Medicine: Singapore PRECISE-SG100K Experience

The whole-of-nation National Precision Medicine (NPM) programme in Singapore aims to integrate a genomics backbone into standard healthcare. Phase II recently completed the sequencing of 102,202 genomes from the diverse Asian population of Singapore, generating the PRECISE-SG100K dataset. The scaling from the pilot SG10K_Health phase to national production scale demanded a central shift: from earlier generation workflows to an integrated, industrial grade "Genomic Engine". The initiative was co-developed under a public-private partnership model. Our Genomic Engine was designed with an automation-first approach, as a modular platform linking a purpose-built, centralized biospecimen processing and DNA extraction facility (EXTRACT Lab), a high-throughput sequencing operation (Illumina Data Generation Infrastructure, IDGI), with the cloud-native analytics (GRIDS) running on a fully integrated PRECISE Sample Tracking System (STS). Here we describe the key design features of our Genomic Engine. Highlighting the Stress Test phase where the automated DNA extraction yields failed to meet expectations. We share the operational adjustments which were implemented to stabilize production and resulted in a three-fold reduction in sample triage due to improved DNA recovery. Without establishing the operational guardrails during the Stress Test, the Engine would not have reached a steady-state production evidenced by >3,000 genomes per month, median turnaround time 11 days, 99.6% operational success, within in six months. This framework offers a practical blueprint for other population-scale initiatives seeking to bridge the gap between scientific piloting and industrial delivery.

genomics↗

bioRxiv2025

Mutation of an insulin-sensitive Drosophila insulin-like receptor mutant requires methionine metabolism reprogramming to extend lifespan

Insulin/insulin growth factor signaling is a conserved pathway that regulates lifespan across many species. Multiple mechanisms are proposed for how this altered signaling slows aging. To elaborate these causes, we recently developed a series of Drosophila insulin-like receptor (dInr) mutants with single amino acid substitutions that extend lifespan but differentially affect insulin sensitivity, growth and reproduction. Transheterozygotes of canonical dInr mutants (Type I) extend longevity and are insulin-resistant, small and weakly fecund. In contrast, a dominant mutation (dInr353, Type II) within the Kinase Insert Domain (KID) robustly extends longevity but is insulin-sensitive, full-sized, and highly fecund. We applied transcriptome and metabolome analyses to explore how dInr353 slows aging without insulin resistance. Type I and II mutants overlap in many pathways but also produce distinct transcriptomic profiles that include differences in innate immune and reproductive functions. In metabolomic analyses, the KID mutant dInr353 reprograms methionine metabolism in a way that phenocopies dietary methionine restriction, in contrast to canonical mutants which are characterized by upregulation of the transsulfuration pathway. Because abrogation of S-adenosylhomocysteine hydrolase blocks the longevity benefit conferred by dInr353, we conclude the methionine cycle reprogramming of Type II is sufficient to slow aging. Metabolomic analysis further revealed the Type II mutant is metabolically flexible: unlike aged wildtype, aged dInr353 adults can reroute methionine toward the transsulfuration pathway, while Type I mutant flies upregulate the trassulfuration pathway continuously from young age. Altered insulin/insulin growth factor signaling has the potential to slow aging without the complications of insulin resistance by modulating methionine cycle dynamics. Author SummaryMutations in the invertebrate insulin/IGF signaling system robustly extend lifespan. Yet these interventions often cause insulin resistance, reduce growth, and impair fertility. In contrast, a dominant gain-of-function mutation in the kinase insert domain of the Drosophila insulin/IGF receptor extends lifespan while maintaining insulin sensitivity, growth, or reproduction. Here, we demonstrate this unique mutation reprograms methionine metabolism pathway in a way that mirrors dietary methionine restriction, which is known to extend lifespan in several animals including Drosophila and mice. Genetic epistasis analysis verifies this methionine cycle inhibition is an underlying cause for how the kinase insert domain mutation slows Drosophila aging while maintaining insulin-sensitivity, growth, and fecundity.

physiology↗

bioRxiv2025

PRRSV evades innate immune cGAS-STING antiviral function via its Nsp5 to deter STING translocation and activation

Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) is an important pathogen that seriously endangers pig breeding, causing significant economic losses to the global swine industry. Our previous study found that the DNA sensing innate cGAS-STING signaling pathway plays an important role in inducing interferon (IFN) upon PRRSV infection and inhibition of PRRSV replication. However, the immune evasion and its mechanism by PRRSV is still unclear. In current study, we found that PRRSV Non-structural protein 5 (Nsp5) strongly inhibits the cGAS-STING-IFN antiviral response. Further, we found that Nsp5 interacts with STING, blocking STING transport from ER to Golgi apparatus and interfering with STING recruitment of TBK1/IKK{varepsilon}/IRF3. Finally, we demonstrated that the Nsp5 36-47 and 58-67 amino acid regions are critical regions for inhibiting STING activity and PRRSV replication. The study describes a novel mechanism by PRRSV for suppression of the host innate antiviral response and have implications for our understanding of PRRSV pathogenesis. Author summaryThe innate immune system is the first line of host defense against infections. PRRSV is an immunosuppressive pathogen that has developed different strategies to evade the host innate immune responses. Here, we demonstrate for the first time that PRRSV utilizes its non-structural protein Nsp5 to inhibit the innate immune cGAS-STING-IFN signaling pathway, promoting the viral replication. Nsp5 can inhibit the cGAS-STING pathway by interacting with STING, blocking STING transport from ER to Golgi apparatus and interfering with STING recruitment of TBK1/IKK{varepsilon}/IRF3. In particular, the 36-47 and 58-67 amino acid regions of the Nsp5 is indispensable. This information will help us understand PRRSV-host interactions and provides new insights into the prevention and control of PRRSV.

immunology↗

bioRxiv2025

Layered stomatal immunity contributes to resistance of Vitis riparia against downy mildew Plasmopara viticola

Downy mildew (DM), caused by Plasmopara viticola, is one of the most serious grapevine diseases. Resistant grapevines are a well-known tool for mitigating pathogen-caused damage. We evaluated 29 global grapevine cultivars from 7 species for the sensitivity to P. viticola. Chardonnay belonging to the sensitive species V. vinifera and Qingdahean belonging to the well-known resistant species V. riparia were chosen for further investigation on the resistance mechanism against DM. Unlike Chardonnay, Qingdahean exerted an inhibitory effect on stomatal targeting, suppression of stomatal closure, stomatal penetration of P. viticola, and the development of primary hyphae and haustoria during the early phase of infection, and contained higher levels of malondialdehyde (MDA), which was significantly increased by P. viticola infection, toxic to the pathogen and had an interfering effect on the stomatal targeting. Furthermore, Qingdahean resisted pathogen invasion through the rapid induction of guard cell death and the hypersensitive responses (HR) of other cell types. These findings suggest that resistance to P. viticola consists of layered stomatal immunity in addition to the well-known HR in V. riparia, which is overcome by the pathogen in V. vinifera.

plant biology↗

bioRxiv2025

Ultradeep N-glycoproteome Atlas of Mouse Reveals Spatiotemporal Signatures of Brain Aging and Neurodegenerative Diseases

The current depth of site-specific N-glycoproteomics is insufficient to fully characterize glycosylation events in biological samples. Herein, we achieved an ultradeep and precision analysis of the N-glycoproteome of mouse tissues by integrating multiple workflows. The largest N-glycoproteomic dataset to date was established on mice, which contained 91,972 precursor glycopeptides, 62,216 glycoforms, 8,939 glycosites and 4,563 glycoproteins. The database consisted of 6.8 million glyco-spectra (containing oxonium ions), among which 160,928 were high-quality spectra with confident N-glycopeptide identifications. The large-scale and high-quality dataset enhanced the performance of current artificial intelligence models for glycopeptide tandem spectrum prediction. Using this ultradeep dataset, we observed tissue specific microheterogeneity and functional implications of protein glycosylation in mice. Furthermore, the region-resolved brain N-glycoproteomes for Alzheimers Diseases, Parkinson Disease and aging mice revealed the spatiotemporal signatures and distinct pathological functions of the N-glycoproteins. A comprehensive database resource of experimental N-glycoproteomic data from this study and previous literatures were further established. This N-glycoproteome atlas serves as a promising tool for revealing the role of protein glycosylation in biological systems.

neuroscience↗

bioRxiv2025

CHIP protects lysosomes from CLN4 mutant-induced membrane damages

Understanding how cells mitigate lysosomal damage is critical for unraveling pathogenic mechanisms of lysosome-related diseases. Here we use organelle-specific proteomics in iPSC-derived neurons (i3Neuron) and an in vitro lysosome-damaging assay to demonstrate that lysosome damage, caused by the aggregation of Ceroid Lipofuscinosis Neuronal 4 (CLN4)-linked DNAJC5 mutants on lysosomal membranes, serves as a critical pathogenic linchpin in CLN4-associated neurodegeneration. Intriguingly, in non-neuronal cells, a ubiquitin-dependent microautophagy mechanism downregulates CLN4 aggregates to counteract CLN4-associated lysotoxicity. Genome-wide CRISPR screens identify the ubiquitin ligase CHIP as a central microautophagy regulator that confers ubiquitin-dependent lysosome protection. Importantly, CHIPs lysosome protection function is transferrable, as ectopic CHIP improves lysosomal function in CLN4 i3Neurons, and effectively alleviates lipofuscin accumulation and neurodegeneration in a Drosophila CLN4 disease model. Our study establishes CHIP-mediated microautophagy as a key organelle damage guardian that preserves lysosome integrity, offering new insights into therapeutic development for CLN4 and other lysosome-related neurodegenerative diseases.

cell biology↗

bioRxiv2025

Pooled screening identifies combinatorial CAR signaling domains for next-generation CAR-M immunotherapies

Chimeric antigen receptor-engineered macrophages (CAR-Ms) hold great promise for solid tumor immunotherapy. The intracellular domains (ICDs) of CARs determine the phenotypic output of therapeutic macrophages but remain largely unexplored. Here, we constructed a CAR library containing 131 unique signaling domains derived from native immune receptors and identified 17 ICDs that enhance macrophage phagocytosis, inflammatory responses, or tumor infiltration in vitro and in vivo. We further developed a scalable 3 barcode technology, CARode, to uniquely label and trace ICD variants within large-scale combinatorial CAR library and applied it to single-cell RNA sequencing and single-cell CAR analysis to assess the synergetic effects of ICD combinations on macrophage activation. Our approach uncovered a novel CD40-LY9-FCRL1 chimeric receptor that modulates the tumor microenvironment and improves solid tumor clearance. In conclusion, our findings demonstrate that pooled screening can accelerate the discovery of complex ICD constructs, providing a powerful platform for engineering macrophage-based immunotherapies.

bioengineering↗

bioRxiv2025

DNAHX: a novel, non-motile dynein heavy chain subfamily, identified by cryo-EM endogenously

Ciliogenesis and cilia motility rely on the coordinated actions of diverse dyneins, yet the complexity of these motor proteins in cilia has posed challenges for understanding their specific roles. Traditional evolutionary analyses often overlook key family members due to technical limitations. Here, we present a cryo-EM-based, bottom-up approach for large-scale, de novo protein identification and functional prediction of endogenous axonemal dynein complexes. This approach led to the identification of a novel dynein heavy chain subfamily (XP_041462850), designated as DNAHX, from sea urchin sperm. Phylogenetic analysis indicates that DNAHX branches from the outer-arm dynein alpha chain during evolution and is found in specific animal lineages with external fertilization. DNAHX contains multiple insertions throughout the protein, locking DNAHX permanently in a pre-powerstroke state. The AAA1 site exhibits poor conservation of essential ATPase motifs, consistent with DNAHXs non-motile nature. DNAHX also forms a heterodimeric dynein complex, which we named dynein-X, with another dynein heavy chain and accessory chains. Furthermore, a subset of dynein-X displays an autoinhibited phi particle conformation, potentially facilitating the intraflagellar transport of axonemal dyneins. Our discovery of the novel, non-motile dynein heavy chain and the dynein-X complex provides valuable insights into the evolution of dyneins and potentially their diverse cellular functions.

biophysics↗

bioRxiv2025

Decoding the Molecular Language of Proteins with Evola

Proteins, natures intricate molecular machines, are the products of billions of years of evolution and play fundamental roles in sustaining life. Yet, deciphering their molecular language--understanding how sequences and structures encode biological functions--remains a cornerstone challenge. Here, we introduce Evolla, an interactive protein-language model designed to transcend static classification by interpreting protein function through natural language queries. Trained on 546 million protein-text pairs and refined via Direct Preference Optimization, Evolla couples high-dimensional molecular representations with generative semantic decoding. Benchmarking establishes Evollas superiority over general large language models in functional inference, demonstrates zero-shot performance parity with the state-of-the-art supervised model, and exposes remote functional relationships invisible to conventional alignment. We validate Evolla through two distinct applications: identifying candidate eukaryotic signature proteins in Asgard archaea, with functional Vps4 homologs validated via yeast complementation; and interactively discovering a novel deep-sea polyethylene terephthalate (PET) hydrolase, PsPETase, confirmed to degrade plastic films. These results position Evolla not merely as a predictor, but as a generative engine capable of complex hypothesis formulation, shifting the paradigm from static annotation to interactive, actionable discovery. The Evolla online service is available at http://www.chat-protein.com/.

bioinformatics↗

bioRxiv2024

Tax1bp1 enhances bacterial virulence and promotes inflammatory responses during Mycobacterium tuberculosis infection of alveolar macrophages

Crosstalk between autophagy, host cell death, and inflammatory host responses to bacterial pathogens enables effective innate immune responses that limit bacterial growth while minimizing coincidental host damage. Mycobacterium tuberculosis (Mtb) thwarts innate immune defense mechanisms in alveolar macrophages (AMs) during the initial stages of infection and in recruited bone marrow-derived cells during later stages of infection. However, how protective inflammatory responses are achieved during Mtb infection and the variation of the response in different macrophage subtypes remain obscure. Here, we show that the autophagy receptor Tax1bp1 plays a critical role in enhancing inflammatory cytokine production and increasing the susceptibility of mice to Mtb infection. Surprisingly, although Tax1bp1 restricts Mtb growth during infection of bone marrow-derived macrophages (BMDMs) (Budzik et al. 2020) and terminates cytokine production in response to cytokine stimulation or viral infection, Tax1bp1 instead promotes Mtb growth in AMs, neutrophils, and a subset of recruited monocyte-derived cells from the bone marrow. Tax1bp1 also leads to increases in bacterial growth and inflammatory responses during infection of mice with Listeria monocytogenes, an intracellular pathogen that is not effectively targeted to canonical autophagy. In Mtb-infected AMs but not BMDMs, Tax1bp1 enhances necrotic-like cell death early after infection, reprogramming the mode of host cell death to favor Mtb replication in AMs. Tax1bp1s impact on host cell death is a mechanism that explains Tax1bp1s cell type-specific role in the control of Mtb growth. Similar to Tax1bp1-deficiency in AMs, the expression of phosphosite-deficient Tax1bp1 restricts Mtb growth. Together, these results show that Tax1bp1 plays a crucial role in linking the regulation of autophagy, cell death, and pro-inflammatory host responses and enhancing susceptibility to bacterial infection. Author SummaryAlthough macrophages are the first innate immune cells to encounter Mycobacterium tuberculosis during infection, M. tuberculosis has evolved the ability to persist in them. Recent studies highlight that some types of macrophages are more permissive to M. tuberculosis replication and survival than others, but the mechanisms for cell type-specific differences in M. tuberculosis growth are only beginning to be understood. We found that the host factor, Tax1bp1 (Tax-1 binding protein 1), supports M. tuberculosis growth during animal infection and in specific subsets of innate immune cells, including alveolar macrophages while restricting M. tuberculosis in bone marrow-derived macrophages. We also found that Tax1bp1 has a similar phenotype in enhancing the pathogenesis of another intracellular pathogen, Listeria monocytogenes. Compared to bone marrow-derived macrophages, in alveolar macrophages, Tax1bp1 enhances the release of inflammatory mediators and leads to necrotic-like host cell death, which is known to enhance M. tuberculosis growth. Phosphorylation of Tax1bp1 in alveolar macrophages promotes M. tuberculosis growth. Our research highlights that Tax1bp1 is a host target for host-directed therapy against M. tuberculosis and controls host responses to M. tuberculosis in a cell type-specific manner.

microbiology↗

bioRxiv2024

RetS senses neurotransmitter norepinephrine and adrenergic agents to regulate Pseudomonas aeruginosa virulence

Bacterial cells employ diverse mechanisms to perceive and respond to host-associated molecules, coordinate intracellular signal transduction, and engage in interkingdom communications. In this study, we firstly discovered that the adrenergic antagonist dronedarone (DDR) effectively suppresses quorum sensing (QS) and pathogenesis of the opportunistic pathogen P. aeruginosa. Further analysis revealed that DDR targets RetS, an bacterial surface receptor critical for suppressing the GacS/GacA two-component regulatory system (TCS). Interestingly, the adrenergic agonist norepinephrine (NE) was found to exert opposite effects against RetS, thereby reversing DDR-induced phenotypes in P. aeruginosa. Moreover, we identified the hinge region of RetSDISMED2 serves as a binding pocket of NE and DDR, which provides potential mechanisms of the antagonistic effect of NE and DDR. Our study demonstrated that NE promotes P. aeruginosa infections while DDR can serve as an anti-virulence compound against P. aeruginosa infections. Thus, RetS might serve as a bacterial adrenergic receptor which keeps the NE under constant surveillance. SignificanceSensing and recognition of host-derived molecules, as well as therapeutic drugs, are crucial for the survival of pathogens during infection process. The adrenergic drugs are well known to affect virulence and pathogenesis of the opportunistic pathogen Pseudomonas aeruginosa (P. aeruginosa). In this work, we have provided in-depth characterization of a Norepinephrine (NE) receptor in P. aeruginosa, the RetS membrane protein. NE is the-adrenergic agonist known as a stress hormone, exerts its activity through interaction with mammalian adrenergic receptors (AR). It has been shown that increased NE concentration during early sepsis can stimulate the growth of various Gram-negative and Gram-positive bacteria, including P. aeruginosa. We revealed that both adrenergic agonist NE and another adrenergic antagonist dronedarone (DDR) are able to interfering the RetS-GacS/GacA signaling cascade of P. aeruginosa and execute opposite impact on the pathogenesis of P. aeruginosa during infections.

microbiology↗

bioRxiv2024

Accurate prediction of CRISPR editing outcomes in somatic cell lines and zygote with few-shot learning

The CRISPR-Cas system has revolutionized gene editing, while its outcome prediction remains unsatisfactory, especially in new cell states due to their distinct DNA repair preferences. In this study, we introduce inDecay, a flexible system for predicting CRISPR editing outcomes from target sequence, returning probabilities of nearly the full spectrum of indel events. Uniquely, inDecay utilizes informative and parameter-efficient features for each indel event and incorporates cell-type-specific repair preferences through a multi-stage design. While both inDecay and existing methods achieve accurate results for prediction within cell lines, only inDecay with transfer learning can retain the high performance for cross-cell line prediction. We then applied inDecay to mouse embryo editing using our newly generated data and observed remarkable accuracy by including as few as 30 fine-tuning embryonic samples. Notably, inDecay is the first software to predict embryonic editing. Therefore, our few-shot learning-supported system may accelerate guide RNA prioritization in mouse model generation, mammal embryonic gene editing, and cellular therapeutics.

bioinformatics↗

bioRxiv2024

Phospholipid signaling and multivesicular endolysosomes modulate membrane dynamics at the biotrophic interface in Rice Blast

Magnaporthe oryzae, the rice blast pathogen, secretes an arsenal of apoplastic and cytoplasmic effector proteins to suppress host immunity during the biotrophy phase. The proposed non-conventional secretion of cytoplasmic effectors into the Biotrophic Interfacial Complex (BIC), remains uncharted except for the involvement of the exocyst complex therein. Recently, the plant endocytic machinery has been implicated in translocation of cytoplasmic effectors to the host. Here, we used live cell imaging and mutant analyses to provide new insight into late endolysosomal mediators as conduits for establishment of the BIC and/or unconventional secretion across the host interface. Rerouting of the late endocytic GTPase Rab7 occurs at the BIC; and the dominant negative Rab7T22N led to significant defects in focal interface formation. FYVE biosensor and pharmacological interventions revealed that Rab7 membrane trafficking and establishment/integrity of the biotrophy interface requires the PI3 kinase Vps34 and phosphoinositide PI3(P) function. Additional pharmacological evidence supports endolysosomal trafficking to the host interface follows a distinct non-conventional secretion route too. Finally, we show that the Multi-Vesicular Bodies and lysosomal membranes act as downstream modules of the Rab7 function in precise sorting and specialised traffic of cytoplasmic effectors across the Magnaporthe-Rice interface during blast disease.

cell biology↗

bioRxiv2024

Phase Separation to Resolve Growth-Related Circuit Failures

Fluctuations in host cell growth poses a significant challenge to synthetic gene circuits, often disrupting circuit function. Existing solutions typically rely on circuit redesign with alternative topologies or additional control elements, yet a broadly applicable approach remains elusive. Here, we introduce a new strategy based on liquid-liquid phase separation (LLPS) to stabilize circuit performance. By engineering a self-activating circuit with transcription factors (TF) fused to an intrinsically disordered region (IDR), we enable the formation of TF condensates at the promoter region, maintaining local TF concentration despite growth-mediated dilution. This condensate formation preserves bistable memory in the self-activating circuit, demonstrating that phase separation can robustly counteract growth fluctuations, offering a novel design principle for resilient synthetic circuits.

synthetic biology↗