Biology subjects

Xiong, L.

Publications and source records attributed to Xiong, L..

At least 19 recordsLinked to original sources

bioRxiv2026

Monothiol glutaredoxin DaGrxB couples iron homeostasis with redox balance to orchestrate development and virulence in the peach shoot blight fungus Diaporthe amygdali

Peach shoot blight (PSB), caused by Diaporthe amygdali, is one of the most devastating fungal diseases affecting peach production worldwide. Glutaredoxins (Grxs) are ubiquitous redox-regulatory proteins involved in antioxidant defense and iron metabolism, yet their functions in phytopathogenic fungi remain largely unknown. Here, we identified four Grxs (DaGrxA-DaGrxD) in D. amygdali. Among them, DaGrxB, a monothiol glutaredoxin, is the sole member essential for vegetative growth, asexual reproduction and virulence. DaGrxB interacts with iron-regulatory transcription factors DaHapX and DaSreA and modulates their activities via iron-dependent nucleocytoplasmic shuttling, coordinating iron acquisition and storage gene expression. During infection, D. amygdali encounters a biphasic iron environment: an inherently iron-limited host environment at early stages (1-2 dpi) followed by host-mediated iron excess (3-4 dpi), and DaGrxB is essential for navigating this transition. Consistently, deletion of either DaHAPX or DaSREA significantly reduced virulence, confirming that iron homeostasis is a critical determinant of D. amygdali virulence. Notably, deletion of DaGRXB disrupts Fe-S cluster biosynthesis, causing mitochondrial Fe2+ accumulation, impaired aconitase activity, and reduced ATP production, which triggers constitutive activation of the GSH-dependent antioxidant system, with elevated GSH/GSSG ratios, increased GR and GST activities, and reduced H2O2 levels. Exogenous GSH and H2O2 depletion both reduced virulence, suggesting that disturbed redox balance directly contributes to attenuated virulence. Additionally, the Trx domain assists the Grx domain in nucleocytoplasmic translocation through intramolecular interactions. Finally, LDH-delivered dsRNA targeting DaGRXB effectively controls PSB. Collectively, these findings reveal how a monothiol glutaredoxin integrates iron homeostasis, redox balance and virulence in a plant pathogen, providing a foundation for RNAi-based disease management.

pathology↗

bioRxiv2026

NLR DNA copy-number expansion outpaces accumulation of predicted coding-intact copies

Whether nucleotide-binding leucine-rich repeat (NLR) DNA copies and predicted coding-intact copies accumulate proportionally within homologous neighbourhoods remains unresolved. We compared these counts across 173 anchor-defined neighbourhoods in 11 Capsicum genomes. After accounting for neighbourhood and genome effects, predicted coding-intact copies increased less than proportionally with DNA copies. DNA-copy doubling corresponded to 1.48-fold and 1.84-fold increases in expected coding counts under strict and inclusive definitions, respectively. The strict association persisted across sequence and sampling controls, with highly expanded arrays and heterogeneous coding recovery affecting interpretation. An independently defined panel of 17 Arabidopsis thaliana accessions reproduced sublinear scaling using curated gene counts. Matched pepper RNA-seq showed a transcript-support gradient across predicted coding states. At DH06, a near-megabase segmental duplication in C. pubescens contained intact and disrupted family pairs. Copy-specific HiFi reads placed the same F396 coding lesions in both physical blocks. Together, these results reveal a recurrent relationship between NLR array size and coding composition in the surveyed plant systems. Interpreting DNA expansion together with coding state provides a more informative basis for comparing NLR repertoires and prioritizing immune-gene candidates.

evolutionary biology↗

bioRxiv2026

DNA methylation regulates a key sex differentiation gene in Pogona vitticeps, a dragon lizard with sex reversal

During embryonic development bipotential gonads differentiate into either testes or ovaries under the direction of mutually exclusive gene networks. DNA methylation has been shown to regulate gene expression and to play a role in many developmental processes. This includes sex differentiation in which DNA methylation is linked to control of key sex genes. Whether DNA methylation regulates sexual differentiation in species where environmental influences such as temperature are involved is less clear. We conducted a genome-wide study in embryonic gonads of the central bearded dragon (Pogona vitticeps), a lizard with temperature induced sex reversal, and compared DNA methylation patterns to gene expression profiles at a stage of early sex differentiation. Overall, sex reversed ZZf females were found to have lower global methylation than both canonical sexes, ZZm males and ZWf females. We found that the expression of a key gene in sex differentiation, Amh, is regulated via DNA methylation. Amh is a driver of testis differentiation and is repressed in genetically determined females, as well as in temperature sex-reversed females, by hypermethylation around its transcription start site. Although the trigger of ovary determination is different in both groups of females, one by genetic complement and one by a temperature signal, downstream regulation of gene expression converges and seems to follow similar mechanisms.

genomics↗

bioRxiv2026

ARID4 Modulates Stomatal ABA Signaling and Stress-Responsive Gene Expression to Confer Drought Tolerance in Arabidopsis

To identify novel regulators of plant drought tolerance, we performed a forward genetic screen using thermal infrared imaging to detect alterations in leaf temperature, a proxy for transpiration. This screen identified AT-rich Interacting Domain Protein 4 (ARID4), a protein involved in chromatin remodeling, as a modulator of transpiration. arid4 loss-of-function mutants exhibited cooler leaves, increased water loss from detached leaves, and heightened susceptibility to drought relative to wild-type plants. Stomata of arid4 mutants were hyposensitive to abscisic acid (ABA), displaying reduced ABA-induced reactive oxygen species (ROS) production and impaired stomatal closure. Transcriptome profiling under drought revealed extensive misregulation of genes involved in stress-responses, redox homeostasis, iron acquisition, and photosynthesis. These findings indicate that ARID4 modulates the expression of key regulatory genes to maintain redox homeostasis and photosynthetic efficiency, thereby conferring drought stress tolerance.

Plant Biology↗

bioRxiv2026

The Function of LncRNA DRIR in Freezing Tolerance by Promoting Autophagic Degradation of CP29A and CP29B to Alter Alternative Splicing Patterns of Pre-mRNAs

O_LIResearch on the functions and molecular mechanisms of long non-coding RNAs (lncRNAs) involved in regulating plant freezing tolerance is still in its infancy. Our previous research work identified that lncRNA DROUGHT INDUCED LNCRNA (DRIR) regulates gene expressions in Arabidopsis. However, the underlying molecular mechanism is still unknown. C_LIO_LIThis study demonstrates that lncRNA DRIR regulates plant freezing tolerance by affecting alternative splicing patterns of pre-mRNAs. C_LIO_LIThrough chromatin isolation by RNA purification followed by mass spectrometry (ChIRP-MS), we identified two DRIR interacting proteins: CP29A and CP29B. We showed that the drirD mutant, which exhibits elevated DRIR expression and DRIR overexpression lines showed increased sensitivity to freezing stress, whereas DRIR RNAi lines were more tolerant to the stress. CP29A and CP29B bind to nuclear transcripts and, together with DRIR, regulate pre-mRNA alternative splicing under freezing stress. Notably, DRIR induces the relocalization of CP29A and CP29B to autophagosomes, leading to autophagy-mediated protein degradation. C_LIO_LICollectively, our findings elucidate the molecular mechanism by which DRIR influences the autophagy-based degradation of its binding proteins CP29A and CP29B, thereby regulating plant freezing tolerance by altering the alternative splicing patterns of pre-mRNAs, providing novel insights into the functions and mechanisms of lncRNAs in plants adapting to freezing environments. C_LI

plant biology↗

bioRxiv2026

Genos-m: a foundation model for human-associated microbial genomes

Human-associated microbial genomes encode extensive strain-level diversity and niche-specific gene repertoires that are critical to host health. However, these complex sequence features remain difficult to capture using general-purpose DNA foundation models, highlighting the need for dedicated representation learning tailored to the human microbiome. Here, we introduce Genos-m, an open-source foundation model for human-associated microbial genome representation. Genos-m was pretrained on approximately 1.2 trillion nucleotide tokens from a curated microbial genome corpus, including human-associated prokaryotic isolates, high-quality metagenome-assembled genomes (MAGs) and bacteriophages, supplemented with GTDB species-level representative genomes to broaden prokaryotic taxonomic breadth. The model uses a sparsely activated Mixture-of-Experts (MoE) Transformer architecture, with 4.7 billion total parameters, approximately 330 million activated parameters per forward pass and a maximum context length of one million base pairs. We evaluated frozen Genos-m representations across short-sequence and gene-level tasks, biosynthetic gene cluster (BGC)-based regional sequence tasks, whole-genome strain phenotype prediction, and zero-shot transfer on prokaryote-related RNAfitness assays. Across these benchmarks, Genos-m consistently ranked among the leading comparison models, with the best performance in five of eight gene-fitness regression tasks and in BGC type classification. Using sparse autoencoders, we identified sparse features in Genos-m hidden activations that aligned with annotated ORFs, intergenic regions, and tRNA and rRNA loci. In downstream applications, Genos-m-derived genome-informed species representations in-corporated into a human microbiome self-supervised learning model improved colorectal cancer (CRC)-control classification over conventional species-abundance random forest models. Genos-m also generated stable sample-level embeddings from as few as 10,000 metagenomic reads, retaining gut microbial community structure that distinguished geographic origin and aligned with enterotypes defined from full-depth taxonomic profiles. Together, these results support Genos-m as a reusable representation model for microbial genomes and metagenomes, with conclusions bounded by the reported datasets, task definitions and evaluation protocols. Genos-m model weights, inference code, and usage documentation are publicly available on GitHub (https://github.com/BGI-HangzhouAI/Genos-m) and Hugging-Face (https://huggingface.co/BGI-HangzhouAI/Genos-m).

bioinformatics↗

bioRxiv2026

A selective Cullin 3 RING E3 ligase inhibitor attenuates hyperglycemia via dual insulin sensitizing and insulinotropic action

Hyperglycemia is a hallmark of type-2 diabetes and a key pathogenic driver of diabetic complications. Cullin RING E3 ligases (CRLs) are multi-subunit E3 ubiquitin ligases that mediate cellular protein turnover. The activity of CRLs requires cullin neddylation, a post-translational modification that can be pharmacologically targeted with therapeutic potentials. By using hyperinsulinemic-euglycemic clamp analysis, we discover that pan neddylation inhibitor exerts both insulin sensitization effect in liver and muscle and insulinotropic effect in pancreatic {beta} cells. This dual action is mediated by Cullin 3 (Cul3), a member of the 7 canonical cullin family proteins. DI-1859, a selective Cul3 neddylation inhibitor, effectively protects against hyperglycemia in obese mice. DI-1859 enhances insulin signaling by preventing Cul3-mediated insulin receptor substrate degradation in liver and muscle cells. DI-1859 increases insulin secretion in a glucagon-like peptide-1-independent manner in mice and directly potentiates glucose-stimulated insulin secretion in INS-1 832/13 {beta} cells and human islets. Mechanistic studies reveal that DI-1859 does not promote glycolytic flux or bioenergetics function but potentiates glucose-stimulated insulin secretion via mechanisms involving RhoA activation and cytoskeleton remodeling in {beta} cells. This study shows that a single agent targeting Cul3 neddylation simultaneously promotes insulin sensitization and insulin secretion to attenuate hyperglycemia in mice. Article Highlightsa. Pan cullin neddylation inhibitors exhibit potent hypoglycemic effect. b. The target organs and mechanisms underlying the hypoglycemia effect of cullin pan neddylation inhibitors are incompletely understood. c. We found that inhibition of Cul3 leads to a dual insulin sensitization and insulinotropic effect. d. Selective inhibition of Cul3 neddylation is a feasible approach to lower hyperglycemia.

physiology↗

bioRxiv2026

Presynaptic control of top-down signaling in neocortical layer 1

The integration of bottom-up and top-down signals is crucial for perception and behavior. Neocortical layer 1 (L1) is a key target for top-down inputs and contains various GABAergic interneurons. Here, we investigated whether GABA can presynaptically regulate neurotransmitter release at top-down synapses targeting L1 of primary somatosensory cortex (S1). Our findings show that presynaptic GABAB receptor activation suppresses corticocortical inputs from primary motor (M1) and secondary somatosensory cortices (S2) more than those from the posterior medial nucleus of the thalamus (POm). This effect varied by target cell type, with GABAergic interneurons being less affected. Finally, we demonstrate that L1 neuron-derived neurotrophic factor (NDNF)-expressing interneurons inhibit top-down synapses more effectively than somatostatin-expressing interneurons, and that POm drives L1 neurogliaform cells, a subtype of NDNF interneuron that elicits unitary GABAB responses. These results reveal a novel circuit in which higher-order thalamus influences top-down corticocortical communication via L1 neurogliaform cells.

neuroscience↗

bioRxiv2025

Dissecting the homeodomain MAT locus and engineering novel tripolar and bipolar mating systems in Cryptococcus amylolentus

Sex in fungi is governed by the mating-type (MAT) locus, which exists as bipolar, pseudobipolar, or tetrapolar systems. The significance and impact of MAT on sexual reproduction, however, remain understudied. Furthermore, the evolution of fungal MAT loci shares features with the evolution of sex chromosomes in plants and animals. Pathogenic Cryptococcus species harbor a bipolar system with a large contiguous MAT locus, whereas closely related species, such as the non-pathogen C. amylolentus possess a tetrapolar system with unlinked P/R and HD loci. The HD locus encodes homeobox domain containing proteins that play an important and evolutionarily conserved role in sexual reproduction. Here, we explored the roles of HD genes in sexual reproduction and determined the implications of a tetrapolar to bipolar MAT transition. With a CRISPR-Cas9 system we developed for C. amylolentus, we generated gene deletion mutants and demonstrated that a single compatible Sxi1-Sxi2 pair is necessary and sufficient for mating. By relocating the HD genes to the P/R locus, we found that the artificially generated bipolar configuration led to defective sexual development, which could be partially restored through additional rounds of sexual reproduction. Transcriptomic profiling further revealed that a Sxi1-Sxi2 heterodimeric complex drives expression of genes required for DNA replication and ergosterol biosynthesis during sexual reproduction. These findings provide the first experimental demonstration of a tetrapolar-to-bipolar transition in a tetrapolar mating species, illuminating MAT locus evolution and homeodomain protein functions in Cryptococcus. ImportanceSexual reproduction is critical for fungal survival and adaptation, yet the mechanisms driving transitions between mating systems remain unclear. With Cryptococcus amylolentus, we provide the first experimental validation of a mating system transition from its original tetrapolar to an intermediate tripolar to a derived bipolar in a tetrapolar species. We show that HD heterodimers phenotypically govern dikaryotic filamentation and also transcriptionally modulate DNA replication. These findings establish a mechanistic basis for how MAT locus reorganization drives bipolar evolution from an ancestral tetrapolar state and reinforce that fertility depends on the coordinated control of MAT locus architecture and regulatory functions.

microbiology↗

bioRxiv2025

Life in the fast lane: Functional consequences of male-female dynamic differences in the renal autoregulation of flow

Tubuloglomerular feedback (TGF) is essential for the renal auto-regulation of flow. TGF is known to induce spontaneous oscillations in single-nephron tubular fluid flow in male kidneys. However, male-female differences in this dynamic behavior have not been studied. Leveraging intravital two-photon microscopy, resting-state magnetic resonance imaging, ultrasound-based and transdermal recordings, we found TGF-mediated oscillations across spatial scales in the rodent kidney, from single-nephron to whole-organ levels, and that male kidneys exhibited higher operating frequencies than females. To understand the mechanisms involved, we developed a dynamical systems model of TGF that agrees with physiological observations. Analysis of the mathematical model indicated that higher reabsorption rate and fluid flow efficiency in male proximal tubules not only result in higher frequencies, but also render male nephrons more susceptible to lose TGF-mediated oscillations. Furosemide abolished TGF-mediated oscillations in male kidneys and upregulated tubular injury marker KIM-1, suggesting that the propensity to lose TGF-mediated oscillations underlies the heightened risk for injury in males. Our analysis also indicated that SGLT-2 inhibition confers renoprotection by preserving TGF-mediated oscillations in hyperglycemia. Combining quantitative imaging and mathematical modeling, this study provides mechanistic insights into the transition from normal physiology to pathophysiology in the kidney.

physiology↗

bioRxiv2025

Cancer cell phagocytosis induces an anti-inflammatory gene regulatory program in macrophages

Macrophages are capable of eliminating cancer cells by phagocytosis, particularly in the presence of monoclonal antibody (mAb) therapies targeting tumor antigens. Paradoxically, tumor-associated macrophages are typically associated with poor patient outcome, and can promote tumor growth by secretion of immunosuppressive cytokines and growth factors. The mechanisms by which these pro-tumor macrophage states arise are poorly understood, and it is unclear how mAb-induced cancer cell phagocytosis may contribute to these states. To understand how antibody-dependent cancer cell phagocytosis (ADCP) alters macrophage state and function, we profiled gene expression and chromatin accessibility changes over time after ADCP. We observed that after ADCP, macrophages upregulate an anti-inflammatory gene regulatory program, characterized by expression of pro-angiogenic and immunosuppressive chemokine genes, and increased activity by cellular, oxidative, and lysosomal stress transcription factors. This gene regulatory program was shared among phagocytic macrophages following either ADCP or apoptotic cancer cell phagocytosis, in addition to substrate-specific pathways. Conditioned media from macrophages promoted EMT in cancer cells, but this pro-EMT macrophage phenotype was attenuated following ADCP, but not following apoptotic cancer cell phagocytosis. The phagocytic gene signature we identified in vitro is also expressed by tumor-associated macrophages across numerous cancer types in vivo. Together, this work identifies an anti-inflammatory and immunosuppressive epigenetic program in macrophages following ADCP upon mAb treatment, and expands our understanding of how phagocytosis influences macrophage heterogeneity in the tumor microenvironment.

cancer biology↗

bioRxiv2025

TRIM32 controls timely cell cycle exit in muscular differentiation through c-Myc down-regulation

Mutations in TRIM32 cause Limb-Girdle Muscular Dystrophy recessive 8 (LGMDR8), a neuromuscular disorder primarily affecting the proximal muscles of hips and shoulders. However, the precise pathogenic mechanism remains unclear. In this study, we used Trim32 knockout C2C12 murine myoblasts to investigate the impact of Trim32 loss on myogenesis. We found that Trim32 deficiency leads to impaired myogenic signaling and reduced expression of key myogenic regulatory factors ultimately leading to delayed and abnormal myotube formation. Further, we discovered that absence of Trim32 disrupts the transition from proliferation to differentiation by limiting the necessary down-regulation of the proto-oncogene c-Myc, thus delaying and altering the onset of differentation. Interestingly, unlike previous reports that emphasized protein-level regulation, our data reveal that Trim32 regulates c-Myc at mRNA stability level. Contrasting the sustained c-Myc level is able to partially recover the myogenesis defects observed in the absence of Trim32, suggesting that the Trim32-c-Myc axis may represent a promising therapeutic target for treating LGMDR8. Altogether these findings underscore the essential role of Trim32 in muscle regeneration within LGMDR8 pathogenesis.

cell biology↗

bioRxiv2025

A telomere to telomere phased genome assembly and annotation for the Australian central bearded dragon Pogona vitticeps

BackgroundThe central bearded dragon (Pogona vitticeps) is widely distributed in central eastern Australia and adapts readily to captivity. Among other attributes, it is distinctive because it undergoes sex reversal from ZZ genotypic males to phenotypic females at high incubation temperatures. Here, we report an annotated telomere to telomere phased assembly of the genome of a female ZW central bearded dragon. ResultsGenome assembly length is 1.75 Gbp with a scaffold N50 of 266.2 Mbp, N90 of 28.1 Mbp, 26 gaps and 42.2% GC content. Most (99.6%) of the reference assembly is scaffolded into 6 macrochromosomes and 10 microchromosomes, including the Z and W microchromosomes, corresponding to the karyotype. The genome assembly exceeds standard recommended by the Earth Biogenome Project (6CQ40): 0.003% collapsed sequence, 0.03% false expansions, 99.8% k-mer completeness, 97.9% complete single copy BUSCO genes and an average of 93.5% of transcriptome data mappable back to the genome assembly. The mitochondrial genome (16,731 bp) and the model rDNA repeat unit (length 9.5 Kbp) were assembled. Male vertebrate sex genes Amh and Amhr2 were discovered as copies in the small non-recombining region of the Z chromosome, absent from the W chromosome. This, coupled with the prior discovery of differential Z and W transcriptional isoform composition arising from pseudoautosomal sex gene Nr5a1, suggests that complex interactions between these genes, their autosomal copies and their resultant transcription factors and intermediaries, determines sex in the bearded dragon. ConclusionThis high-quality assembly will serve as a resource to enable and accelerate research into the unusual reproductive attributes of this species and for comparative studies across the Agamidae and reptiles more generally. Species TaxonomyEukaryota; Animalia; Chordata; Reptilia; Squamata; Iguania; Agamidae; Amphibolurinae; Pogona; Pogona vitticeps (Ahl, 1926) (NCBI:txid103695). Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=102 SRC="FIGDIR/small/651798v1_ufig1.gif" ALT="Figure 1"> View larger version (45K): org.highwire.dtl.DTLVardef@1a9d5fforg.highwire.dtl.DTLVardef@1206061org.highwire.dtl.DTLVardef@97acf7org.highwire.dtl.DTLVardef@1dc939d_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2025

Homeodomain protein Sxi1α regulates cell-cell fusion during distinct sexual reproduction modes in Cryptococcus deneoformans

Sex-specific homeodomain (HD) proteins are key regulators of cell identity and sexual development in fungi, typically functioning as heterodimers to govern transcription. In the human fungal pathogens Cryptococcus neoformans and Cryptococcus deneoformans, the HD proteins Sxi1 and Sxi2a (Sex-inducer 1 and 2a) have been characterized as interacting components that play critical roles in sexual development during x a sexual reproduction. cells are the predominant mating type in natural populations of Cryptococcus, and unisexual (same-sex) mating can also occur in certain genetic backgrounds. The roles of Sxi1 and Sxi2a in unisexual reproduction are not fully understood. To elucidate the functions of Sxi1 and Sxi2a, we first applied AlphaFold3 prediction, which identified potential heterodimeric and homodimeric complexes. Formation of a Sxi2a homodimer was then experimentally validated through yeast two-hybrid assays. We subsequently deleted SXI1 and SXI2a in the hyper-filamentous self-fertile C. deneoformans strains XL280 and XL280a. Disruption of these genes did not result in noticeable defects in vegetative growth, virulence-associated traits, colony morphology, sporulation, or competitive fitness during unisexual crosses. Interestingly, both bilateral (mutant x mutant) and unilateral (mutant x wildtype) crosses involving the sxi1{Delta} mutant significantly increased - cell fusion efficiency, suggesting a previously unrecognized inhibitory role for Sxi1 in regulating same-sex cell fusion. Consistently, genes encoding mating pheromones and the -pheromone receptor Ste3 were upregulated in the sxi1{Delta} fusion assays. Transcriptomic analysis of sxi1{Delta} and sxi2a{Delta} mutants led to the identification of unique subsets of genes negatively regulated by each transcription factor during unisexual reproduction. Additionally, x a crosses between null mutants of sxi1{Delta} and sxi2a{Delta} revealed differential regulation of mating-type (MAT) loci genes dependent only on Sxi1 or Sxi2a. Together, our findings reveal a novel role for Sxi1 in governing cell fusion and demonstrate that Sxi1 and Sxi2a have distinct transcriptional control during unisexual and x a sexual reproduction, potentially exerting opposing regulation of sex-specific MAT genes. Author SummaryAnalogous to processes observed in many eukaryotic organisms, fungal sexual reproduction reshuffles genetic material, producing novel genetic variants with increased fitness and/or adaptive potential. The human fungal pathogen Cryptococcus deneoformans is widespread in the environment and can cause severe infections in immunocompromised individuals. Sexual reproduction in this species occurs between cells of either opposite mating types ( x a mating) or the same mating type ( x and a x a unisexual reproduction). The mating type-specific homeodomain transcription factors Sxi1 and Sxi2a are known to form a complex that broadly controls sexual development following and a cell fusion. Here, we find that Sxi1 inhibits cell-cell fusion during unisexual reproduction, while its partner, Sxi2a, shows only limited inhibition in certain genetic contexts. Further, we show that Sxi1 and Sxi2a regulate transcription of distinct gene sets during both x a and unisexual reproduction, including many within the mating-type (MAT) loci, often in opposing manners. Together, these findings reveal that Sxi1 and Sxi2a function both cooperatively and independently, thereby allowing for a more flexible and finely-tuned mating control system than previously recognized, providing insight into how this organism generates genetic variation and adapts to changing environments.

microbiology↗

bioRxiv2024

Astrocyte-specific secretome profiling reveals its correlation with neurological disorders

Secreted proteins mediate intercellular communication throughout the lives of multicellular organisms. However, due to the lack of new technology for secreted protein capturing, the progress of "secretomics lags behind. Here, we report a two-step secretome enrichment method (tsSEM) combining unnatural amino acid labeling and click chemistry-based biorthogonal reaction, which enables in vitro secretome profiling in the presence of serum. Using this novel method, we systematically investigated the secretome of human iPSCs-derived astrocytes (iAst) in different disease models and identified a panel of astrocyte-secreted proteins that are responsible for its non-cell autonomous toxicity under disease conditions. Furthermore, we validated two astrocytes-derived novel neurotrophic proteins, FAM3C and KITLG, which we identified from disease models, and found that they could boost neurite outgrowth, protect neurons, and promote neural progenitor proliferation. Our study highlights the utility of secretome profiling of iAst and demonstrates its applications in disease study and target identification and validation in drug development.

neuroscience↗

bioRxiv2024

NLGN3 contributes to angiogenesis in myocardial infarction via activation of the Gαi1/3-Akt Pathway

BACKGROUNDAngiogenesis is an important repair mechanism for myocardial infarction. Neuroligin-3 (NLGN3) can promote angiogenesis by activating Gi1/3-Akt signaling following ischemic brain injury. This study investigated the role of NLGN3 in myocardial infarction (MI). METHODS AND RESULTSOn the 7th day after MI, the plasma level of NLGN3 in patients was significantly higher than in the control group. A mouse model of MI also showed significantly increased expression of NLGN3 in heart tissue. Single-nucleus transcriptome analysis revealed that NLGN3 was located predominantly in cardiac fibroblasts and endothelial cells (ECs). Endothelial-specific knockdown of NLGN3, or inhibition of NLGN3 using ADAM10i, significantly increased the ischemic area, reduced angiogenesis, and worsened cardiac function. Co-immunoprecipitation (Co-IP) experiments showed that NLGN3 interacted with Gi1/3. The Gi1/3 knockout (Gi1/3-KO) mouse model of MI showed an increased ischemic area, decreased angiogenesis, and impaired cardiac function. Mechanistic studies showed that the NLGN3-Gi1/3 signaling pathway exerts cardioprotective effects by promoting EC proliferation and tube formation through the PI3K-Akt-mTOR pathway. Silencing of Gi1/3 largely eliminated the ability of NLGN3-promoting cardiac ECs to proliferate and form tubes. CONCLUSIONOur findings suggest the endothelial NLGN3-Gi1/3 signaling pathway promotes angiogenesis and reduces the ischemic area following MI, which is critical for maintaining cardiac function and repairing tissues. Targeting of the NLGN3-Gi1/3 signaling pathway may have clinical therapeutic potential in protecting the heart from ischemic injury. What Is Known?O_LIMyocardial infarction (MI) leads to a significantly increased risk of recurrent events in individually vascularized regions, which then stimulates adverse systemic vascular effects. C_LIO_LIGi proteins play a key role in receptor tyrosine kinase (RTK) and other non-GPCR receptor-mediated signaling. C_LIO_LINeuroligin 3 (NLGN3) is a major member of the NLGN family of proteins that help to form synapses between neurons. It is known be cleaved and secreted in an activity-dependent manner, but its function in heart disease is still poorly understood. C_LI What New Information Does This Article Contribute?O_LINLGN3 expression was found to be markedly increased in the heart after MI, especially in cardiac ECs. C_LIO_LIKnockdown of Gi1/3 significantly reduced angiogenesis and impaired cardiac function, while knockdown of NLGN3 increased the ischemic area and reduced angiogenesis through Gi1/3. C_LIO_LINLGN3-Gi1/3 signaling promotes vascular EC proliferation and tube formation through the PI3K-Akt-mTOR pathway, thereby protecting the ischemic heart. C_LI

molecular biology↗

bioRxiv2024

B cells targeting parasites capture spatially linked antigens to secure T cell help

Our understanding of T-cell-dependent humoral responses has been largely shaped by studies involving model antigens such as recombinant proteins and viruses 1,2. In these contexts, B cells internalize the entire antigen or pathogen, and present a range of antigens to helper CD4+ T cells to initiate the humoral response. However, this model does not account for large pathogens (such as parasites) that are too large to be taken up by individual B cells, and the mechanisms by which B cells acquire and present antigens from large complex pathogens to T cells remain poorly understood. Here we used Plasmodium, the causative parasite of malaria, as a model to investigate the requirements for T cell help for B cells targeting the Plasmodium surface circumsporozoite protein (CSP). Upon Plasmodium sporozoite (SPZ) immunization, CSP-specific B cells can form a synapse-like structure with SPZs and take up CSP and non-CSP surface antigens. As a result, CSP-specific B cells can receive help from CD4+ T cells specific to antigens that are located on the surface but not cytosol of the Plasmodium SPZ. Therefore, B cells can obtain help, not only from T cells with the same protein specificity, but also from T cells specific for spatially linked antigens. This flexibility in T cell help may enhance the initiation and maintenance of humoral immune responses to complex pathogens.

immunology↗

bioRxiv2024

Covalent DNA-Encoded Library Workflow Drives Discovery of SARS-CoV-2 Non-structural Proteins Inhibitors

The global coronavirus disease 2019 (COVID-19) pandemic persists, with the ongoing mutation of the virus. Consequently, the development of inhibitors with diverse binding modes and mechanisms of action, along with the elucidation of novel binding sites is of paramount importance. The 3-chymotrypsin-like protease (3CLpro) and papain-like protease (PLpro) are two validated cysteine proteases that cleave the viral polyprotein and are essential for viral replication. In this study, we utilized covalent DNA-Encoded libraries (CoDELs) workflow to identify two series of triazine-based covalent inhibitors targeting 3CLpro and PLpro. Molecular docking facilitated the identification of optimization pathways, further refined through medicinal chemistry efforts, leading to the development of the non-peptide 3CLpro inhibitor LU9, which exhibited an IC50 value of 0.34 M, and crystal structure of LU10 revealed a unique binding mode within the active site. Additionally, the X-ray cocrystal structure of SARS-CoV-2 PLpro with XD5 uncovered a previously unexplored binding site, adjacent to the catalytic pocket, providing an opportunity for further development of PLpro inhibitors. Overall, these novel compounds serve as valuable chemical probes for target validation and represent promising drug candidates for the continued development of SARS-CoV-2 antivirals.

biochemistry↗