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Peng, C.

Publications and source records attributed to Peng, C..

At least 73 records · Page 4Linked to original sources
bioRxiv2024

Collaboration between IL-7 and IL-15 enables adaptation of tissue-resident and circulating memory CD8+ T cells

Interleukin-7 (IL-7) is considered a critical regulator of memory CD8+ T cell homeostasis, but this is primarily based on analysis of circulating and not tissue-resident memory (TRM) subsets. Furthermore, the cell-intrinsic requirement for IL-7 signaling during memory homeostasis has not been directly tested. Using inducible deletion, we found that Il7ra loss had only a modest effect on persistence of circulating memory and TRM subsets and that IL-7R was primarily required for normal basal proliferation. Loss of IL-15 signaling imposed heightened IL-7R dependence on memory CD8+ T cells, including TRM populations previously described as IL-15-independent. In the absence of IL-15 signaling, IL-7R was upregulated, and loss of IL-7R signaling reduced proliferation in response to IL-15, suggesting cross-regulation in memory CD8+ T cells. Thus, across subsets and tissues, IL-7 and IL-15 act in concert to support memory CD8+ T cells, conferring resilience to altered availability of either cytokine. HighlightsTissue-resident and circulating memory CD8+ T cells modestly decline after loss of IL-7R IL-7R is required for normal self-renewal of memory CD8+ T cells Combined loss of IL-7 and IL-15 causes a profound defect across memory CD8+ T cell subsets Cross-regulation of IL-7 and IL-15 signaling occurs in memory CD8+ T cells O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=70 SRC="FIGDIR/small/596695v1_ufig1.gif" ALT="Figure 1"> View larger version (17K): org.highwire.dtl.DTLVardef@8e439corg.highwire.dtl.DTLVardef@1f8e42forg.highwire.dtl.DTLVardef@14cfceforg.highwire.dtl.DTLVardef@171e4eb_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗
bioRxiv2024

Molecular basis for the host range function of the poxvirus PKR inhibitor E3

The antiviral protein kinase R (PKR) is activated by viral double-stranded RNA and phosphorylates translation initiation factor eIF2, thereby inhibiting translation and virus replication. Most poxviruses contain two PKR inhibitors, called E3 and K3 in vaccinia virus (VACV), which are determinants of viral host range. The prevailing model for E3 function is that it inhibits PKR through the non-specific sequestration of double-stranded (ds) RNA. Our data revealed that Syrian hamster PKR was resistant to E3, which is at odds with the sequestration model. However, Syrian hamster PKR was still sensitive to K3 inhibition. In contrast, Armenian hamster PKR showed opposite sensitivities, being sensitive to E3 and resistant to K3 inhibition. Mutational analyses of hamster PKRs showed that sensitivity to E3 inhibition was largely determined by the region linking the dsRNA-binding domains and the kinase domain of PKR, whereas two amino acid residues in the kinase domain (helix G) determined sensitivity to K3. Expression of PKRs in congenic cells showed that Syrian hamster PKR containing the two Armenian hamster PKR residues in helix-G was resistant to wild type VACV infection, and that cells expressing either hamster PKR recapitulated the phenotypes observed in species-derived cell lines. The observed resistance of Syrian hamster PKR to E3 explains its host range function and challenges the paradigm that dsRNA-binding PKR inhibitors mainly act by the sequestration of dsRNA. SignificanceThe molecular mechanisms that govern the host range of viruses are incompletely understood. A small number of poxvirus genes have been identified that influence the host range of poxviruses. We show that the host range functions of E3 and K3, two host range factors from vaccinia virus, are a result of species-specific interactions with the antiviral protein kinase R (PKR) and that PKR from closely related species displayed dramatic differences in their sensitivities to these viral inhibitors. While there is a substantial body of work demonstrating host-specific interactions with K3, the current model for E3-mediated PKR inhibition is that E3 non-specifically sequesters dsRNA to prevent PKR activation. This model does not predict species-specific sensitivity to E3; therefore, our data suggest that the current model is incomplete, and that dsRNA sequestration is not the primary mechanism for E3 activity.

microbiology↗
bioRxiv2024

Asymmetric binding of coactivator SRC1 to FXR-RXR and allosteric communication within the complex

Farnesoid X receptor (FXR) is a promising target for treatment of metabolic associated fatty liver disease (MAFLD). In this study, we employed an integrative approach to investigate the interaction between FXR-RXR-hSHP-1 complex and the entire coactivator SRC1-NRID (nuclear receptor interaction domain). We constructed a multi-domain model of FXR120-RXR98-hSHP-1, highlighting the interface between FXR-DBD and LBD. Using HDX-MS, XL-MS, and biochemical assays, we revealed the allosteric communications in FXR-RXR-hSHP-1 upon agonist and DNA binding. We then demonstrated that SRC1 binds only to the coactivator binding surface of FXR within the FXR-RXR heterodimer, with the NR-box2 and NR-box3 of SRC1 as the key binding motifs. Our findings, which provide the first model of SRC1-NRID in complex with FXR-RXR-hSHP-1, shed light on the molecular mechanism through which the coactivator asymmetrically interacts with nuclear receptors and provide structural basis for further understanding the function of FXR and its implications in diseases.

biophysics↗
bioRxiv2024

Wampee chromosome-level reference genome elucidates fruit sugar-acid metabolism

Wampee (Clausena lansium) is an economically significant subtropical fruit tree widely cultivated in Southern China. High-quality genomic resources are unavailable, but they are essential for functional genomics and germplasm enhancement of wampee. Here, we provide a chromosome-level genome sequence for the wampee cultivar JinFeng and a population genomic analysis of 266 accessions. The 297.1 Mb wampee genome, containing nine chromosomes with a scaffold N50 of 29.2 Mb and encoding 23,468 protein-coding genes, showed a significant improvement over the previous version. We dissected the wampee population structure and genetic differentiation in China using population genomic analysis, which detected 110 and 671 genes under a selective sweep associated with sour and sweet wampee evolution in domesticated clones, respectively. Homozygous non-synonymous single nucleotide polymorphisms are likely associated with fruit flavor differentiation. A genome-wide association study identified 220 remarkable marker-trait associations for total acid content, harboring 289 genes encoding transcription factors, transporters, and enzymes involved in sugar and acid metabolism, which are potentially useful for sour and sweet taste development in wampee fruit. Furthermore, the ethylene response factor family gene ClERF061 and the SWEET family gene ClSWEET7 were identified. Linkage assessment between the relative expression levels of ClERF061 or ClSWEET7 and the total acid/total sugar contents implied their potential involvement in sugar-acid metabolism in wampee fruits. High-quality genome resources are valuable for expediting wampee research and genome-assisted breeding.

genomics↗
bioRxiv2024

The iPhylo Suite: An Interactive Platform for Building and Annotating Biological and Chemical Taxonomic Trees

Accurate and rapid taxonomic classifications are essential for systematically exploring organisms and metabolites in diverse environments. Many tools have been developed for biological taxonomic trees, but limitations apply, and a streamlined method for constructing chemical taxonomic trees is lacking. We present the iPhylo suite (https://www.iphylo.net/), a comprehensive, automated, and interactive platform for biological and chemical taxonomic analysis. The iPhylo suite includes the Tree and Visual web-based modules for interactive building and annotating biological and chemical taxonomic trees, and a stand- alone command-line interface (CLI) module for local use and deployment on high-performance computing clusters. Furthermore, the iPhylo visualization module is fully implemented in R, allowing users to save progress locally and modify the underlying R codes. Finally, the iPhylo CLI module extends the analysis to all hierarchical relational databases. In summary, the iPhylo suite provides an integrated interactive framework for in-depth taxonomic analyses of biological and chemical features and beyond. HighlightsO_LIA platform for both biological and chemical classification analysis and visualizations. C_LIO_LIA stand-alone Command line interface module for local or HPC deployment. C_LIO_LIR-based interactive visualization module with session-saving options. C_LIO_LISupports customized hierarchical databases for analysis beyond taxonomy. C_LI

bioinformatics↗
bioRxiv2024

MALAT1 mediates miR-206/CDC42/PAK1/Paxillin signaling axis to alleviate erectile dysfunction in rats

Erectile dysfunction (ED) is a male sexual dysfunction with a gradually increasing prevalence, and current treatments are ineffective. This study aims to find a safer and more effective treatment for erectile dysfunction. Bone marrow-derived mesenchymal stem cells (BM-MSCs) treated with VEGFA. The expression of endothelial markers vWF, VE-cadherin and eNOS were determined by overexpressing MALAT1 and CDC42, respectively. The results showed that interference with CDC42 and MALAT1 expression inhibited the differentiation of BM-MSCs to ECs and the expression of endothelial marker-related proteins. Moreover, MALAT1 induced differentiation of BM-MCs to ECs through the CDC42/PAK1/Paxillin pathway was explored by transfecting si-MALAT1 and overexpressing CDC42 as well as PAK1 in the BM-MSCs. Next, miR-206 was overexpressed within BM-MCs to determine CDC42 expression. The binding sites of MALAT1, miR-206 and CDC42 were reported using luciferase. it was found that the MALAT1 competes with CDC42 3-UTR for binding miR-206, which in turn is involved in differentiating BM-MSCs to ECs. Finally, DMED rat modeling success was demonstrated by APO experiments. MALAT1 overexpression-modified BM-MSCs had significant therapeutic effects in DMED rats.

molecular biology↗
bioRxiv2024

Mechanistic dissection of the CHD4 enhancers reveals cooperative functions among the homotypic ZNF410 clustered motifs

Transcription factors often regulate numerous target genes. However, ZNF410 controls only a single gene, CHD4, in human erythroid cells by its highly restricted chromatin occupancy to the CHD4 locus via two clusters of ZNF410 binding motifs. Here, we uncover that ZNF410 controls chromatin accessibility and activity of the two CHD4 enhancer regions. Combining CRISPR/Cas9 genomic deletion with CRISPRi approaches, we demonstrate that both enhancer regions additively contribute to CHD4 gene expression. Mutations of the di-adenine nucleotides within the ZNF410 binding motif fully disrupt ZNF410-DNA interaction. Luciferase assays, ChIP-seq, and ATAC-seq studies reveal that the homotypic clustered motifs within the CHD4 enhancers are recognized by ZNF410 in a collaborative fashion. Especially, the three ZNF410 motifs located in the 3 end of the distal enhancer act as "switch motifs" to control the chromatin accessibility and activity of the whole distal enhancer region. Together, our findings expose a complex functional hierarchy of motifs, where clustered motifs bound by the same transcription factor cooperate to fine-tune the expression of target gene. HighlightsO_LIZNF410 maintains the accessibility of the CHD4 enhancer regions C_LIO_LIThe two enhancer elements additively modulate CHD4 gene expression C_LIO_LIThe dinucleotide "AA" is the key bases within ZNF410 binding motif C_LIO_LIZNF410 binds to homotypic clustered motifs in a cooperative manner C_LI

molecular biology↗
bioRxiv2024

Unfolded Von Willebrand Factor Binds Protein S and Reduces Anticoagulant Activity

Protein S (PS), the critical plasma cofactor for the anticoagulants tissue factor (TF) pathway inhibitor (TFPI) and activated protein C (APC), circulates in two functionally distinct pools: free (anticoagulant) or bound to complement component 4b-binding protein (C4BP) (anti-inflammatory). Acquired free PS deficiency is detected in several viral infections, but its cause is unclear. Here, we identified a shear-dependent interaction between PS and von Willebrand Factor (VWF) by mass spectrometry. Consistently, plasma PS and VWF comigrated in both native and agarose gel electrophoresis. The PS/VWF interaction was blocked by TFPI but not APC, suggesting an interaction with the C-terminal sex hormone binding globulin (SHBG) region of PS. Microfluidic systems, mimicking arterial laminar flow or disrupted turbulent flow, demonstrated that PS stably binds VWF as VWF unfolds under turbulent flow. PS/VWF complexes also localized to platelet thrombi under laminar arterial flow. In thrombin generation-based assays, shearing plasma decreased PS activity, an effect not seen in the absence of VWF. Finally, free PS deficiency in COVID-19 patients, measured using an antibody that binds near the C4BP binding site in SHBG, correlated with changes in VWF, but not C4BP, and with thrombin generation. Our data suggest that PS binds to a shear-exposed site on VWF, thus sequestering free PS and decreasing its anticoagulant activity, which would account for the increased thrombin generation potential. As many viral infections present with free PS deficiency, elevated circulating VWF, and increased vascular shear, we propose that the PS/VWF interaction reported here is a likely contributor to virus-associated thrombotic risk. Key PointsO_LIVon Willebrand Factor (VWF) binds Protein S (PS) in a shear-dependent manner, reducing the free PS pool and its anticoagulant activity. C_LIO_LIThe PS/VWF complex forms under turbulent flow conditions, is stable in whole blood, and localizes to growing platelet thrombi. C_LI

biochemistry↗
bioRxiv2024

ViraLM: Empowering Virus Discovery through the Genome Foundation Model

MotivationViruses, with their ubiquitous presence and high diversity, play pivotal roles in ecological systems and have significant implications for public health. Accurately identifying these viruses in various ecosystems is essential for comprehending their variety and assessing their ecological influence. Metagenomic sequencing has become a major strategy to survey the viruses in various ecosystems. However, accurate and comprehensive virus detection in metagenomic data remains difficult. Limited reference sequences prevent alignment-based methods from identifying novel viruses. Machine learningbased tools are more promising in novel virus detection but often miss short viral contigs, which are abundant in typical metagenomic data. The inconsistency in virus search results produced by available tools further highlights the urgent need for a more robust tool for virus identification. ResultsIn this work, we develop a Viral Language Model, named ViraLM, to identify novel viral contigs in metagenomic data. By employing the latest genome foundation model as the backbone and training on a rigorously constructed dataset, the model is able to distinguish viruses from other organisms based on the learned genomic characteristics. We thoroughly tested ViraLM on multiple datasets and the experimental results show that ViraLM outperforms available tools in different scenarios. In particular, ViraLM improves the F1-score on short contigs by 22%. AvailabilityThe source code of ViraLM is available via: https://github.com/ChengPENG-wolf/ViraLM. Contactyannisun@cityu.edu.hk

bioinformatics↗
bioRxiv2024

Microplastics from disposable paper cups are enriched in the placenta and fetus, leading to metabolic and reproductive toxicity during pregnancy

The health implications of microplastics (MPs), especially those originating from hot drinks in disposable paper cups (DPCs), are increasingly alarming. We investigated the accumulation and metabolic and reproductive toxicological effects of MPs from DPCs filled with hot water in various tissues in a pregnant mouse model. Simulating human intake of 0.3, 3.3, and 33.3 cups daily, we found MPs exposure-induced dose-responsive harmful effects on murine fetal development and maternal physiology. MPs were detected in all 13 examined tissues, with the highest accumulation in the cecal contents, followed by significant depositions in the fetus, placenta, kidney, spleen, lung, and heart. A higher proportion of smaller MPs (90.35% < 10 m) was identified in brain tissues. Dose-responsive changes in functional microbiome and gene pathways were observed. Moderate MPs intake of 3.3 cups daily significantly altered cecal microbiome composition and metabolic functions. The transcriptomic functional variations in maternal blood, placenta, and mammary gland underscore the significant impacts of realistic MPs exposure on metabolic and immune health and posing neurodegenerative and miscarriage risks. The benchmark dose framework analysis using tissue-specific gene biomarkers revealed safe exposure limits at 2 to 4 cups/day during pregnancy. Our results indicate selective tissue accumulation and potential metabolic and reproductive toxicities of MPs at exposure levels presumed non-hazardous. Such risks remain unaddressed within current food safety regulations, impacting vulnerable groups such as pregnant women and fetuses. Research HighlightsO_LIMicroplastics released from disposable paper cups filled with hot water showed preferential accumulations in the murine fetus, placenta, kidney, spleen, lung, and heart, with significant adverse impacts on fetal development. C_LIO_LIMicroplastic exposure led to dose-responsive maternal microbiome changes associated with increased fatty acid biosynthesis and elevated expressions of genes related to viral infections, neurodegenerative diseases, oxidative stress, and miscarriage risk. C_LIO_LIA consumption level of 3.3 cups/day was sufficient to elicit systemic metabolic and reproductive toxicity, with a predicted safe exposure limit of 2 to 4 cups/day during pregnancy by benchmark dose framework analysis with molecular biomarkers. C_LI

molecular biology↗
bioRxiv2024

Neonatal sensitivity to vocal emotions: A milestone at 37 weeks of gestational age

Emotional responsiveness in neonates, particularly their ability to discern vocal emotions, plays an evolutionarily adaptive role in human communication and adaptive behaviors. The developmental trajectory of emotional sensitivity in neonates is crucial for understanding the foundations of early social-emotional functioning. However, the precise onset of this sensitivity and its relationship with gestational age (GA) remain subjects of investigation. In a study involving 120 healthy neonates categorized into six groups based on their GA (ranging from 35 and 40 weeks), we explored their emotional responses to vocal stimuli. These stimuli encompassed disyllables with happy and neutral prosodies, alongside acoustically matched nonvocal control sounds. The assessments occurred during natural sleep states using the odd-ball paradigm and event-related potentials. The results reveal a distinct developmental change at 37 weeks GA, marking the point at which neonates exhibit heightened perceptual acuity for emotional vocal expressions. This newfound ability is substantiated by the presence of the mismatch response, akin to an initial form of adult mismatch negativity, elicited in response to positive emotional vocal prosody. Notably, this perceptual shifts specificity becomes evident when no such discrimination is observed in acoustically matched control sounds. Neonates born before 37 weeks GA do not display this level of discrimination ability. This developmental change has important implications for our understanding of early social-emotional development, highlighting the role of gestational age in shaping early perceptual abilities. Moreover, while these findings introduce the potential for a valuable screening tool for conditions like autism, characterized by atypical social-emotional functions, it is important to note that the current data are not yet robust enough to fully support this application. This study makes a substantial contribution to the broader field of developmental neuroscience and holds promise for future research on early intervention in neurodevelopmental disorders. Significance statementThis study illuminates a key developmental change, pinpointing the emergence of heightened emotional perceptual acuity at 37 weeks of gestational age. Employing rigorous methods, we reveal that neonates at this stage exhibit remarkable discrimination abilities for emotional vocal prosody, a vital turning point in early social-emotional functioning. These findings emphasize the pivotal role of gestational age in shaping neonatal perception and provides a potential pathway for early screening of neurodevelopmental disorders, particularly autism. This insight holds profound implications for understanding the foundations of early social-emotional development in humans, offering a potential tool for early intervention in neurodevelopmental disorders, thereby enhancing child health and well-being.

neuroscience↗
bioRxiv2023

Multivalent DNA-encoded lectins on phage enable detecting compositional glycocalyx differences

Selective detection of disease-associated changes in the cellular glycocalyx is a foundation of modern targeted therapies. Detecting minor changes in the density and identity of glycans on the cell surface is a technological challenge exacerbated by lack of 1:1 correspondence between cellular DNA/RNA and glycan structures on cell surface. We demonstrate that multivalent displays of up to 300 lectins on DNA-barcoded M13 phage on a liquid lectin array (LiLA), detects subtle differences in composition and density of glycans on cells ex vivo and in immune cells or organs in animals. For example, constructs displaying 73 copies of diCBM40 lectin per 700x5 nm virion ({varphi}-CBM73) exhibit non-linear ON/OFF-like recognition of sialoglycans on the surface of normal and cancer cells. In contrast, a high-valency {varphi}-CBM290 display, or soluble diCBM40, exhibit canonical progressive scaling in binding with increased epitope density; these constructs cannot amplify the subtle differences detected by {varphi}-CBM73. Similarly, multivalent displays of diCBM40 and Siglec-7 detect differences in the glycocalyx between stem-like and non-stem populations in cancer cells that are not detected with soluble lectins. Multivalent display of lectins on M13 scaffold with protected DNA inside the phage offer non-destructive detection of minor differences in glycocalyx in cells in vitro and in vivo not feasible to currently available technologies.

biochemistry↗
bioRxiv2023

HDAC10 blockade upregulates SPARC expression thereby repressing melanoma cell growth and BRAF inhibitor resistance

Secreted Protein Acidic and Rich in Cysteine (SPARC), a highly conserved secreted glycoprotein, is crucial for various bioprocesses. Here we demonstrate that histone deacetylase 10 (HDAC10) is a key regulator of SPARC expression. HDAC10 depletion or inhibition upregulates, while overexpression of HDAC10 downregulates, SPARC expression. Mechanistically, HDAC10 coordinates with histone acetyltransferase p300 to modulate the acetylation state of histone H3 lysine 27 (H3K27ac) at SPARC regulatory elements and the recruitment of bromodomain-containing protein 4 (BRD4) to these regions, thereby tuning SPARC transcription. HDAC10 depletion and resultant SPARC upregulation repress melanoma cell growth, primarily by induction of autophagy via activation of AMPK signaling. Moreover, SPARC upregulation due to HDAC10 depletion partly accounts for the resensitivity of resistant cells to a BRAF inhibitor. Our work reveals the role of HDAC10 in gene regulation through epigenetic modification and suggests a potential therapeutic strategy for melanoma or other cancers by targeting HDAC10 and SPARC. HighlightsO_LIHDAC10 is the primary HDAC member that tightly controls SPARC expression. C_LIO_LIHDAC10 coordinates with p300 in modulating the H3K27ac state at SPARC regulatory elements and the recruitment of BRD4 to these regions. C_LIO_LIHDAC10 depletion and resultant SPARC upregulation inhibit melanoma cell growth by inducing autophagy via activation of AMPK signaling. C_LIO_LISPARC upregulation as a result of HDAC10 depletion resensitizes resistant cells to BRAF inhibitors. C_LI

cancer biology↗
bioRxiv2023

Generalized Reporter Score-based Enrichment Analysis for Diverse Omics Data

Enrichment analysis contextualizes biological features in pathways to facilitate a systematic understanding of high-dimensional data and is widely used in biomedical research. The emerging reporter score-based analysis (RSA) method shows more promising sensitivity, as it relies on p-values instead of raw values of features. However, RSA cannot be directly applied to multi-group experimental designs and is often misused due to the lack of a proper tool. Here, we propose the Generalized Reporter Score-based Analysis (GRSA) method for multi-group and longitudinal omics data. A comparison with other popular enrichment analysis methods demonstrated that GRSA had increased sensitivity across multiple benchmark datasets. We applied GRSA to microbiome, transcriptome, and metabolome data and discovered new biological insights in omics studies. Finally, we demonstrated the application of GRSA beyond functional enrichment using a taxonomy database. We implemented GRSA in an R package, ReporterScore, integrating with a powerful visualization module and updatable pathway databases (https://github.com/Asa12138/ReporterScore). We believe the ReporterScore package will be a valuable asset for broad biomedical research fields.

bioinformatics↗
bioRxiv2023

Progressive community, biogeochemical and evolutionary remodeling of the soil microbiome underpins long-term desert ecosystem restoration

Ecological restoration of degraded lands is essential to human sustainability. Yet, an in-depth community, functional, and evolutionary microbial perspective of long-term restoration of damaged ecosystems is lacking. Herein, we comprehensively assessed the impact of long-term (up to 17 years) restoration of Tengger Desert, China, by multi-omic profiling of 1,910 topsoil samples. The soil biophysiochemical properties, especially soil hydraulics, microbiome stability, and functional diversity, significantly improved during restoration. The soil microbiome transitioned from an extreme oligotrophic and autotrophic community to a diverse copiotrophic ecosystem. The soil microbiota, including fungi, could mediate the soil physicochemical changes through metabolites. Importantly, the systematic rewiring of nutrient cycles featured the multi-domain preference of an efficient carbon fixation strategy in the extreme desert environment. Finally, the microbiome was evolving via positive selections of genes of biogeochemical cycles, resistance, and motility. In summary, we present a comprehensive community, functional, biogeochemical, and evolutionary landscape of the soil microbiome during the long-term restoration of desert environments. We highlight the crucial microbial role in restoration from soil hydraulic and biogeochemical perspectives, offering promising field applications. HighlightsO_LIThe desert soil microbiome transformed from simple oligotrophic to a diverse, stable, and nutrient-rich ecosystem with expanded functional diversity. C_LIO_LIRestoration led to systematically rewired biogeochemical cycles, which are highly efficient in carbon fixation in the desert environment. C_LIO_LIThe microbiome was evolving via positive selections of genes involved in biogeochemical cycles and environmental adaptations. C_LIO_LIMicrobes and metabolites could facilitate desert restoration from hydraulic and biogeochemical aspects, offering promising field applications. C_LI

microbiology↗
bioRxiv2023

The 14-3-3/spastin pathway coordinates neurite regeneration and recovery of spinal cord injury by targeting microtubule dynamics

Axon regeneration is abortive in the central nervous system following injury. Orchestrating microtubule dynamics has emerged as a promising approach to improve axonal regeneration. The microtubule severing enzyme spastin is essential for axonal development and regeneration through remodeling of microtubule arrangement. To date, however, little is known regarding the mechanisms underlying spastin action in neural regeneration after spinal cord injury. Here, we use glutathione transferase pulldown and immunoprecipitation assays to demonstrate that 14-3-3 interacts with spastin, both in vivo and in vitro, via spastin Ser233 phosphorylation. Moreover, we show that 14-3-3 protects spastin from degradation by inhibiting the ubiquitination pathway and upregulates the spastin-dependent severing ability. Furthermore, the 14-3-3 agonist Fusicoccin (FC-A) promotes neurite outgrowth and regeneration in vitro which needs spastin activation. Western blot and immunofluorescence results revealed that 14-3-3 protein is upregulated in the neuronal compartment after spinal cord injury in vivo. In addition, administration of FC-A not only promotes locomotor recovery, but also nerve regeneration following spinal cord injury in both contusion and lateral hemisection models; however, application of spastin inhibitor spastazoline successfully reverses these phenomena. Taken together, these results indicate that 14-3-3 is a molecular switch that regulates spastin protein levels, and the small molecule 14-3-3 agonist FC-A effectively mediates the recovery of spinal cord injury in mice which requires spastin participation. HighlightsO_LIThe formation of the 14-3-3 and spastin protein complex requires phosphorylation of spastin at the S233 site. C_LIO_LI14-3-3 is involved in the phosphorylation-ubiquitination crosstalk of spastin, thus impacting the protein stability. C_LIO_LI14-3-3 agonists Fusicoccin-A can facilitate the repair of spinal cord injury in mice. C_LIO_LIActivation of spastin function is a prerequisite for nerve regeneration and recovery of spinal cord injury in mice. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/550600v3_ufig1.gif" ALT="Figure 1"> View larger version (66K): org.highwire.dtl.DTLVardef@ebabb6org.highwire.dtl.DTLVardef@5651caorg.highwire.dtl.DTLVardef@113df6borg.highwire.dtl.DTLVardef@1b0ea5a_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗
bioRxiv2023

Identification and Characterization of CtUGT3 as the Key Player of Astragalin Biosynthesis in Safflower

Safflower (Carthamus tinctorius L.) flowers are used as a traditional Chinese medicine for a long history. Flavonoids are the main bioactive components in safflower flowers, and most of them exist in the form of flavonoid glycosides. Only few glycosyltransferases have been identified in safflower. To reveal the uridine diphosphate glycosyltransferase (UGT) involved in flavonoid glycosides biosynthesis in safflower, a metabolomics and transcriptome analysis was performed by using the flowers under different light qualities treatments. Three differentially expressed UGT genes were screened, and their expressions were significantly related with concentrations of 9 flavonoid O-glycosides. Safflower corolla protoplasts were further used to confirm flavonoid O-glycosylation ability of UGT candidates. The astragalin (kaempferol 3-O-glucoside) content was only significantly increased when CtUGT3 was overexpressed in protoplasts. The biochemical properties and kinetic parameters of CtUGT3 were determined. CtUGT3 also showed flavonoid 3-OH and 7-OH glycosylation activities in vitro. Molecular modeling and site-directed mutagenesis revealed that E384 and S276 were critical catalytic residues for the 3-OH glycosylation of CtUGT3. These results demonstrate that CtUGT3 has a flavonoid 3-OH glycosylation function and is involved in the biosynthesis of astragalin in safflower. This study provides insights into the catalytic mechanisms of flavonoid O-glycosyltransferases, and makes a reference for flavonoid biosynthesis genes research in medicinal plants.

plant biology↗