Search Results (438)

Bioactive peptides are an important source of therapeutic molecules and molecular scaffolds involved in defense, signaling, and immune regulation. Despite the extraordinary diversity of Coleoptera, the structural landscape of beetle-derived bioactive peptides remains largely unexplored, limiting our understanding of their evolutionary diversity and biotechnological potential. Here, we performed a large-scale structural survey of predicted toxin-like peptide scaffolds across publicly available Coleoptera transcriptomes by integrating transcriptome mining, peptide maturation prediction, physicochemical characterization, AlphaFold 3 structural modeling, structural similarity analyses, and interpretable machine learning. We identified 291 candidate peptides, of which 155 contained canonical signal peptides and 273 produced mature peptides within the expected size range of known bioactive peptides. Structural analyses revealed that, despite extensive sequence diversity, many candidates were organized into a comparatively restricted repertoire of compact cysteine-rich architectures, indicating that structural similarity is retained across peptides exhibiting substantial primary-sequence variation. Comparative structural analyses further identified recurrent protein architectures shared across multiple beetle lineages, while machine learning prioritization integrated structural and biochemical descriptors to identify high-confidence candidates for future functional characterization. These analyses establish the first structural atlas of predicted toxin-like peptides across Coleoptera and demonstrate that structure-guided transcriptome mining provides a powerful framework for uncovering recurrent bioactive peptide scaffolds that would remain largely undetected using sequence-based approaches alone. Beyond expanding our understanding of peptide evolution in beetles, this resource is a foundation for future structural, functional, and biotechnological exploration of bioactive peptides in underexplored animal groups. Full article

Verticillium wilt, caused by Verticillium dahliae, is a major constraint on cotton production. The GASA (Gibberellic Acid-Stimulated in Arabidopsis) proteins are cysteine-rich peptides induced by gibberellin signaling; however, their systemic role in upland cotton (Gossypium hirsutum) resistance to Verticillium wilt remains elusive. Here, we identified 40 GhGASA members from the G. hirsutum genome, which were clustered into three subfamilies. All members possessed a typical gibberellin-regulated domain, and segmental duplication was the primary driver of family expansion. Transcriptomic analyses revealed tissue-specific expression, with high abundance in pistils and roots, and distinct responsive patterns to salt, drought, and cold stresses. Quantitative real-time PCR (qRT-PCR) showed that multiple GhGASA genes were significantly upregulated following V. dahliae inoculation, with transcript levels being much higher in roots and stems than in leaves. Notably, GhGASA17, harboring ethylene-responsive elements, was identified as a key candidate involved in the defense response. Overall, this study provides a comprehensive characterization of the GhGASA gene family, offering a theoretical foundation for understanding its regulatory functions in Verticillium wilt resistance and delivering potential genetic resources for molecular breeding in cotton. Full article

Properly folded proteins are essential for studying protein function and developing therapeutic applications. However, when recombinant proteins are overexpressed in bacterial systems such as Escherichia coli, they often accumulate as inclusion bodies, insoluble protein aggregates in non-native conformations. Here, we report a refolding protocol for mouse Frizzled8 cysteine-rich domain (mouse FZD8 CRD) recovered from inclusion bodies, yielding a soluble, folded CRD preparation suitable for NMR analysis. Because mouse FZD8 CRD contains multiple conserved cysteine residues that must form an appropriate intramolecular disulfide-bonding pattern, the refolding strategy combines denaturant-mediated solubilization, dilution into a redox-buffered refolding solution, concentration, dialysis, lyophilization, and HPLC purification. Lyophilization followed by HPLC purification enables the resolution of the desired CRD species from aggregated and misfolded products. In this workflow, HPLC served as a key purification step, providing the resolution needed to enrich a folded, multi-disulfide-bond-containing CRD species from closely related misfolded species. NMR spectroscopy supported the presence of a folded, conformationally homogeneous CRD preparation. This workflow was also applied to mouse secreted Frizzled-related protein 3 cysteine-rich domain (mouse sFRP3 CRD), suggesting that it may be useful for other Frizzled-type CRDs with conserved cysteine-rich architectures, although additional validation will be required before broader application to more distantly related cysteine-rich proteins. Full article

Hydrophobins are small, cysteine-rich amphipathic proteins predominantly produced by filamentous fungi, known for their ability to self-assemble at hydrophobic–hydrophilic interfaces. These proteins are essential for fungal development, surface interactions, pathogenicity, and environmental adaptation, and they have attracted growing interest for biotechnological applications. In this work, we provide a narrative review of fungal hydrophobins, based on a systematic literature search and manual curation of eligible studies integrated with protein database records from Ascomycota and Basidiomycota. Information was compiled from peer-reviewed publications selected according to predefined eligibility criteria and complemented with UniProt records, covering taxonomic distribution, functional and biophysical properties, and physiological and pathogenic roles. Significant diversity in molecular features and physicochemical profiles was observed, indicating functional specialization across different ecological niches and lifestyles. Additionally, the compiled data highlight various biotechnological applications, such as surface modification, enzyme immobilization, drug delivery systems, biomaterial development, and environmentally sustainable technologies. By consolidating molecular, functional, and applied information across a wide range of fungal species, this review provides a comprehensive reference framework for hydrophobin research and biotechnological innovation. Full article

The Wnt gene family encodes highly conserved signaling molecules involved in vertebrate development, tissue homeostasis, and cell fate regulation. However, the evolutionary characteristics and potential biological functions of Wnt genes in the domestic donkey (Equus asinus) remain poorly understood. In this study, we performed the first genome-wide identification and comprehensive characterization of the donkey Wnt gene family. A total of 19 Wnt genes were identified, consistent with the conserved Wnt repertoire reported in other mammalian species. Phylogenetic and synteny analyses revealed strong evolutionary conservation of Wnt genes among mammals, particularly between donkey and horse, indicating conserved orthologous relationships within the genus Equus. Structural analyses showed that all donkey Wnt proteins contained the conserved cysteine-rich domain (CRD), whereas sequence variation was mainly concentrated in the N- and C-terminal regions. Transcriptomic analysis across 13 adult tissues revealed tissue-biased expression patterns, with Wnt3, Wnt5a, and Wnt4 showing relatively higher expression in the testis, epididymis, and skin, respectively, suggesting their potential involvement in reproductive regulation and tissue homeostasis. PPI network and GO enrichment analyses indicated potential associations between donkey Wnt proteins and conserved regulators involved in developmental processes, cell fate regulation, and Wnt signaling pathways. Integration of expression profiles with predicted interaction networks further suggested tissue-associated regulatory patterns among Wnt pathway components. Overall, this study provides a comprehensive resource for understanding the genomic organization, evolutionary conservation, and expression diversity of the donkey Wnt gene family, and offers insights into potential Wnt-mediated regulatory mechanisms in donkey biology. Full article

The growing demand for sustainable protein sources has intensified interest in plant-based food systems. Pulses are well established and recognized as nutritious, sustainable protein sources (~15–30% dry weight), rich in lysine and limited in cysteine, methionine, and tryptophan amino acids. In contrast, Spirulina is characterized by a complete amino acid profile, high protein content (~50–70% dry weight), and bioactive potential. This comprehensive review explores the role of Spirulina as a complementary protein source in plant-based systems, with emphasis on its structural, technofunctional, and processing characteristics. A literature analysis was conducted to identify advances, trends, and gaps in their incorporation into food matrices. The findings indicate that, despite their nutritional and functional advantages, the application of microalgae and beans remains constrained by sensory and technological challenges, such as off-flavors and textural issues. The combination of microalgae with pulse-based ingredients represents a promising strategy to enhance amino acid balance, improve functionality, and develop nutritionally enriched products. Studies addressing multi-component systems remain scarce, especially in bakery and flour-based applications, as well as in understanding synergistic interactions between microalgae and pulse proteins in food matrices. This review highlights the potential of microalgae as key ingredients in next-generation food formulations. It identifies critical gaps that must be addressed to enable their broader application in food products. Full article

Aortic dissection (AD) is a severe vascular condition marked by abrupt onset, rapid progression, and heightened mortality rates. Vascular smooth muscle cells (VSMCs), the predominant cellular component of the arterial media, are essential for maintaining the structural integrity and functionality of blood vessels. Recent studies have associated Cysteine-rich protein 2 (CSRP2) with the advancement of several vascular diseases. The involvement of CSRP2 in AD progression is unclear. Aortic tissues were collected from patients for RNA sequencing and histological analysis. A mouse model of AD was created using β-aminopropionitrile monofumarate (BAPN), while VSMC phenotypic switching was induced by platelet-derived growth factor BB (PDGF-BB). Adeno-associated virus vector was used to overexpress CSRP2 in aorta. A variety of histopathological assays and biochemical analyses were applied to determine gene and protein expression patterns as well as uncover underlying molecular mechanisms. CSRP2 was significantly downregulated in both human and murine AD, and CSRP2 gene overexpression dramatically reduced BAPN-induced AD incidence and prevented animal mortality. CSRP2 could preserve a contractile VSMC phenotype, even though under PDGF-BB stimulation. Mechanistically, our findings reveal that CSRP2 directly interacts with p130 Crk-associated substrate (p130Cas; also known as BCAR1) and reduces its phosphorylation, which in turn inhibits the activation of extracellular signal-regulated kinase (ERK) signaling pathways, thereby preventing VSMC phenotypic switching induced by PDGF-BB. Our findings identify CSRP2 as a novel regulator of VSMC phenotypic modulation and a significant modulator of AD development, suggesting its potential as a target for early intervention for AD. Full article

Cotton is an economically important cash crop severely affected by Verticillium wilt caused by Verticillium dahliae. Secreted cell-wall-degrading enzymes act as key virulence factors of this pathogen, yet the biological function of glucan 1,3-β-glucosidase remains largely uncharacterized. The gene VDAG_02814 (designated Vdbgl1) was previously found to be strongly induced during host infection. Here, combined approaches including gene knockout, host-induced gene silencing (HIGS), and transcriptomic analysis were utilized to characterize the function of Vdbgl1. The results showed that Vdbgl1 deletion retarded colony growth on PDA medium by 20.1–21.6%, decreased conidial yield by 20.2–46.7%, lowered spore germination rate by 30.7–32.9%, and weakened utilization of diverse carbon sources by 5.0–14.9%. The ΔVdbgl1 mutants also exhibited significantly increased sensitivity to cell wall-perturbing, osmotic, and membrane-damaging agents. Additionally, the ΔVdbgl1 mutants exhibited reduced disease index by approximately 19.1–27.7% and decreased fungal biomass by 42.7–61.8% in cotton; consistently, HIGS-mediated silencing of Vdbgl1 reduced the disease index by 25.7% and 26.7% and decreased fungal biomass by 48.1–71.5%. Transcriptome profiling of cotton roots infected by the ΔVdbgl1 mutant and the wild-type strain revealed 1007 down-regulated genes enriched in carbohydrate metabolism, cell wall degradation, and energy pathways, including 27 carbohydrate-active enzyme genes and 58 genes encoding cysteine-rich secreted proteins. Collectively, these findings indicated that Vdbgl1 coordinates carbon utilization, cell wall remodeling, and stress responses to regulate fungal development and pathogenicity, representing a promising target for cotton Verticillium wilt control. Full article

Hulless barley (HB) is a nutrient-rich cereal gaining renewed research interest in the Tibetan regions of China. Grain protein content (GPC) is a quality-determining trait in HB that significantly affects end-use quality and is strongly influenced by environmental factors. Identifying stable association regions and developing breeding-applicable molecular markers are breeding objectives for high-GPC HB variety improvement. This study analyzed 266 HB accessions grown across six environments (2018–2021). Phenotypic GPC ranged from 5.02% to 18.31%, with moderate heritability (H² = 0.473) and significant G×E interaction. A genome-wide association study (GWAS) using 168,983 GBS-derived SNPs and three models (GLM, MLM, FarmCPU) revealed no Bonferroni-significant SNPs in the BLUP meta-analysis. At the suggestive threshold (p < 1 × 10⁻⁵), 40 SNPs (18 loci) showed nominal association, with chromosome 7H harboring the most robust cross-model signal. Two SNPs on chromosome 6H and 7H exceeded Bonferroni correction in Yangma-2018 under GLM and FarmCPU. Candidate gene mining revealed seven functional genes, including 30S ribosomal protein S13 and glutamate-cysteine ligase. The lead SNP S7H_6966377 was located 165 kb from 30S ribosomal protein S13. These findings establish chromosome 7H as a regulatory hub for GPC and provide markers for marker-assisted selection. Full article

The Cysteine-rich Polycomb-like Protein (CPP) gene family is a class of transcription factors containing conserved CXC domains that are widely involved in the regulation of plant growth and development, cell division, and stress responses. Based on the ginseng genome and transcriptome database, all members of the PgCPP gene family in Panax ginseng were systematically identified, and comprehensive bioinformatics analyses, including phylogenetic, conserved domain, chromosomal localization and collinearity, cis-acting element, and expression pattern analyses, were conducted. In this study, we identified 44 PgCPP gene family members in ginseng, which were unevenly localized on multiple chromosomes. The phylogenetic tree divided them into three subfamilies, with members in the same subfamily being highly conserved. Conserved domain analysis revealed that all PgCPPs contain typical CXC motifs. Cis-acting elements were abundant in light response, hormone responses (abscisic acid, methyl jasmonate, salicylic acid), and stress response elements. Expression heatmaps demonstrated that different members have specific expression patterns across different ages, tissues, and species. After treatment with MeJA, transcriptional suppression of PgCPP03-4 and PgCPP03-13 was observed, and their expression levels demonstrated significant negative correlations with the contents of six protopanaxadiol-type ginsenosides. These findings suggest that PgCPP03-4 and PgCPP03-13 may act as negative regulators of protopanaxadiol-type ginsenoside biosynthesis within the MeJA signaling pathway. This systematic characterization and identification of the CPP gene family members in P. ginseng establishes a foundational framework for future functional validation and molecular breeding initiatives. Full article

Alternaria brassicicola is a necrotrophic fungal pathogen causing black spot disease on cruciferous crops worldwide. In this study, we comprehensively characterized the pathogenic isolate Ab0920a from diseased broccoli in Shanghai using morphological, phylogenetic, host range, fungicide sensitivity, genomic, and functional analyses. Pathogenicity tests on 27 cruciferous varieties revealed a broad host range with varying resistance levels. Fungicide sensitivity assays showed that fluxapyroxad (EC₅₀ = 0.0695 µg/mL), prochloraz (0.0711 µg/mL), and difenoconazole (0.0863 µg/mL) were highly effective, whereas fluazinam was least effective. Genome-wide annotation identified 941 secreted proteins (8.95% of the proteome) and 237 candidate effectors, including 31 small cysteine-rich secreted proteins and 68 homologs of known virulence factors. Conserved effector-associated motifs (e.g., RXLR, [Y/F/W]xC) were detected, and carbohydrate-active enzyme CAZyme annotation revealed diverse families potentially involved in plant cell wall degradation. Functional validation using the pSUC2 yeast system confirmed that N-terminal signal peptides of tested effectors are competent for secretion. A PVX-based transient expression assay in Nicotiana benthamiana identified two effectors that suppress Bax-induced programmed cell death, suggesting their potential roles in modulating host immunity. Overall, this study provides comprehensive insights into the pathogen Ab0920a, offering resources for disease management and functional studies on necrotrophic fungal pathogenesis. Full article

Basonuclin 2 (BNC2) is a highly conserved cysteine–histidine (C2H2)-type zinc-finger nuclear regulatory protein characterized by three pairs of zinc-finger domains, a putative nuclear localization signal, a serine-rich region, broad tissue distribution, and remarkable transcript diversity generated through alternative promoter usage, alternative splicing, and polyadenylation. Increasing evidence from human genetics, animal models, functional genomics, and transcriptomic studies indicates that BNC2 links nuclear regulatory mechanisms to tissue-specific developmental and disease phenotypes. In the nervous system, BNC2-positive neuronal populations and BNC2-derived circular RNAs have been implicated in energy-balance circuits and neuroinflammatory regulation. In the skeletal system, BNC2 contributes to osteochondral development, periosteal stem-cell activation, chromatin remodeling, fracture repair, and genetic susceptibility to adolescent idiopathic scoliosis. BNC2 variants have also been associated with congenital lower urinary tract obstruction, whereas its expression and regulatory landscape are closely related to germ-cell development, epithelial ovarian cancer susceptibility, pigmentation traits, fibrosis, and several tumor contexts. Mechanistically, BNC2-associated phenotypes appear to involve cysteine–histidine zinc-finger-mediated transcriptional regulation, non-coding enhancer activity, epigenetic alterations, RNA-processing-associated nuclear functions, and chromatin-remodeling-dependent control of cell proliferation, differentiation, and stromal activation. This review integrates current evidence on the molecular architecture and regulatory functions of BNC2, critically discusses its context-dependent roles across development and disease, and highlights unresolved questions regarding isoform-specific activity, cell-type-specific regulation, downstream target networks, and clinical translation. A clearer understanding of these mechanisms may support the future evaluation of BNC2 as a biomarker, genetic susceptibility locus, molecular stratification factor, and potential therapeutic regulatory node. Full article

Expansin-like A (EXLA) proteins belong to one of the four main families within the expansin superfamily, a group of plant proteins essential for cell wall loosening. Here, we report, for the first time, the purification of a novel EXLA, named cpEXLA, from canihua seeds. cpEXLA (yield ~0.16 mg per 100 g of seeds) is a 29 kDa glycoprotein with a high melting temperature (Tm of 86.75 ± 1.06 °C). Elucidation of its primary structure reveals that the mature protein consists of 246 amino acids, ten of which are cysteine residues forming five disulphide bridges. Structural studies based on 3D model prediction reveal the presence of N- and C-terminal domains, which are typical of EXLAs and rich in β-sheets, as confirmed by circular dichroism (CD) spectroscopy. Furthermore, comparative analysis of amino acid sequences between cpEXLA and 219 similar EXLAs, retrieved from dicotyledonous genomes and transcriptomes, identified eighteen invariant amino acid residues: eleven in the N-terminal domain and seven in the C-terminal domain. Finally, phylogenetic analysis of EXLAs in dicotyledonous species shows a close relationship with other EXLAs from the Amaranthaceae family, confirming that EXLA proteins are highly conserved among dicotyledonous plants. Overall, cpEXLA represents an intriguing native tool for studying cell wall evolution and the functional role of EXLAs. Full article

Kelch-like ECH-associated protein 1 (Keap1) acts as a repressor of nuclear factor-erythroid 2-related factor 2 (Nrf2), a major transcription factor regulating cellular antioxidant response. Keap1 is the substrate adaptor subunit of the cullin 3-RING E3 ubiquitin ligase complex that specifically facilitates Nrf2 ubiquitination and its proteasomal degradation. Keap1 is rich in cysteine residues, and several of them undergo various modifications, such as sulphydration, nitrosylation and glutathionylation under cellular stress conditions. Some of these modifications alter the conformation of Keap1, preventing Nrf2 from ubiquitination and subsequent proteasome-mediated degradation. As a result, newly synthesised Nrf2 translocates to the nucleus to induce the expression of diverse genes involved in protecting cells against oxidative stress. Protein CoAlation is a reversible redox-dependent post-translational modification (PTM) in which coenzyme A (CoA) forms disulphide bonds with oxidised cysteine residues under oxidative or metabolic stress. In this study, we demonstrate for the first time that disulphide Keap1 dimer undergoes CoAlation in cellular response to oxidative stress induced by various oxidising compounds. Furthermore, glucose deprivation also induces CoAlation of the disulphide Keap1 dimer in HEK293/Pank1β cells. We also demonstrate that the Keap1^{11 Cys-less} mutant is not CoAlated in response to diamide treatment or glucose deprivation. In summary, this study uncovers a novel PTM of Keap1 by the key metabolic integrator CoA, which provides new insights into the regulation of the Keap1-Nrf2 antioxidant pathway under oxidative and metabolic stress. Full article

Animal venoms induce systemic inflammatory response syndrome through their interaction, inter alia, with pattern recognition receptors (PRRs) of the innate immune system. CD5 and CD6 are lymphoid members of the scavenger receptor cysteine-rich superfamily, endowed with PRR activity against microbial-associated molecular patterns (MAMPs) derived from bacteria, fungi, viruses and/or parasites. In this study, we aimed to investigate CD5 and CD6 interaction with cobra (Naja haje), scorpion (Androctonus australis) and honeybee (Apis mellifera) venoms. Binding assays revealed direct, dose-dependent and specific interaction of soluble human CD5 and CD6 receptors with protein nature components from the three venoms. Proteomic analysis identified venom nerve growth factor, basic phospholipase A2 (PLA2) and cobra venom factor, in cobra venom, and scorpion venom toxins targeting potassium (α-KTx 8.1) and sodium channels (Neurotoxin-1″ and G-TI) as potentially interacting components with CD5 and CD6. Further studies confirmed direct binding of bee venom main components, phospholipase A2 and melittin, to both soluble CD5 and CD6 receptors. Interestingly, in vitro PLA2 activity from cobra and bee venom was significantly reduced by both soluble CD5 and CD6 receptors. These findings broaden the PRR properties of CD5 and CD6 and support their potential involvement in envenomation pathophysiology. Full article