Protein structure bridges the sequence-function relationship, enabling deep exploration of biological processes across diverse organisms. Insects, the most diverse animal lineage, accounting for over 50% of all described animal species, provide an exceptional system for exploring sequence-structure-function relationships. Here, we reconstructed a comprehensive and well-resolved phylogeny of 4854 insects, spanning all orders. Leveraging this framework, we created an atlas of 13.29 million predicted protein structures from 824 representative species, including 11.63 million newly predicted structures. Structural clustering revealed that proteins with divergent sequences but similar structures could be effectively grouped together. Structural similarity searches against proteins with well-characterized functions yielded annotations for 7.61 million insect proteins, including up to 14% of previously unannotated proteins. We further identified 750 million remote homologs between insect proteins, many of which trace back to ancient branches of the insect phylogeny. Remarkably, despite extensive sequence divergence, cGAS-like receptors (cGLRs) were structurally conserved across all 824 insects. Experimental assays demonstrated that these structurally identified cGLRs play a crucial role in antiviral defense in the yellow fever mosquito. Our findings highlight the significance of structural genomics for understanding protein function and evolution across the tree of life.

Eukaryotic messenger RNA (mRNA) homeostasis requires precise coordination between synthesis and decay, yet the mechanisms governing this balance in fungal pathogens remain elusive. Here we provide a comprehensive characterization of the Lsm1-7 complex in the cereal pathogen Fusarium graminearum. We show that Lsm1-7 assembles into a conserved hetero-heptameric module that localizes to processing bodies (P-bodies) and is required for fungal growth, virulence, and mycotoxin biosynthesis. Mechanistically, Lsm1-7/Pat1 binds U/A-rich 3' termini of a defined set of transcripts enriched for central metabolism and restrains their 3'-5' decay. Genetic suppressor analyses and mechanistic dissection identify two parallel decay routes antagonized by Lsm1-7, including the exosome recruited by the uridyltransferase Cid1 and the Ski-exosome complex mediated by the newly identified scaffold protein Lsp1. Moreover, loss of Lsm1-7 elicits a compensatory transcriptional response involving the Rpd3L (Sin3) histone deacetylase complex, in which elevated histone H4 acetylation at affected loci partially restores transcript output. Together, our results define an integrated cytoplasmic-nuclear regulatory axis in F. graminearum that couples 3'-end protection to chromatin-based transcriptional buffering to maintain mRNA homeostasis. While Lsp1 appears lineage-adapted, the underlying logic may reflect a broader principle of gene-expression buffering that supports fungal fitness and pathogenicity.

Fusarium graminearum, the causal agent of Fusarium head blight (FHB), poses a major threat to global food security by contaminating cereals with the mycotoxin deoxynivalenol (DON). Although transcriptional and protein-level regulation of its stress response and virulence has been extensively studied, the functional significance of mRNA processing in these critical processes remains largely unexplored. Here, we identify Lsm8, a highly conserved core subunit of the nuclear Lsm2-8 complex, as a pivotal regulator linking RNA splicing fidelity to fungal growth, stress adaptation, and virulence. Deletion of LSM8 disrupted Lsm2-8 assembly and nuclear localization, resulting in widespread intron retention in genes essential for stress signaling (HOG1, ATF1), development (GPA1, STE12), and trichothecene biosynthesis. Consequently, osmoadaptation was impaired, sexual reproduction was abolished, and both DON production and virulence were drastically reduced. We further demonstrate that intron-retained transcripts are predominantly degraded by the RNA exosome, revealing a conserved Lsm8-exosome module that maintains splicing fidelity and RNA surveillance. Given the deep evolutionary conservation of Lsm8 across eukaryotes, these findings uncover a fundamental post-transcriptional regulatory layer governing fungal stress response, virulence, and mycotoxin biosynthesis, and highlight RNA-processing factors as universal determinants of virulence and promising antifungal targets across eukaryotic pathogens.

In both plant and animal innate immunity, rapid biosynthesis of cytokine-like peptides is essential for activating immune responses through plasma membrane receptor kinases. Peptide homeostasis is tightly controlled at multiple regulatory levels to avoid adverse effects caused by excessive phytocytokine production. However, the mechanisms underlying the rapid transition between transcriptional activation and repression of phytocytokine genes in response to immune stimuli remain largely unknown. Here we identify previously uncharacterized phytocytokine clusters in wheat, termed TaFIPs, which are rapidly induced by Fusarium infection. TaFIP genes are epigenetically regulated by H3K4me3 and H3K27me3 bivalent histone marks enabling rapid transcriptional shifts. Functionally, the leucine-rich repeat receptor kinase TaFIPR directly recognizes and binds TaFIPs to activate multiple immune responses, thereby enhancing wheat resistance to Fusarium infection. Our findings reveal an epigenetic mechanism that fine-tunes phytocytokine-mediated immunity in wheat.

Cellulose, a primary component of plant cell walls, is synthesized by cellulose synthase complexes (CSCs) at the plasma membrane. Targeting this process with cellulose biosynthesis inhibitors (CBIs) has significantly advanced our understanding of plant cell wall formation and provided valuable compounds for herbicide development. Here, we identified a fungal natural product, 8-methyldichlorodiaporthin (MDD), as a broad-spectrum plant CBI. Structure-activity relationship analyses demonstrate that methylation modifications on the isocoumarin ring and chlorination of the side chain are crucial for MDD-induced growth inhibition. A chemical forward genetic screen in Arabidopsis thaliana revealed two semidominant CESA1 mutations, causing A903T and H1024Y substitutions, that confer insensitivity to MDD. Both mutations locate to transmembrane domains of CESA1, and we show that MDD depletes CSCs from the plasma membrane and reduces cellulose content. Further genetic analyses indicate that the cesa1mddi1-1 A903T mutant also confers resistance to CBIs quinoxyphen and C17, but not to CBIs isoxaben, indaziflam, or ES20. Stacking additional point mutations conferring resistance to other CBIs, cesa3ixr1-1 G998D, and cesa6es20-r3 G935E into the cesa1mddi1-1 A903T background yields multiple-drug-resistant lines that maintain normal growth. These findings establish MDD, as a natural CBI that likely targets CESA1, thereby extending our understanding of CSC regulation and abilities to develop multidrug-resistant crop varieties. These findings offer unique perspectives for weed management and plant biotechnology.

Filamentous fungi harbor a vast potential for secondary metabolite (SM) biosynthesis, yet the biological functions of numerous biosynthetic gene clusters (BGCs) remain obscure. In Fusarium graminearum, a devastating cereal pathogen, SMs are best known as virulence factors, but their broader contributions to fungal physiology are poorly defined. Here, we present a genome-scale functional dissection of 53 predicted SM-BGCs by constructing a knockout library targeting cluster backbone genes and systematically quantifying 24 phenotypic traits, generating 1,272 phenotypic measurements. This dataset reveals that secondary metabolism is not a dispensable metabolic burden; instead, SM-BGCs are broadly integrated into vegetative growth, asexual development, and abiotic stress adaptation. Transcriptome analyses further uncover pronounced spatiotemporal regulation and tissue-dependent requirements of SM-BGCs during infection of wheat heads versus coleoptiles, thereby revealing an ecological dimension of pathogenesis. Mechanistic investigation identified two previously uncharacterized clusters, PKS-type BGC36 and NRPS-type BGC47, as critical for full virulence. BGC36 positively regulates deoxynivalenol (DON) biosynthesis, whereas disruption of BGC47 compromises cell wall/membrane stress tolerance and is associated with reduced phosphorylation of the kinase Mgv1, impaired DON-toxisome formation, and reduced DON production. Together, our findings establish fungal secondary metabolism as a core physiological buffer against environmental fluctuations that supports homeostasis and virulence, and they provide a comprehensive genetic resource for dissecting the chemical biology of Fusarium.

Understanding how fungi regulate mycotoxin production is critical for managing crop diseases and reducing contamination in food systems. Here, we elucidate the mechanism of the transcription factor FgSge1 in Fusarium graminearum, a significant fungal pathogen responsible for Fusarium head blight in cereal crops, in regulating mycotoxin biosynthesis and pathogenicity. FgSge1 specifically binds to the 8-bp cis-element TAARGTTT. Under mycotoxin-induced conditions, FgSge1 binds to this cis-element within its own promoter, activating its own transcription. Additionally, FgSge1 connects with this cis-element within the promoters of mycotoxin biosynthesis genes and interacts directly with the scaffold protein FgAda2 of the Spt-Ada-Gcn5-Acetyltransferase (SAGA) complex. This interaction recruits the histone acetyltransferase FgGcn5 to the promoters of DON biosynthetic genes, promoting histone acetylation and facilitating jet-like chromatin architecture, thereby activating transcription. In contrast, the FgSge1 mutant fails to recruit the SAGA complex, leading to reduced histone acetylation, disrupted chromatin structure, and impaired DON biosynthesis. Together, these findings establish FgSge1 as a critical self-activating regulatory factor that links histone acetylation to higher-order chromatin remodeling, orchestrating mycotoxin gene activation, and providing a framework for understanding epigenetic control of mycotoxin biosynthesis and virulence in fungi.

Thermo-sensitive genic male sterility (TGMS) lines play a pivotal role in two-line hybrid rice breeding. However, the availability of elite TGMS germplasm remains limited, and the molecular mechanisms underlying fertility transition of TGMS lines are still poorly understood. Here, we identified a novel TGMS line, Osgpat6.1, which exhibits temperature-dependent fertility transition, with male sterility under high-temperature (HT) and restored fertility under low-temperature (LT). Cytological observations revealed that the TGMS line exhibits disrupted programmed cell death (PCD) of the tapetum under HT conditions. This disruption leads to delayed tapetum degradation and defective pollen exine formation. Map-based cloning identified a premature termination mutation in glycerol-3-phosphate acyltransferase 6.1 (OsGPAT6.1), which is highly expressed in anthers and essential for lipid biosynthesis. Lipidomic profiling revealed that disruption of OsGPAT6.1 perturbed lipid metabolism during pollen development. Genetic studies further revealed that male sterility in Osgpat6.1 mutant was restored through functional redundancy with its homologues OsGPAT6.2 and OsGPAT6.3 under LT. Overexpression of OsGPAT6.2 or OsGPAT6.3 partially restored the fertility of Osgpat6.1 mutant under HT. However, simultaneous disruption of all three homologues abolished fertility restoration even under LT. Notably, introgression of the Osgpat6.1 allele into diverse rice cultivars produced stable TGMS lines without compromising hybrid vigor. Collectively, these findings demonstrate that dysfunction of GPAT confers TGMS in rice by disrupting glycerolipid metabolism and provides TGMS germplasm to facilitate two-line hybrid rice breeding.

Bacterial leaf blight (BLB), caused by Xanthomonas oryzae pv. oryzae (Xoo), remains a major threat to global rice production. In this study, we identified the novel small-molecule compound PK150 with potent antibacterial activity against Xoo. PK150 exhibits a minimum inhibitory concentration (MIC) of 0.15 μg/mL in vitro, and achieved 78% protective efficacy at 200 μg/mL in planta. Mechanistic studies revealed that PK150 targets menaquinone biosynthesis by binding to demethylmenaquinone methyltransferase (MenG), which is a key enzyme in this pathway. Surface plasmon resonance (SPR) analysis confirmed that PK150 binds to MenG, with a dissociation constant (Kd) of 6.42 × 10-5 M. The antibacterial effect of PK150 was markedly reduced by the addition of 100 μg/mL exogenous menaquinone-4 (MK-4) and overexpression of MenG. Moreover, molecular docking analysis identified ALA-73, THR-76, ASP-97, and ILE-98 as key residues involved in the PK150-MenG interaction, which was further validated by SPR assays showing loss of binding to a MenG mutant. Phylogenetic analysis revealed that the sequence of MenG in Xoo showed significant evolutionary differences from those of other Gram-negative bacteria. These findings highlight PK150 as a promising candidate for the development of novel agrochemicals to manage BLB.

Lipid composition represents a significant differentiator across the three domains (eukaryotes, bacteria, and archaea) of cellular life. Eukaryotes possess distinct lipids, such as sterols and sphingolipids, generally, these are not commonly found in typical bacteria and archaea. Sterols play a pivotal role in eukaryotic cellular functions, lanosterol, a key precursor for animal and fungal steroids, has well established functions in eukaryotes, while its potential functions in bacteria remain largely uninvestigated. In this study, we genetically engineered Escherichia coli (E. coli) to reconstruct the biosynthesis of lanosterol, and successfully developed a novel E. coli strain capable of synthesizing lanosterol, although its specific location, such as whether it is incorporated into the cell membrane, remains to be further determined. Comprehensive characterization of the observed phenotypic changes has unveiled that, despite an unaltered growth rate under normal condition, the engineered E. coli strain displayed notably enhanced tolerance to various stresses. Subsequent analysis has indicated that lanosterol plays a role in preserving membrane integrity, fluidity, hydrophobicity, and ATP production, mirroring the functions of sterols in eukaryotes. This study unveils the unexpected capacity of E. coli to synthesize sterols, not only underscores the importance of lanosterol as a precursor for essential cellular lipids but also offers fresh insights into the potential functions of sterols within bacterial systems.

Fusarium crown rot (FCR) poses a major threat to wheat production in the Huanghuai wheat region of China. This study aims to enhance understanding of pathogen populations causing FCR, focusing on their pathogenicity, trichothecene genotypes, and fungicide resistance. During the 2022-2023 growing seasons, we collected 1820 fungal isolates from 233 locations in this region. Our results identified Fusarium pseudograminearum, Fusarium graminearum, and Fusarium asiaticum as the primary pathogens, with F. pseudograminearum exhibiting the highest virulence. Three trichothecene genotypes were identified, including nivalenol, 3-acetyldeoxynivalenol, and 15-acetyldeoxynivalenol. No correlation was observed between trichothecene genotype and virulence, except in F. asiaticum. Antifungal assays demonstrated that all six tested fungicides effectively inhibited F. pseudograminearum, with fludioxonil being particularly effective. Field surveys identified isolates resistant to difenoconazole and pyraclostrobin. Laboratory analysis also revealed strains with FpSdhC1A83 V and FpSdhC1S80N mutations conferring resistance to cyclobutrifluram. These findings offer critical insights for developing effective control strategies to manage FCR.

Histone methylation, catalyzed by SET domain-containing lysine methyltransferases, is a conserved epigenetic mechanism regulating gene expression in eukaryotes. However, the evolutionary dynamics of SET domain proteins and their functional interplay in fungi remain poorly understood. Here, we analyzed 18,718 SET domain proteins from 1038 fungal genomes and identified three major clusters, with Cluster 1 enriched for canonical histone methyltransferases. The evolution of the SET domain protein family coordinates with genome expansion in fungi. Functional characterization of seven Cluster 1 proteins in Fusarium graminearum, a globally significant fungal pathogen, reveals diverse roles in growth, development, and virulence. In-depth analyses of two H3K36-specific methyltransferases, Set2 and Ash1, uncover their distinct regulatory functions. Set2-mediated H3K36me3 is enriched in gene bodies of euchromatic regions and facilitates transcription elongation. In contrast, Ash1-mediated H3K36me3 localizes to promoters within facultative heterochromatin and represses transcription. Notably, Ash1-mediated H3K36me3 cooperates with Polycomb repressive complex 2 (PRC2)-dependent H3K27me3 to silence secondary metabolite (SM) gene clusters. Deletion of ASH1 reduces H3K27me3 levels and derepresses SM gene expression. Conversely, Set2-mediated H3K36me3, facilitated by Ctk1-dependent RNA polymerase II phosphorylation, promotes transcriptional elongation of SM genes. Together, these findings reveal evolutionary features of fungal SET domain proteins and uncover a synergistic interplay between H3K36me3 and H3K27me3 in regulating fungal secondary metabolism and virulence. This study advances our understanding of epigenetic regulation in fungi and provides potential targets for controlling fungal pathogens.

Ferroptosis occurs after myocardial ischemia-reperfusion (I/R) injury. Long non-coding RNA (lncRNA) AK020546 possesses the effect of protecting the myocardium from I/R injury; however, the underlying mechanisms remain not fully understood. This study aimed to investigate the effect of AK020546 on ferroptosis in cardiomyocytes and underlying molecular mechanisms. Hypoxia/reoxygenation (H/R) was used to induce H9C2 injury, and an I/R rat model was generated. Ferroptosis was evaluated by detecting lipid reactive oxygen species, Fe2+, glutathione, and malondialdehyde levels. The N(6)-methyladenosine (m6A) methylation of Mst1 was assessed by RNA binding protein immunoprecipitation (RIP), methylated-RIP, luciferase reporter assay, and RNA stability assay. The results showed that AK020546 inhibited H/R-induced ferroptosis. It also decreased Mst1 expression through binding with METTL14. Moreover, METTL14 promoted m6A methylation of Mst1, thereby enhancing its stability. METTL14 and Mst1 were involved in AK020546-mediated ferroptosis. Besides, AK020546 alleviated myocardial damage in I/R rats. In conclusion, the AK020546/METTL14/m6A-Mst1 axis protects against myocardial I/R injury by suppressing cardiomyocyte ferroptosis, suggesting a promising target for myocardial I/R injury.

The SWI/sucrose non-fermentable (SWI/SNF)-facilitated removal of nucleosomes and Spt-Ada-Gcn5 acetyltransferase (SAGA) complex-mediated histone acetylation are crucial for the activation of transcription initiation. However, the mechanism by which these two complexes coordinate to regulate gene expression involved in cell wall remodeling during infection process or in response to external stimuli remains largely unknown in plant pathogenic fungi. Here, we demonstrate that the cell wall integrity (CWI) pathway is activated under toxin (deoxynivalenol)-inducing conditions in the phytopathogenic fungus Fusarium graminearum. This treatment results in the phosphorylation and nuclear translocation of the mitogen-activated protein kinase FgMgv1 in the CWI signaling pathway. Once in the nucleus, the activated FgMgv1 phosphorylates the downstream transcription factor FgRlm1, which binds to a 12- or 14-bp cis-element in the promoters of target genes. Notably, FgMgv1 forms a polymer and interacts with FgRlm1 via its kinase domain. Crucially, this polymerization enables FgMgv1 to recruit both the SWI/SNF and SAGA complexes simultaneously through its C-terminal domain at the target promoters. This coordinated action among FgMgv1, FgRlm1, SWI/SNF, and SAGA ultimately facilitates the transcriptional activation of target genes. Collectively, these findings illuminate a regulatory framework in which Mgv1–Rlm1 axis serves as a key regulatory hub, integrating CWI signals with epigenetic modifications to ensure transcriptional responsiveness to external stimuli.

Although cellular sterol sensing and regulation of sterol biosynthesis are essential processes for eukaryotes, the mechanisms governing ergosterol homeostasis remain largely unknown in pathogenic fungi. In this study, we identify the transcription factor FfSR as a key regulator of sterol homeostasis in Fusarium fujikuroi, the causative agent of rice bakanae disease worldwide. Deletion of FfSR results in reduced ergosterol levels, increasing the susceptibility of F. fujikuroi to azole fungicides. Mechanistically, azole-induced ergosterol depletion promotes FfSR phase separation, which facilitates its binding to cis-elements at target promoters, subsequently activating the expression of ergosterol biosynthesis genes. Conversely, when ergosterol levels are high, ergosterol binds to FfSR, inhibiting its phase separation and transcriptional activation. Additionally, we identify a natural compound, natamycin, as a direct inhibitor of FfSR, suppressing its phase separation and transcriptional capability. These findings highlight a novel mechanism by which fungal pathogens regulate ergosterol homeostasis through transcription factor phase separation, indicating that small molecules targeting FfSR could serve as a synergist to enhance azole efficacy against pathogenic Fusarium.

The forkhead box (FOX) proteins are a conserved family of eukaryotic transcription factors with diverse biological functions. However, the role and evolution of FOX proteins in filamentous fungi have not been fully explored. In this study, we identified and characterized four FOX transcription factors and two forkhead-associated domain (FHA) proteins in the fungal pathogen Fusarium graminearum, the causative agent of Fusarium head blight on cereal crops. Our results reveal that the FOX transcription factors FgHcm1 and FgFoxO4, along with the FHA domain-containing protein FgFha1, are essential for the full virulence of F. graminearum. These proteins regulate fungal penetration of wheat spikelets by modulating the expression of penetration-related genes. In addition, FgHcm1, FgFoxO4, and FgFha1 influence both asexual and sexual reproduction. Deletion of these genes also confers increased sensitivity to various stressors including fungicides. Notably, the gene deletion mutants ΔFgHCM1, ΔFgFOXO4, and ΔFgFHA1 exhibited heightened sensitivity to tebuconazole, and these proteins regulate the inducible expression of the FgCYP51A gene in response to tebuconazole treatment. Furthermore, ΔFgHCM1 mutants showed increased sensitivity to rapamycin with impaired autophagic responses. This study represents the first comprehensive analysis of FOX and FHA proteins in a filamentous fungal pathogen, highlighting their critical roles in virulence, stress tolerance, and fungicide resistance in F. graminearum.

The methane-dependent arsenate reduction (M-AsR) process is recently demonstrated to enhance the release of mobile and toxic arsenite [As(III)], with critical implications for ecosystem safety in wetland ecosystem. However, the key functional microorganisms and the underlying metabolic mechanisms of M-AsR in wetland remain unclear. In this study, 13C-labeled methane (CH4) was used as the sole carbon source to track the active microorganisms responsible for M-AsR. DNA-stable isotope probing (DNA-SIP) combined with amplicon and metagenomic sequencing was further employed to identify the microorganisms involved in M-AsR. The results showed that arsenate [As(V)] reduction occurred exclusively in the treatment amended with both CH₄ and As(V). After a 50-day incubation, significant shifts in the relative abundance of functional genes (pmoA, ANME-2d mcrA and arrA) were observed in the heavy DNA fractions from the treatment amended with 13CH4 and As(V), indicating the incorporation of 13C into M-AsR microorganisms. Furthermore, the Methanobacterium, Methylobacter and arsenate-reducing bacteria (Chryseobacterium and Hydrogenophaga) were the predominant genera in 13CH4-As heavy fractions and identified as the potential microorganisms responsible for M-AsR in wetland. Metagenomic analysis further confirmed that most of these microorganisms contained genes for CH4 oxidation and As(V) reduction. This multi-omics approach provides mechanistic insights into microbial mediated As(V) reduction in methane-rich wetland area.

Organisms exposed to environmental stimuli can develop "memory" of those experiences, a phenomenon known as priming, which allows them to better adapt to subsequent stimuli. Growing evidence has shown that fungi can "remember" past encounters, but the priming effect remains poorly understood in phytopathogenic fungi. In this study, we examined the priming effect in Fusarium graminearum, the causative agent of Fusarium head blight (FHB), by culturing its conidia in the presence of a median effective concentration (EC50) of triazole fungicide. We observed that primed conidia exhibited significantly higher germination rates and longer hyphal length than unprimed conidia when exposed to double EC50 concentration of triazole. The triazole priming effect in F. graminearum was retained in conidia for an extended period but was not stably heritable. Further investigations revealed that this priming effect was linked to increased over-expression of the fungicide target genes (FgCYP51s) above the level seen in non-primed F. graminearum. This study reveals that F. graminearum develops adaptive resistance following treatment with triazole fungicides, and elucidates the mechanism behind priming, which is regulated by the transcription factor FgSR. This regulation leads to the upregulation of FgCYP51 expression, thereby mediating the observed adaptive resistance. This provides a theoretical basis for understanding the development of resistance in pathogens and offers relevant guidance for the use of triazole fungicides in the control of FHB.

Phenazines are highly prevalent, natural bioactive substances secreted by microbes. However, their mode of action and potential involvement in shaping microbiomes remain elusive. Here we performed a comprehensive analysis of over 1.35 million bacterial genomes to identify phenazine-producing bacteria distributed across 193 species in 34 families. Analysis of rhizosphere microbiome and public rhizosphere metagenomic datasets revealed that phenazines could shape the microbial community by inhibiting Gram-positive bacteria, which was verified by pairwise interaction assays using Phenazine-1-carboxamide (PCN)-producing Pseudomonas chlororaphis. PCN induced DNA damage in Bacillus subtilis, a model Gram-positive target, where it directly bound to the bacterial topoisomerase IV, inhibiting its decatenation activity and leading to cell death. A two-species consortium of phenazine-producing Pseudomonas and resistant B. subtilis exhibited superior synergistic activity in preventing Fusarium crown rot in wheat plants. This work advances our understanding of a prevalent microbial interaction and its potential for biocontrol.