BIOINFORMATIC AND FUNCTIONAL CHARACTERIZATION OF THE VITELLOGENIN RECEPTOR IN DIPETALOGASTER MAXIMA, A VECTOR OF CHAGAS DISEASE.

RNAi-mediated knockdown of vitellogenin genes disrupts ovary development and alters ovarian bacterial composition in Zeugodacus tau (Diptera: Tephritidae).

The vitellogenesis pathway of the tick host Ixodes scapularis regulates Rickettsia buchneri transcriptional state and persistence

Obligate endosymbionts relying on transovarial transmission must coordinate with host reproduction, yet the regulatory mechanisms remain poorly understood. Here we show that the vitellogenesis pathway of the tick Ixodes scapularis regulates the transcriptional state of its endosymbiont Rickettsia buchneri (Rb), separately from its abundance. In males, Rb DNA remained detectable, but bacterial transcription was strongly reduced across all examined genes, with markedly lower RNA/DNA ratios. In females, Rb was restricted to ovarian tissues (developing oocytes and interstitial cells), with no detection in salivary glands or midgut by TEM, FISH, or PCR. After blood feeding, Rb density within size-matched early-stage oocytes was significantly reduced. We further characterized two gene families mediating vitellogenesis: vitellogenin synthesis genes (Vgs, n = 20) and vitellogenin receptor genes (Vgr, n = 15). Vgs proteins showed conserved domain organization, whereas Vgr paralogs showed greater structural diversification. RNAi silencing of Vgs20 or Vgr12 altered Rb transcriptional profiles in both tick cells and ticks, although changes in bacterial load were not consistent between the two systems. Antibiotic depletion of Rb increased expression of both host genes. Together, these findings show that tick vitellogenesis pathways contribute to Rb regulation during reproduction and identify the I. scapularis-Rb system as a useful model for studying host control of obligate endosymbionts. IMPORTANCE Maternally inherited bacterial symbionts are commonly characterized by bacterial abundance. This study shows that bacterial abundance alone does not necessarily reflect symbiont functional state. In the blacklegged tick, the inherited symbiont Rickettsia buchneri persists in males but exhibits little transcriptional activity. In females, the host reproductive pathway determines whether the symbiont is active: silencing one component of this pathway changed bacterial gene expression without changing bacterial abundance. Hosts therefore regulate not only the abundance of inherited symbionts, but also their functional state. The same bacterial abundance can correspond to very different functional states. Understanding inherited symbioses therefore requires considering bacterial function alongside bacterial abundance.

Oosporein in Phlebia centrifuga: Prediction of Key Enzymes by a Combination of Multi-Omics and Molecular Dynamics

The red bibenzoquinone oosporein, a promising biocontrol agent with potential to replace conventional pesticides in insect pest management in crops, was produced by the Basidiomycota Phlebia centrifuga P. Karst isolated from the Black Forest National Park. A submerged system was established, yielding up to 1.60 g L–1 oosporein in the culture supernatant upon supplementation with the key intermediate orsellinic acid, which strongly induced oosporein-specific biosynthetic genes. Using a multiomics approach, genes encoding enzymes for all necessary conversions were predicted, including a type I polyketide synthase, two monooxygenases, and a heme peroxidase. Enzymatic functions were investigated by extensive docking analyses and molecular dynamics simulations, ultimately leading to the prediction of the underlying biosynthetic pathway. In summary, a spore-free, high-yield, scalable production platform for oosporein was established, highlighting the potential of rare, protected fungal species as sources for valuable enzymes and bioactive secondary metabolites for efficient microbial biomanufacturing systems.

The versatile TRAF3 from sea urchin (Strongylocentrotus intermedius) interacts with MyD88 and regulates host immunity.

Tumor necrosis factor receptor-associated factor 3 (TRAF3) serves as a versatile protein in innate immune signaling cascades, playing a pivotal role in modulating inflammatory responses, cellular homeostasis, and anti-infection immunity. Here, we cloned and identified a novel TRAF3 homolog (SiTRAF3) from the sea urchin Strongylocentrotus intermedius. Sequence analysis revealed that the open reading frame (ORF) of SiTRAF3 is 1914 bp in length, encoding a polypeptide of 637 amino acids. Structural prediction indicated the presence of a typical N-terminal RING finger domain, zinc finger motifs, and a highly conserved C-terminal MATH domain. Phylogenetic analysis demonstrated that SiTRAF3 clustered closely with the TRAF3-like protein from the sea urchin Strongylocentrotus purpuratus, underscoring the evolutionary conservation of TRAF3 among echinoderms. Quantitative reverse transcription PCR (qRT-PCR) assays revealed ubiquitous expression of SiTRAF3 transcripts across various tissues of S. intermedius, with the highest levels detected in the gill and coelomocytes. Following immune challenges with lipopolysaccharide (LPS), poly(I:C), and peptidoglycan, SiTRAF3 mRNA expression was significantly upregulated, suggesting an active, broad-spectrum response to diverse pathogenic stimuli. In addition, RNA interference of SiTRAF3 suppresses the expression of IL-17 family members and strongylocins following LPS stimulation. Co-immunoprecipitation (Co-IP) assays further verified a direct physical interaction between SiTRAF3 and the core adaptor protein SiMyD88. Functional analyses in HEK293T cells demonstrated that ectopic expression of SiTRAF3 notably elevated the phosphorylation levels of JNK, Erk1/2, and p38 MAPKs, and synergistically enhanced LPS-induced signaling activation. Furthermore, dual-luciferase reporter (DLR) assays revealed the broad-spectrum transcriptional activation capacity of SiTRAF3, as it significantly drove the promoter activities of nuclear factor kappa-B, activator protein-1, signal transducer and activator of transcription 3, along with multiple inflammatory cytokines (tumor necrosis factor-alpha, interleukin-6) and interferon-related elements (interferon α/β/γ, interferon-stimulated response element). Taken together, these findings identify SiTRAF3 as a pivotal scaffold protein in the echinoid innate immune system. By assembling a signaling complex with SiMyD88, SiTRAF3 mediates the activation of MAPK cascades and multiple downstream immune transcriptional pathways. This study not only elucidates the functional significance of SiTRAF3 within the invertebrate immune regulatory network but also provides a theoretical foundation for the molecular breeding of disease-resistant sea urchin strains.

The first dipteran luciferase from Keroplatus testaceus evolved by repurposing of a storage protein.

Bioluminescence has independently evolved multiple times during animal evolution, yet the biochemical mechanisms underlying light production in true flies (Diptera) remain poorly understood. Here we describe the identification, cloning and functional characterization of the first dipteran luciferase from the fungus gnat Keroplatus testaceus. The 76‑kDa enzyme, KerLuc, is active when expressed heterologously in yeast, yielding a blue emission spectrum indistinguishable from native larvae. Sequence and domain analyses place the protein within the hemocyanin/hexamerin superfamily, but intriguingly, the protein lacks the canonical copper‑binding histidines. Structure prediction points to a hydrophobic cavity consistent with binding of the known Keroplatus oxyluciferin (3‑hydroxykynurenic acid), suggesting a novel catalytic mechanism. The luciferase is encoded by a single‑exon gene adjacent to a closely related paralogue within the conserved Enhancer of Split complex locus, indicating recent duplication and possible neofunctionalisation. Phylogenetic comparisons highlight proximity to Orfelia homologues and support independent origins of bioluminescence within Keroplatidae. Our results provide compelling evidence for a novel evolutionary origin for a luciferase from a storage protein, thereby closing a long-standing gap in understanding the molecular mechanisms of bioluminescence in Diptera.

Identification of Partitivirus-like RdRPs in the Brevipalpus yothersi Genome Supports Viral-to-Arthropod Horizontal Gene Transfer

RNA-dependent RNA polymerases (RdRPs) are essential enzymes involved in RNA virus replication and eukaryotic RNA silencing. They are generally absent in vertebrates but present in some invertebrate lineages, such as nematodes and certain arthropods. Brevipalpus yothersi is a phytophagous mite of agricultural relevance due to its role as a vector of plant-infecting viruses. We have identified two RdRPs on the genome of this mite species that, unexpectedly, are not of eukaryotic origin. Phylogenetic reconstruction and comparisons of 3D protein structures revealed similarity with viral RdRPs of the Partitiviridae family. Both RdRPs retain conserved catalytic motifs at the protein sequence level, and expression was confirmed by RNAseq and qPCR across mite developmental stages, with a peak during the nymphal stage. K-mer profiles showed similarity with mite endogenous genes, suggesting gene amelioration after the integration, or being derived from a viral donor already adapted to the mite. Our study also identified orthologs in other Brevipalpus species, but not in other Acari relatives, supporting that the horizontal gene transfer event is circumscribed to the Brevipalpus genus. These findings highlight an intriguing case of viral gene domestication in arthropods that might influence their developmental biology and the host–virus interaction.