Insect Biochemistry and Molecular Biology最新文献

Ovarian development and reproductive maturation in Aedes aegypti critically depend on 20-hydroxyecdysone (20E) signaling and energy metabolic homeostasis. Knockdown of EF1A2 (AAEL017096) profoundly affects lipid nutrient metabolism in the fat body and ovarian development, yet whether this effect involves 20E signaling has remained unclear. Here, we show that EF1A2 silencing significantly suppresses transcription of the Halloween genes CYP314A1, CYP315A1, and CYP307A1, reduces endogenous 20E titers, and blocks downstream hormonal signaling. Differential proteomic analysis reveals that EF1A2 broadly participates in metabolic reprogramming, endoplasmic reticulum protein processing, and mRNA regulation. The poly(A)-binding protein PABP (AAEL010318) is identified as a core downstream effector and is specifically highly expressed in ovaries of post-blood-meal mosquitoes. PABP knockdown causes aberrant lipid accumulation in the fat body, disrupts glycolipid metabolism, and ultimately arrests ovarian development and impairs reproductive output, recapitulating EF1A2 knockdown phenotypes. Mechanistically, EF1A2 directly binds full-length PABP via its domain I, positively regulating PABP protein abundance. PABP regulates transcription of the Halloween genes CYP302A1, CYP306A1, and CYP315A1, thereby modulating 20E titer and, in turn, Vg transcription. PABP also associates with the 3' UTR of E75 mRNA, stabilizing the transcript and activating downstream 20E signaling. E75 positively regulates CYP302A1, CYP306A1, and CYP314A1 through a positive feedback mechanism; E75 downregulation impairs ovarian development and reduces egg production. Collectively, the EF1A2-PABP-E75 axis coordinates hormone biosynthesis and metabolic homeostasis to sustain ovarian development and reproductive capacity, offering molecular targets for mosquito vector control.

Insect herbivores employ many adaptive mechanisms to successfully colonize host plants. Transcriptional plasticity is one mechanism that enables broad host use in polyphagous insect herbivores; however, whether specialist herbivores use transcriptional plasticity or undergo epigenome modifications to cope with host shifts remains largely unexplored. We measured the transcriptome and DNA methylome of the specialist aphid herbivore, Daktulosphaira vitifoliae, after transfer to new hosts using RNA-seq and whole-genome bisulfite sequencing. We found the transcriptome of D. vitifoliae is highly plastic and responsive to host genotype - 3,706 genes (28% of expressed genes) are significantly differentially expressed in at least one contrast comparing host genotypes. Most host-responsive genes (2,667 genes) were upregulated when feeding on the non-native host Vitis vinifera and enriched for genes encoding predicted secretory effectors, detoxification enzymes, and cuticle- and chitin-associated proteins. We found gene co-expression modules that correlated with variation in fitness suggesting plasticity in transcriptional networks contributes to D. vitifoliae adaptation to host plants. We also identified a previously undescribed family of proline-rich secretory proteins conserved across the Aphidomorpha infraorder of insects. Our analysis of DNA methylation revealed a negative correlation between gene-level CpG methylation and host-induced transcriptional plasticity, suggesting a lack of gene body methylation may enable transcriptional plasticity when insects move to new host environments. Together, these results provide new insight into the molecular mechanisms of host use in insect herbivores.

Although cytochrome P450-mediated xenobiotic detoxification has been well characterized, comparatively little is known about how naturally occurring host-plant phytochemicals regulate the expression of insect detoxification genes. Capsaicin, a defensive alkaloid produced by chili pepper (Capsicum annuum), deters many vertebrate and invertebrate herbivores, whereas the generalist crop pest Helicoverpa armigera tolerates and metabolizes this compound. In this species, the cytochrome P450 enzyme CYP6AN1 contributes to capsaicin metabolism, and the corresponding gene is strongly induced by capsaicin; however, the regulatory mechanism underlying this induction has remained unknown. Using dual-luciferase reporter assays, we identified several cis-regulatory regions with positive or negative effects on the capsaicin responsiveness of the CYP6AN1 promoter. An element essential for capsaicin induction of CYP6AN1, designated the xenobiotic response element to capsaicin (XRE-Cap), was fine-mapped to positions -57 to -44 and found to consist of a direct repeat of the CAAAGTC motif. Internal deletions and site-directed mutagenesis demonstrated that both the nucleotide sequence and direct-repeat architecture of XRE-Cap contribute to maximal induction. Conversely, converting a single distal CAAAGTC motif into a second XRE-Cap copy significantly enhanced capsaicin induction, thereby providing gain-of-function evidence for the regulatory importance of this direct-repeat architecture. Electrophoretic mobility shift assays showed that XRE-Cap is specifically bound by nuclear proteins from Helicoverpa zea fat body cells. Together, these findings identify and functionally characterize XRE-Cap as a cis-regulatory element required for capsaicin-inducible expression of CYP6AN1 and provide mechanistic insight into phytochemical-responsive regulation of detoxification genes in lepidopterans.

Symbiotic microorganisms and endogenous viruses play fundamental roles in shaping host physiology, yet the non-coding RNA-mediated mechanisms underlying viral-driven metabolic reprogramming remain poorly understood. Here, we demonstrate a novel mechanism of cross-kingdom metabolic manipulation mediated by symbiotic viral microRNAs. Using the tobacco cutworm Spodoptera litura, its endoparasitoid wasp Snellenius manilae, and the wasp-associated bracovirus SmBV as a model system, we identified two virus-derived microRNAs, SmBV-miR-2733 and SmBV-miR-3772, that target the host lipogenic genes encoding AGPAT and lipin, leading to the suppression of glycerolipid biosynthesis and reduced lipid accumulation. Functional analyses revealed that these symbiotic viral miRNAs concurrently attenuate host innate immune signaling pathways (Toll and JAK/STAT) and cellular encapsulation, establishing a mechanistic link between viral metabolic coercion and host immunomodulation. Crucially, this viral microRNA-mediated regulation is transient and highly dynamic; miRNA expression declines during later stages of infection, coinciding with an intermediate recovery of host lipogenic gene expression and lipid reserves, which may help sustain the developing parasitoids. Prolonged experimental maintenance of viral miRNAs disrupted host energetic homeostasis and compromised symbiotic success, underscoring the evolutionary necessity of temporally coordinated metabolic regulation. Comparative genomic analysis identified miR-2733- and miR-3772-like candidate loci with miRNA-precursor-like structures that are unevenly distributed across related bracovirus lineages. Together, our findings provide significant mechanistic insights into how symbiotic viral effectors post-transcriptionally rewire host metabolic and immune networks, offering a broader conceptual framework for understanding homeostasis in diverse host-microbe symbioses.

Triacylglycerol (TAG) is the main lipid reserve in insect eggs and represents a critical energy source for embryonic development. In the hematophagous vector Rhodnius prolixus, oocyte growth relies heavily on lipids transported from the fat body through the hemolymph, which are delivered mainly as diacylglycerol and free fatty acids. Therefore, the ovary must catalyze the final esterification step converting these substrates into TAG to ensure proper lipid storage in developing oocytes. This reaction is mediated by diacylglycerol acyltransferases (DGATs), yet the specific roles of DGAT1 and DGAT2 in insect reproduction remain poorly understood. Here, we investigated the functions of RpDGAT1 and RpDGAT2 in ovarian physiology and reproduction in R. prolixus. RNAi gene knockdown efficiently reduced transcript levels of both enzymes without affecting blood digestion, survival, or oviposition rates. However, RpDGAT1 knockdown caused pronounced reproductive defects. Females produced morphologically abnormal eggs with collapsed appearance and reduced size, leading to an approximately 70% reduction in hatching rates. Ultrastructural analyses revealed alterations in eggshell organization as seen by SEM, but no changes in the hydrocarbon contents of the eggshell were detected. Lipid analyses showed that RpDGAT1 knockdown reduced ovarian TAG levels at day 10 after blood feeding, but not at day 5, while increasing ovarian phosphatidylethanolamine at day 10. Ovarian free fatty acid levels remained unchanged at both time points. In phenotypic eggs, TAG levels were reduced and free fatty acids were increased. Consistently, oocytes from RpDGAT1-knocked down females exhibited smaller lipid droplets, reflecting impaired neutral lipid storage. Hemolymph lipid composition and ovarian incorporation of radiolabeled palmitate were unchanged, indicating that reduced ovarian TAG was not associated with detectable changes in circulating lipid availability or overall ovarian fatty acid uptake. In contrast, RpDGAT2 knockdown produced no detectable reproductive phenotype. Together, our findings demonstrate that RpDGAT1, but not RpDGAT2, is essential for TAG accumulation, proper oocyte development, and egg hatchability in R. prolixus, highlighting the importance of lipid metabolism in insect reproductive physiology.