Search Results (292)

Grapholita molesta is a globally significant fruit pest. Females achieve maximal reproductive output through efficient sperm utilization following a single copulation. Post-mating maturation of eupyrene sperm is a critical step in reproductive success. Here, we report that a male accessory gland-derived serine protease (named GmAGSP1) is essential for this process. GmAGSP1 was only distantly related to other identified sperm-activating SPs, and its transcript was highly expressed in the AG at 48 h after emergence. RNAi-mediated knockdown of GmAGSP1 in males did not affect courtship rate, copulation duration, or mating frequency, whereas male fertility decreased significantly. Mating with GmAGSP1-knockdown males markedly impaired eupyrene sperm maturation in the spermatophores, with phenotypes including failure of eupyrene sperm bundles to dissociate normally and marked reduction in viability of the dissociated eupyrene sperm. Finally, untargeted metabolomic analysis preliminarily demonstrated marked alterations in multiple metabolic pathways within the spermatophore following mating with GmAGSP1-knockdown males. This study advances our understanding of the regulatory mechanism of “sperm activation in the spermatophore’s metabolic microenvironment mediated by male AG-derived SP” while providing critical insights for the development of novel genetic control strategies targeting G. molesta. Full article

The brown planthopper (BPH), Nilaparvata lugens, is the most destructive insect pest of rice. BPH infestations severely threaten rice yield worldwide. The gustatory receptor NlGr23 plays a critical role in mediating the repulsive reaction to oxalic acid of the BPH. We integrated transcriptomic and proteomic analyses to determine the metabolic and behavioral consequences of NlGr23 silencing. The RNAi-mediated knockdown of NlGr23 increased body weight and honeydew production, indicating enhanced feeding activity. The results of multiomics profiling revealed disrupted lipid homeostasis, identifying 187 differentially expressed genes and 150 differentially expressed proteins. These genes were enriched in pathways including glycerophospholipid metabolism, fatty acid biosynthesis, and AMPK signaling. The results of biochemical assays showed that NlGr23 silencing elevated triacylglycerol levels by 68.83%, and reduced glycerol and free fatty acid levels, suggesting impaired lipolysis. The NlGr23 loss-of-function mutation mechanistically activates the AMPK pathway, suppresses lipid breakdown, and promotes energy storage. This study established NlGr23 as a key regulator linking chemosensation to metabolic reprogramming, providing new insights into gustatory receptor-mediated energy homeostasis in the BPH. Full article

Soybean meal (SBM) is extensively used as a predominant protein source in animal feed. However, raw soybean cannot be directly utilized in animal feed, due to the presence of the Kunitz trypsin inhibitor (KTi) and the Bowman–Birk protease inhibitor (BBi). These antinutritional factors inhibit the digestive enzymes in animals, trypsin and chymotrypsin, resulting in poor animal performance. To inactivate the activity of protease inhibitors, SBM is subjected to heat processing, a procedure that can negatively impact the soybean protein quality. Thus, it would be beneficial to develop soybean varieties with little or no trypsin inhibitors. In this study, we report on the creation of experimental soybean lines with significantly reduced levels of Bowman–Birk protease inhibitors. RNA interference (RNAi) technology was employed to generate several transgenic soybean lines. Some of these BBi knockdown soybean lines showed significantly lower amounts of both trypsin and chymotrypsin inhibitor activities. Western blot analysis revealed the complete absence of BBi in selected RNAi-derived lines. RNA sequencing (RNAseq) analysis demonstrated a drastic reduction in the seed-specific expression of BBi genes in the transgenic soybean lines during seed development. Confocal fluorescence immunolabeling studies showed that the accumulation of BBi was drastically diminished in BBi knockdown lines compared to wild-type soybeans. The absence of BBi in the transgenic soybean did not alter the overall protein, oil, and sulfur amino acid content of the seeds compared to wild-type soybeans. The seed protein from the BBi knockdown lines were more rapidly hydrolyzed by trypsin and chymotrypsin compared to the wild type, indicating that the absence of BBi enhances protein digestibility. Our study suggests that these BBi knockdown lines could be a valuable resource in order for plant breeders to incorporate this trait into commercial soybean cultivars, potentially enabling the use of raw soybeans in animal feed. Full article

As a commercially valuable aquaculture species, rainbow trout (Oncorhynchus mykiss) urgently require solutions to growth inhibition associated with reproductive development. To elucidate the function of the cell cycle regulator Cyclin E genes (CCNE1 and CCNE2) in this process, we cloned the genes and analyzed their relative expression across various tissues and gonadal developmental stages. Using RNA interference (RNAi) and overexpression in RTG2 cells, we examined the effects of CCNE on cell viability, proliferation, and meiotic gene expression. Results showed that the open reading frame lengths of CCNE1 and CCNE2 were 1230 bp and 1188 bp, encoding 408 and 395 amino acids, respectively. Both proteins contain two conserved cyclin boxes, exhibit high structural similarity, and are phylogenetically most closely related to Oncorhynchus tshawytscha and Oncorhynchus kisutch. Expression and localization analyses revealed that CCNE1 was highly expressed in the ovary, while CCNE2 was highly expressed in the testis. Both proteins were expressed during fertilized egg development and key gonadal stages (at 13, 21, and 35 months post-fertilization). CCNE expression positively correlated with RTG2 cell viability and proliferation, with immunofluorescence confirming that CCNE is localized in the nucleus. Knockdown or overexpression of CCNE induced the differential expression of reproductive-related genes and key meiotic regulators. These findings suggest that CCNE1 and CCNE2 balance meiosis and gamete development through specific regulatory mechanisms, and their dysregulation may be a key factor underlying meiosis inhibition and reproductive development abnormalities. Full article

The Colorado potato beetle (Leptinotarsa decemlineata, CPB) is a major pest in potato crops, notorious for its rapid dispersal and insecticide resistance, which are enabled by its robust elytra and flight-capable hindwings. The Miniature (Mi) gene, encoding a protein with a zona pellucida (ZP) domain, is involved in wing development and cuticle integrity, yet its functional role in beetles remains underexplored. In this study, we cloned and characterized the LdMi gene in the CPB and investigated its function using RNA interference (RNAi), morphological analyses, and spectroscopy. LdMi encodes a 146.35 kDa transmembrane protein with a conserved ZP domain, clusters with coleopteran homologs, and exhibits relative conservation across insect species. Expression profiling showed high LdMi transcript levels in the hindwings, the elytra, and the pupal stages. RNAi knockdown in fourth-instar larvae resulted in severe eclosion defects, including malformed wings and reduced adult weight. Scanning electron microscopy (SEM) revealed disrupted elytral patterns and deformed hindwing veins in knockdown individuals. Spectroscopic analyses using Fourier-transform infrared (FTIR) and Raman spectroscopy indicated a reduction in protein–chitin crosslinking and diminished hydrogen bonding, suggesting compromised cuticular integrity. These results highlight the essential role of LdMi in cuticle formation and the surface morphology of the elytra and hindwings, offering new insights into ZP domain proteins in insects. Full article

Cytoplasmic polyadenylation element-binding proteins (CPEBs) are critical regulators of maternal mRNA translation during oogenesis, yet their roles in insect reproduction remain underexplored. Here, we characterized CmCPEB2, a CPEB homolog in the rice leaf roller Cnaphalocrocis medinalis, a destructive lepidopteran pest insect, and elucidated its role in mating-induced oviposition. The CmCPEB2 protein harbored conserved RNA recognition motifs and a ZZ-type zinc finger domain and was phylogenetically clustered with lepidopteran orthologs. Spatiotemporal expression profiling revealed CmCPEB2 was predominantly expressed in ovaries post-mating, peaking at 12 h with a 6.75-fold increase in transcript levels. Liposome-mediated RNA interference targeting CmCPEB2 resulted in a 52% reduction in transcript abundance, leading to significant defects in ovarian maturation, diminished vitellogenin deposition, and a 36.7% decline in fecundity. The transcriptomic analysis of RNAi-treated ovaries identified 512 differentially expressed genes, with downregulated genes enriched in chorion formation and epithelial cell development. Tissue culture-based hormonal assays demonstrated the juvenile hormone-dependent regulation of CmCPEB2, as JH treatment induced its transcription, while knockdown of the JH-responsive transcription factor CmKr-h1 in the moths suppressed CmCPEB2 expression post-mating. These findings established CmCPEB2 as a juvenile hormone-dependent regulator of mating-induced oviposition that orchestrates vitellogenesis through yolk protein synthesis and ovarian deposition and choriogenesis via transcriptional control of chorion-related genes. This study provides novel evidence of CPEB2-mediated reproductive regulation in Lepidoptera, highlighting its dual role in nutrient allocation and structural eggshell formation during insect oogenesis and oviposition. Full article

Abiotic stressors threaten honeybee health, jeopardizing pollination services critical to agriculture and biodiversity. Here, we identified the AmVgR gene, which encodes a member of the low-density lipoprotein receptor family, and examined its function in the response of Apis mellifera to adverse abiotic stress. AmVgR exhibited peak expression in adult workers and was significantly upregulated under heat, cold, heavy metal, and pesticide exposure. RNAi-mediated knockdown of AmVgR suppressed antioxidant enzyme activities, elevated the levels of oxidative damage markers, and downregulated antioxidant gene expression. Crucially, AmVgR silencing reduced survival under H₂O₂-induced oxidative stress, indicating its essential role in stress resilience. Our findings highlight AmVgR as a key regulator of antioxidant defense during development and environmental adaptation in Apis mellifera. This study provides mechanistic insights into bee stress physiology and proposes AmVgR as a novel target for enhancing pollinator protection strategies. Further research should elucidate its molecular pathways and translational applications in mitigating abiotic stress impacts. Full article

TDP-43 is an RNA-binding protein linked to amyotrophic lateral sclerosis (ALS), possibly associated with a role in miRNA biogenesis, which is still not fully understood. Herein we investigated the impact of the Drosophila homolog of TDP-43, TBPH, on genes related to miRNA biogenesis. A TBPH knockout significantly reduced mRNA transcription and protein levels of DCR-1 and DCR-2, whereas an overexpression of DCR-1 and DCR-2 in a TBPH knockdown background exacerbated compound eye damage, with variations in severity that were sex-dependent. Neuronal TBPH RNAi consistently shortened lifespan, with males and females exhibiting distinct survival profiles. DCR-1 and DCR-2 knockdown worsened the locomotor defects induced by TBPH deficiency, thus reinforcing the functional link between TBPH and DCR. In TBPH-deficient flies, the pharmacological activation of Dicer promoted reverse locomotion behavior, with a preference for backward movement. Overall, we show that TBPH is a key regulator of DCR protein expression, highlighting its conserved role in miRNA dysregulation associated with motor function and cytotoxicity in ALS-like pathology in Drosophila models. Full article

The diamide insecticide tetraniliprole is a valuable tool for managing major insect pests like the invasive tomato pinworm, Tuta absoluta (Meyrick). However, the mechanisms underlying tetraniliprole resistance, as well as its associated fitness costs, remain unclear. In this study, we assessed the fitness of tetraniliprole-resistant (TetraRS) and susceptible (SS) strains of T. absoluta and conducted Illumina RNA-seq to compare their transcriptomes. We also used nanocarrier-mediated RNA interference (RNAi) to knockdown P450 genes and evaluate their role in tetraniliprole resistance. After eight generations of selection, T. absoluta developed a 20.80-fold resistance to tetraniliprole, accompanied by fitness costs. RNA-seq analysis revealed 3332 differentially expressed genes (DEGs), with 1707 upregulated and 1625 downregulated in the TetraRS compared to the SS strain. Gene Ontology (GO) annotations showed significant enrichment in categories related to metabolic processes, cellular processes, catalytic activity, cellular anatomical entity, and binding. These genes were also identified in key KEGG pathways such as cytochrome P450, drug metabolism, carbon metabolism, oxidative phosphorylation, fatty acid metabolism, and protein processing. RT-qPCR analysis confirmed that P450 genes (CYP405D1, CYP6AB269, and CYP4AU1) were upregulated in TetraRS insects, in line with the RNA-seq results. Cytochrome P450 activity was significantly higher in the TetraRS strain than in the SS strain. Notably, nano-encapsulated dsRNA targeting these overexpressed P450 genes increased the susceptibility of T. absoluta to tetraniliprole. Further, cytochrome P450 activity was significantly reduced following silencing of P450 genes. These findings suggest that multiple genes and pathways, particularly P450 genes, contribute to tetraniliprole resistance in T. absoluta. This study provides valuable insights into the molecular mechanisms underlying insecticide resistance in this key pest species. Full article

10-Hydroxy-2-decenoic acid (10-HDA), a major fatty acid (FA) component of royal jelly, is synthesized in the mandibular glands (MGs) of worker honeybees. Despite its well-documented nutritional and therapeutic significance, the biosynthetic pathway and regulatory mechanisms of 10-HDA production remain largely unresolved. In this study, the molecular basis of 10-HDA biosynthesis and regulation in the MGs of newly emerged bees (NEBs), nurse bees (NBs), and forager bees (FBs) were investigated using RNA sequencing and weighted gene co-expression network analysis (WGCNA). A five-step biosynthetic pathway for 10-HDA was proposed, and cross-species analysis of Apis mellifera and A. cerana revealed the conserved expression patterns of 15 key enzymes involved. Functional validation via RNA interference (RNAi) demonstrated that knockdown of acyl-CoA Delta(11) desaturase (d11ds, LOC551527), a key enzyme in FA desaturation, led to a 50% reduction in 10-HDA levels. Protein–protein interaction (PPI) network analysis further identified transcriptional regulators Kay and Drep-2 as potential modulators of 10-HDA metabolism. This study provides the first comprehensive mechanistic model of 10-HDA biosynthesis in honeybee MGs and highlights the labor-specific regulation of FA metabolism. These findings offer promising genetic targets for improving the royal jelly quality through genetic technology. Full article

In eukaryotes, a lot of proteins are transported across the endoplasmic reticulum by the heterotrimeric Sec61 channel. And post-translational transport needs another Sec62/Sec63 complex. However, functions of these genes are poorly explored in insects. In this study, we first identified five Sec genes, named Sec61α, Sec61β, Sec61γ, Sec62 and Sec63, in Locusta migratoria. Quantitative reverse-transcription polymerase chain reaction (RT-qPCR) analysis showed that these five genes were expressed in muti-tissues, including wing pad, leg, foregut, midgut, gastric cecum, hindgut, and highly expressed in the integument. Knockdown of LmSec61α and LmSec61γ by RNA interference (RNAi) lead to the feeding cessation with a mortality rate of 100%. However, there is only 13.4% of dsLmSec61β-injected nymphs died before molting. All nymphs injected with dsLmSec61α and dsLmSec61γ died before molting with the gut atrophy. Furthermore, hematoxylin–eosin staining indicated that the cells of the midguts and gastric caecum were defective, and the microvilli and peritrophic matrix were destroyed seriously after silencing LmSec61α and LmSec61γ. Knockdown of LmSec62 and LmSec63 resulted in high mortality before and during molting. The hematoxylin–eosin (HE) staining and transmission electron microscopy (TEM) results showed that both the formation of the new cuticle and the degradation of the old cuticle were inhibited in dsLmSec63-injected insects compared to the controls. Especially, there was no obvious plaques on microvillar tips of the epidermal cells after silencing of LmSec63. These results revealed that Sec61s and Sec62/Sec63 genes are required in the gut and cuticle development of locusts. Therefore, these genes are potential targets for the control of locusts. Full article

Background/Objectives: Essential genes are those required for an organism’s survival and reproduction. However, gene essentiality is not absolute; it can be highly context-dependent, varying across genetic and environmental conditions. Most previous studies have assessed gene essentiality in a single genetic background, limiting our understanding of its variability. The objective of this study was to investigate how genetic background influences gene essentiality in the multicellular model organism Caenorhabditis elegans. Methods: We examined gene essentiality in three genetically distinct C. elegans strains: N2, LKC34, and MY16. A total of 294 genes were selected for RNA interference (RNAi) knockdown: 101 previously classified as essential, 175 as nonessential and 18 as conditional (condition-dependent essentiality). Each gene–strain combination was tested in multiple biological and technical replicates, and rigorous quality control and statistical analyses were used to identify strain-specific effects. Results: Our results demonstrate substantial variation in gene essentiality across genetic backgrounds. Among the 101 genes previously identified as essential in the N2 strain, only 56% were consistently essential in all three strains. We identified 23 genes that were newly essential across all strains, 13 genes essential in two strains, and 9 genes essential in only one strain. These results reveal that a significant proportion of essential genes exhibit strain-dependent essentiality. Conclusions: This study underscores the importance of genetic context in determining gene essentiality. Our findings suggest that relying on a single genetic background, such as N2, may lead to an incomplete or misleading view of gene essentiality. Understanding context-dependent gene essentiality has important implications for functional genomics, evolutionary biology, and potentially for translational research where genetic background can modulate phenotypic outcomes. Full article

The Colorado Potato Beetle (CPB, Leptinotarsa decemlineata Say, Coleoptera: Chrysomelidae) remains a destructive agricultural pest worldwide that continually overcomes conventional control methods. In recent years, RNA interference (RNAi) has emerged as an alternative for its management; however, although promising results have been reported, its effectiveness has been influenced by several factors, including the length of double-stranded RNA (dsRNA), the delivery method, stability, and especially the selection of the target gene. In this study, we designed and synthesized 290 bp dsRNAs targeting the SERCA and CPR genes from L. decemlineata, which encode the Sarco/Endoplasmic Reticulum Ca²⁺-ATPase and NADPH–Cytochrome P450 Reductase, respectively. Both dsRNAs successfully reduced transcript levels in larvae, with dsSERCA achieving ~60% knockdown by day 3 and dsCPR achieving ~50% knockdown by day 7. Furthermore, both treatments affected the larval growth and survival rate. However, while the dsCPR-treated larvae showed a 59% reduction in weight gain, the administration of dsSERCA had a strong phenotypic effect on the larvae, leading to decreased feeding, a 50.4% reduction in weight gain, and ultimately, 100% mortality. These results suggest that the SERCA and CPR genes could be promising targets for L. decemlineata control and emphasize the importance of appropriate target gene selection for RNAi silencing, as well as the need to explore and validate new genes for RNAi-mediated pest management. Full article

The intestinal epithelium, which is protected by mucosal surfaces composed of mucins and other glycoproteins, functions as a selective barrier that absorbs nutrients while preventing the translocation of harmful substances. To understand the mechanisms between mucosal disruption and tissue inflammation, we orally administrated a mucus-disrupting agent, dextran sodium sulfate, to Drosophila melanogaster and screened 63 differentially expressed genes (DEGs). Through a database search using bioinformatics tools (CHEA3 and WebGestalt), we identified ELK1 as a potential key transcription factor for the selected DEGs, and among the 63 DEGs, ELK1-related genes, B3GAT3, FIBP, and TENT2 (GlcAT-S, Fibp, and Wisp in Drosophila), were selected as the relevant genes that respond to mucus disruption. We confirmed that enterocyte (EC)-specific GlcAT-S knockdown by RNAi significantly reduced gut length and increased intestinal stem cell proliferation in Drosophila. Additionally, in EC-specific GlcAT-S-knockdown flies, it was observed that the mucus-production-related genes, Muc68D and Mur29B, were specifically reduced, whereas the inflammatory cytokines egr and upd3 were overexpressed. This study provides evidence that GlcAT-S is involved in the regulation of intestinal inflammation in Drosophila and plays a protective role against mucus disruption. Our findings suggest that GlcAT-S may be a potential therapeutic target for the treatment of intestinal inflammatory diseases such as IBD. Full article

The brown planthopper Nilaparvata lugens, a major rice pest, threatens global food security through rapid reproduction. This study investigates the role of the tramtrack (ttk) gene in male reproductive development and spermatogenesis using RNA interference (RNAi). Gene expression analysis revealed higher ttk levels in testes. RNAi-mediated knockdown of ttk in fourth-instar male nymphs reduced its expression by up to 80%, leading to severely impaired gonad development. Testes, vas deferens, and accessory glands in treated males exhibited 8–89% volume reductions compared to controls, accompanied by a 51–69% decline in sperm count and 60–85% reduction in sperm motility. Consequently, eggs fertilized by treated males showed a 73% decrease in hatching rates, with arrested embryonic development. These findings demonstrate ttk’s critical role in spermatogenesis and gonad maturation in N. lugens, highlighting its potential as an RNAi target for sustainable pest control strategies. Full article