Parasitoid wasps (Hymenoptera) play a crucial role in ecosystems and agroforestry pest management as biological control agents. These wasps utilize venom proteins to suppress host immunity and regulate physiology, facilitating offspring development. Although venom functions have been studied in some parasitoids, their roles in egg parasitoids remain poorly understood. In this study, we employed genomic and transcriptomic sequencing to identify venom proteins in Anastatus japonicus and Anastatus fulloi, two egg parasitoids used in biological control. We discovered a significant expansion of GH19 chitinase in their genomes, with phylogenetic analysis indicating acquisition via lateral gene transfer (LGT) from microsporidian. Functional characterization revealed that four highly expressed GH19 chitinases, Aj13071/Aj13072 in A. japonicus and Af23628/Af23629 in A. fulloi are essential for host egg penetration; silencing these genes increased penetration time and resulted in smaller or incomplete holes. Additionally, silencing Aj13071 and Aj13072 in A. japonicus impaired female fecundity, while Af23628 and Af23629 in A. fulloi affected venom reservoir development and egg load, respectively. These findings underscore the critical roles of GH19 chitinases in host penetration and reproduction, offering new insights into the molecular mechanisms driving parasitism in egg parasitoids. This study advances our understanding of venom evolution and supports the development of targeted biological control strategies.
{"title":"Laterally acquired chitinase genes in venom facilitate parasitism in egg parasitoid wasps","authors":"Xu Chen , Guy Smagghe , Yong-Ming Chen , Lian-Sheng Zang","doi":"10.1016/j.ibmb.2025.104362","DOIUrl":"10.1016/j.ibmb.2025.104362","url":null,"abstract":"<div><div>Parasitoid wasps (Hymenoptera) play a crucial role in ecosystems and agroforestry pest management as biological control agents. These wasps utilize venom proteins to suppress host immunity and regulate physiology, facilitating offspring development. Although venom functions have been studied in some parasitoids, their roles in egg parasitoids remain poorly understood. In this study, we employed genomic and transcriptomic sequencing to identify venom proteins in <em>Anastatus japonicus</em> and <em>Anastatus fulloi</em>, two egg parasitoids used in biological control. We discovered a significant expansion of GH19 chitinase in their genomes, with phylogenetic analysis indicating acquisition via lateral gene transfer (LGT) from microsporidian. Functional characterization revealed that four highly expressed GH19 chitinases, <em>Aj13071</em>/<em>Aj13072</em> in <em>A. japonicus</em> and <em>Af23628</em>/<em>Af23629</em> in <em>A. fulloi</em> are essential for host egg penetration; silencing these genes increased penetration time and resulted in smaller or incomplete holes. Additionally, silencing <em>Aj13071</em> and <em>Aj13072</em> in <em>A. japonicus</em> impaired female fecundity, while <em>Af23628</em> and <em>Af23629</em> in <em>A. fulloi</em> affected venom reservoir development and egg load, respectively. These findings underscore the critical roles of GH19 chitinases in host penetration and reproduction, offering new insights into the molecular mechanisms driving parasitism in egg parasitoids. This study advances our understanding of venom evolution and supports the development of targeted biological control strategies.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"183 ","pages":"Article 104362"},"PeriodicalIF":3.2,"publicationDate":"2025-07-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144648153","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-12DOI: 10.1016/j.ibmb.2025.104360
Yu-Chen Tsai , Yun-Chen Chung , Cheng-Chan Yang , Cheng-Hsien Lee , Che-Wei Liu , Shih-Ming Huang , Lan-Hsin Wang
Spatiotemporal gene expression is fundamental to cellular identity and function, ensuring proper development and tissue homeostasis. Histone modifications, such as H3K4 methylation (associated with active transcription) and H3K27 methylation (linked to repression), act as molecular switches that fine-tune gene expression. However, it remains largely unclear whether and how the histone modifying enzymes are regulated during normal development. In Drosophila, Utx, the sole H3K27 demethylase, plays a crucial role in removing di- and trimethylation marks on H3K27 across the genome. Here, we provide the first evidence that Utx transcription is dynamically regulated, with its regulatory elements exhibiting distinct temporal and spatial activity throughout development. Despite this variability at the transcriptional level, Utx protein is ubiquitously expressed and relatively stable. We found that the regulatory elements of Utx are highly active during embryogenesis but become largely inactivated in wing, eye and leg progenitor tissues during larval and pupal stages. Intriguingly, these regulatory elements are persistently active in the brain into adulthood. Disrupting this dynamic regulation activates a surveillance mechanism that limits excess Utx from translocating into the nucleus, thereby ensuring optimal nuclear protein levels. Moreover, while the Jumonji C (JmjC) demethylase activity of Utx is essential for Drosophila viability, we also discovered that the integrity of this domain is crucial for Utx protein expression. Our findings uncover a previously unrecognized aspect of Utx regulation, highlighting how precise control of its expression and localization safeguards developmental processes and maintains epigenetic stability.
时空基因表达是细胞身份和功能的基础，确保正常发育和组织稳态。组蛋白修饰，如H3K4甲基化（与主动转录相关）和H3K27甲基化（与抑制相关），作为微调基因表达的分子开关。然而，在正常发育过程中是否以及如何调节组蛋白修饰酶仍不清楚。在果蝇中，唯一的H3K27去甲基化酶Utx在去除整个基因组中H3K27的二甲基化和三甲基化标记中起着至关重要的作用。在这里，我们提供了第一个证据，证明Utx转录是动态调控的，其调控元件在整个发育过程中表现出不同的时间和空间活动。尽管在转录水平上存在这种可变性，但Utx蛋白普遍表达且相对稳定。我们发现Utx的调控元件在胚胎发生期间高度活跃，但在幼虫和蛹阶段的翅膀、眼睛和腿祖组织中大部分失活。有趣的是，这些调节因素在成年后仍在大脑中持续活跃。破坏这种动态调节激活了一种监视机制，该机制限制了过量的Utx转运到细胞核中，从而确保了最佳的核蛋白水平。此外，虽然Utx的Jumonji C （JmjC）去甲基酶活性对果蝇的生存至关重要，但我们也发现该结构域的完整性对Utx蛋白的表达至关重要。我们的研究结果揭示了Utx调控的一个以前未被认识到的方面，强调了其表达和定位的精确控制如何保护发育过程并维持表观遗传稳定性。
{"title":"Spatiotemporal dynamics of histone H3K27 demethylase during development","authors":"Yu-Chen Tsai , Yun-Chen Chung , Cheng-Chan Yang , Cheng-Hsien Lee , Che-Wei Liu , Shih-Ming Huang , Lan-Hsin Wang","doi":"10.1016/j.ibmb.2025.104360","DOIUrl":"10.1016/j.ibmb.2025.104360","url":null,"abstract":"<div><div>Spatiotemporal gene expression is fundamental to cellular identity and function, ensuring proper development and tissue homeostasis. Histone modifications, such as H3K4 methylation (associated with active transcription) and H3K27 methylation (linked to repression), act as molecular switches that fine-tune gene expression. However, it remains largely unclear whether and how the histone modifying enzymes are regulated during normal development. In <em>Drosophila</em>, Utx, the sole H3K27 demethylase, plays a crucial role in removing di- and trimethylation marks on H3K27 across the genome. Here, we provide the first evidence that <em>Utx</em> transcription is dynamically regulated, with its regulatory elements exhibiting distinct temporal and spatial activity throughout development. Despite this variability at the transcriptional level, Utx protein is ubiquitously expressed and relatively stable. We found that the regulatory elements of <em>Utx</em> are highly active during embryogenesis but become largely inactivated in wing, eye and leg progenitor tissues during larval and pupal stages. Intriguingly, these regulatory elements are persistently active in the brain into adulthood. Disrupting this dynamic regulation activates a surveillance mechanism that limits excess Utx from translocating into the nucleus, thereby ensuring optimal nuclear protein levels. Moreover, while the Jumonji C (JmjC) demethylase activity of Utx is essential for <em>Drosophila</em> viability, we also discovered that the integrity of this domain is crucial for Utx protein expression. Our findings uncover a previously unrecognized aspect of <em>Utx</em> regulation, highlighting how precise control of its expression and localization safeguards developmental processes and maintains epigenetic stability.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"183 ","pages":"Article 104360"},"PeriodicalIF":3.2,"publicationDate":"2025-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144635826","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-09DOI: 10.1016/j.ibmb.2025.104359
Longhao Tang , Xiangyi Wei , Liying Zhang , Zhidan Peng , Zulian Liu , Yongping Huang , Pinhua Rao , Lili Yan , Dehong Yang
Lepidoptera, a major threat to global agriculture, possesses robust reproductive strategies, making understanding reproduction crucial for developing sterile insect techniques (SIT). The silkworm (Bombyx mori), a model lepidopteran, exhibits polytrophic meroistic oogenesis involving trophocyte-mediated nutrient transport and vitellogenin receptor (VgR)-dependent yolk protein uptake. However, epigenetic regulation of these processes remains unclear. This study investigates the role of BmLid, a histone demethylase, in silkworm oogenesis. Using CRISPR/Cas9, we generated BmLid knockout mutants (ΔBmLid) and confirmed gene disruption via sequencing and western blotting. ΔBmLid females exhibited developmental defects and complete sterility, despite unaffected mating behavior or sperm migration. The H3K4me2/me3 and H3K9me2/me3 levels were significantly increased in ΔBmLid ovaries, indicating BmLid has broad demethylase activity in vivo, too. Transcriptomic analysis showed significant downregulation of VgR, critical for vitellogenin transport, alongside dysregulation of cell junction and apoptosis pathways. These defects disrupted follicular epithelium integrity and yolk protein deposition, mirroring similar phenotypes observed in VgR mutants. Our findings demonstrate that BmLid regulates oogenesis by modulating histone methylation to control VgR expression, cell adhesion, and cell apoptosis, thereby ensuring vitellogenesis. This study highlights epigenetic mechanisms underlying lepidopteran reproduction and identifies BmLid as a potential target for SIT-based pest management strategies.
{"title":"BmLid has histone lysine demethylase activity and regulates oogenesis in the silkworm, Bombyx mori","authors":"Longhao Tang , Xiangyi Wei , Liying Zhang , Zhidan Peng , Zulian Liu , Yongping Huang , Pinhua Rao , Lili Yan , Dehong Yang","doi":"10.1016/j.ibmb.2025.104359","DOIUrl":"10.1016/j.ibmb.2025.104359","url":null,"abstract":"<div><div>Lepidoptera, a major threat to global agriculture, possesses robust reproductive strategies, making understanding reproduction crucial for developing sterile insect techniques (SIT). The silkworm (<em>Bombyx mori</em>), a model lepidopteran, exhibits polytrophic meroistic oogenesis involving trophocyte-mediated nutrient transport and vitellogenin receptor (VgR)-dependent yolk protein uptake. However, epigenetic regulation of these processes remains unclear. This study investigates the role of <em>BmLid</em>, a histone demethylase, in silkworm oogenesis. Using CRISPR/Cas9, we generated <em>BmLid</em> knockout mutants (Δ<em>BmLid</em>) and confirmed gene disruption via sequencing and western blotting. Δ<em>BmLid</em> females exhibited developmental defects and complete sterility, despite unaffected mating behavior or sperm migration. The H3K4me2/me3 and H3K9me2/me3 levels were significantly increased in Δ<em>BmLid</em> ovaries, indicating <em>BmLid</em> has broad demethylase activity <em>in vivo,</em> too. Transcriptomic analysis showed significant downregulation of <em>VgR</em>, critical for vitellogenin transport, alongside dysregulation of cell junction and apoptosis pathways. These defects disrupted follicular epithelium integrity and yolk protein deposition, mirroring similar phenotypes observed in <em>VgR</em> mutants. Our findings demonstrate that <em>BmLid</em> regulates oogenesis by modulating histone methylation to control <em>VgR</em> expression, cell adhesion, and cell apoptosis, thereby ensuring vitellogenesis. This study highlights epigenetic mechanisms underlying lepidopteran reproduction and identifies <em>BmLid</em> as a potential target for SIT-based pest management strategies.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"183 ","pages":"Article 104359"},"PeriodicalIF":3.2,"publicationDate":"2025-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144615628","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-04DOI: 10.1016/j.ibmb.2025.104356
Yatao Zhu , Suijie Kuang , Zhuotian Yao , Yingjuan Sun , HongShuai Gao , Qiao Gao , Wenbing Ding , HuaLiang He , Youzhi Li , Lin Qiu
The doublesex (dsx) gene has a conserved role in sex determination in insects, controlling sexual development and mating behavior. Although dsx is known to participate in these critical functions, its role in insect sex determination remains not fully elucidated. Given the economic importance of the rice pest Chilo suppressalis, we employed this species as a model to investigate the function of its dsx homolog. We cloned and characterized the Csdsx gene, which is 1123 bp in length and encodes four sex-specific proteins: three female-specific isoforms of 252, 258, and 254 amino acids, respectively, and one male-specific isoform of 290 amino acids. Phylogenetic analysis revealed that Csdsx is highly conserved within Lepidoptera, containing two domains: DM DNA binding domain and DSX dimer domain. Analysis of transcripts produced from a mini-dsx construct transfected into human HEK293T cells indicates that the female-splicing pattern is the default mode. Quantitative real-time PCR showed that among various developmental stages, Csdsx expression peaked at the first instar larval stage and showed tissue-specific, stage-dependent patterns, with notably high levels in the larval midgut, pupal fat body, and adult thorax in both sexes. In comparison to wild-type (WT) adults, Csdsx-knockout individuals exhibited malformations in their external genitalia, and female wing patterns became masculinized. Dissections revealed that knockout females had a reduced number of mature oocytes, while knockout males displayed a smaller testis area. Furthermore, when Csdsx-knockout females were paired with wild-type males, their mating behavior was significantly impaired. RNA-seq revealed that Csdsx disruption led to sex-biased gene expression shifts, including upregulation of male-associated genes (e.g., OBPs, PBPs, trypsin) and downregulation of female-specific genes (vitellogenin, FAS), indicating partial masculinization at the transcriptional level. These findings underscore the critical role of the dsx gene in reproductive development and sexual dimorphism in C. suppressalis.
{"title":"Doublesex knockout via CRISPR/Cas9 disrupts fertility and sexual dimorphism of wings in the rice stem borer, Chilo suppressalis","authors":"Yatao Zhu , Suijie Kuang , Zhuotian Yao , Yingjuan Sun , HongShuai Gao , Qiao Gao , Wenbing Ding , HuaLiang He , Youzhi Li , Lin Qiu","doi":"10.1016/j.ibmb.2025.104356","DOIUrl":"10.1016/j.ibmb.2025.104356","url":null,"abstract":"<div><div>The <em>doublesex</em> (<em>dsx</em>) gene has a conserved role in sex determination in insects, controlling sexual development and mating behavior. Although <em>dsx</em> is known to participate in these critical functions, its role in insect sex determination remains not fully elucidated. Given the economic importance of the rice pest <em>Chilo suppressalis</em>, we employed this species as a model to investigate the function of its <em>dsx</em> homolog. We cloned and characterized the <em>Csdsx</em> gene, which is 1123 bp in length and encodes four sex-specific proteins: three female-specific isoforms of 252, 258, and 254 amino acids, respectively, and one male-specific isoform of 290 amino acids. Phylogenetic analysis revealed that <em>Csdsx</em> is highly conserved within Lepidoptera, containing two domains: DM DNA binding domain and DSX dimer domain. Analysis of transcripts produced from a mini-<em>dsx</em> construct transfected into human HEK293T cells indicates that the female-splicing pattern is the default mode. Quantitative real-time PCR showed that among various developmental stages, <em>Csdsx</em> expression peaked at the first instar larval stage and showed tissue-specific, stage-dependent patterns, with notably high levels in the larval midgut, pupal fat body, and adult thorax in both sexes. In comparison to wild-type (WT) adults, <em>Csdsx</em>-knockout individuals exhibited malformations in their external genitalia, and female wing patterns became masculinized. Dissections revealed that knockout females had a reduced number of mature oocytes, while knockout males displayed a smaller testis area. Furthermore, when <em>Csdsx</em>-knockout females were paired with wild-type males, their mating behavior was significantly impaired. RNA-seq revealed that <em>Csdsx</em> disruption led to sex-biased gene expression shifts, including upregulation of male-associated genes (e.g., <em>OBPs</em>, <em>PBPs</em>, <em>trypsin</em>) and downregulation of female-specific genes (<em>vitellogenin, FAS</em>), indicating partial masculinization at the transcriptional level. These findings underscore the critical role of the <em>dsx</em> gene in reproductive development and sexual dimorphism in <em>C. suppressalis</em>.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104356"},"PeriodicalIF":3.2,"publicationDate":"2025-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144563084","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-04DOI: 10.1016/j.ibmb.2025.104358
Jianming Zhang , Ting Tang , Dongdong Lin , Liya Ma , Fengsong Liu
Hemocytes represent a pivotal element of insect defense against pathogens through the mechanisms of cellular and humoral immunity. However, hemocyte types and functions exhibit variation across different insect taxa, leading to the lack of a standardized classification system for insect hemocytes. To gain insight into the immune mechanisms of the house fly Musca domestica, we used a combination of morphological observations and single-cell RNA sequencing techniques (scRNA-seq) to taxonomically characterize house fly larval hemocytes and analyze their immune function. As a result, five different types of hemocytes were identified from house fly larvae, granulocytes (GR), spherulocytes (SP), plasmatocytes (PL), prohemocytes (PR), and oenocytoids (OE). On the basis of microscopic observations, flow cytometry analysis and gene expression profiles, the immune functions of house fly hemocytes were hypothesized to be as follows: the GRs are responsible for phagocytosis, the SPs are highly expressive of lectins, and the PLs are highly expressive of antimicrobial peptides (AMPs) and are involved in nodule formation, the PRs act as progenitor cells and retain the differentiation potential of stem cells, and the OEs are involved in melanization reactions mainly through the expression of phenoloxidase (PO). Based on the scRNA-seq data, the marker genes for each type of hemocyte were identified. The present study unveils the heterogeneity of house fly hemocytes in terms of morphology, gene expression characteristics, and function, thereby establishing the foundation for an in-depth understanding of the immune mechanisms in house fly.
{"title":"Identification of hemocyte types and characterization of their immune function in the house fly based on morphological observation and single-cell RNA sequencing","authors":"Jianming Zhang , Ting Tang , Dongdong Lin , Liya Ma , Fengsong Liu","doi":"10.1016/j.ibmb.2025.104358","DOIUrl":"10.1016/j.ibmb.2025.104358","url":null,"abstract":"<div><div>Hemocytes represent a pivotal element of insect defense against pathogens through the mechanisms of cellular and humoral immunity. However, hemocyte types and functions exhibit variation across different insect taxa, leading to the lack of a standardized classification system for insect hemocytes. To gain insight into the immune mechanisms of the house fly <em>Musca domestica</em>, we used a combination of morphological observations and single-cell RNA sequencing techniques (scRNA-seq) to taxonomically characterize house fly larval hemocytes and analyze their immune function. As a result, five different types of hemocytes were identified from house fly larvae, granulocytes (GR), spherulocytes (SP), plasmatocytes (PL), prohemocytes (PR), and oenocytoids (OE). On the basis of microscopic observations, flow cytometry analysis and gene expression profiles, the immune functions of house fly hemocytes were hypothesized to be as follows: the GRs are responsible for phagocytosis, the SPs are highly expressive of lectins, and the PLs are highly expressive of antimicrobial peptides (AMPs) and are involved in nodule formation, the PRs act as progenitor cells and retain the differentiation potential of stem cells, and the OEs are involved in melanization reactions mainly through the expression of phenoloxidase (PO). Based on the scRNA-seq data, the marker genes for each type of hemocyte were identified. The present study unveils the heterogeneity of house fly hemocytes in terms of morphology, gene expression characteristics, and function, thereby establishing the foundation for an in-depth understanding of the immune mechanisms in house fly.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104358"},"PeriodicalIF":3.2,"publicationDate":"2025-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144566874","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-04DOI: 10.1016/j.ibmb.2025.104357
Shuai Wang , Yuan Xie , Chong Liu , Jin-Wei Liu , Fang-Rui Guo , Song-Tao Qiao , Yu-Ji Xing , Shun-Fan Wu , Chris Bass , Cong-Fen Gao
Abamectin serves as a key alternative for managing diamide-resistant Chilo suppressalis. However, resistance to abamectin itself presents a significant challenge to this pest control, and its genetic basis remains poorly understood. In this study, we report an increase in field resistance to abamectin in populations of C. suppressalis and provide evidence for a cytochrome P450-mediated metabolic resistance mechanism, supported by enzyme activity assays and synergist bioassays. Transcriptomic profiling analysis revealed six candidate P450 genes, among which the duplicated gene CYP6SN2 showed a strong correlation with resistance. Variant CYP6SN2vB was highly expressed in detoxification tissues and mainly produced isoforms with a short 3′-UTR. Functional validation in transgenic Drosophila melanogaster confirmed that overexpression of CYP6SN2vB conferred 2.4- and 1.9-fold resistance to abamectin and emamectin benzoate, whereas CYP6SN2vA had no detectable effect. Molecular docking further revealed that CYP6SN2vB could potentially hydroxylate abamectin via interactions near oxygen-binding motifs. In contrast, the inward-folded substrate recognition site 1 region of CYP6SN2vA compresses the ligand-binding cavity, precluding abamectin interaction. These findings suggest neofunctionalization of CYP6SN2vB following duplication and provide valuable insights into the molecular basis of abamectin resistance in C. suppressalis, with implications for resistance management.
{"title":"Adaptive neofunctionalization of ancestral CYP6SN2vB following duplication contributes to abamectin resistance in Chilo suppressalis Walker","authors":"Shuai Wang , Yuan Xie , Chong Liu , Jin-Wei Liu , Fang-Rui Guo , Song-Tao Qiao , Yu-Ji Xing , Shun-Fan Wu , Chris Bass , Cong-Fen Gao","doi":"10.1016/j.ibmb.2025.104357","DOIUrl":"10.1016/j.ibmb.2025.104357","url":null,"abstract":"<div><div>Abamectin serves as a key alternative for managing diamide-resistant <em>Chilo suppressalis</em>. However, resistance to abamectin itself presents a significant challenge to this pest control, and its genetic basis remains poorly understood. In this study, we report an increase in field resistance to abamectin in populations of <em>C. suppressalis</em> and provide evidence for a cytochrome P450-mediated metabolic resistance mechanism, supported by enzyme activity assays and synergist bioassays. Transcriptomic profiling analysis revealed six candidate P450 genes, among which the duplicated gene <em>CYP6SN2</em> showed a strong correlation with resistance. Variant <em>CYP6SN2vB</em> was highly expressed in detoxification tissues and mainly produced isoforms with a short 3′-UTR. Functional validation in transgenic <em>Drosophila melanogaster</em> confirmed that overexpression of <em>CYP6SN2vB</em> conferred 2.4- and 1.9-fold resistance to abamectin and emamectin benzoate, whereas <em>CYP6SN2vA</em> had no detectable effect. Molecular docking further revealed that CYP6SN2vB could potentially hydroxylate abamectin via interactions near oxygen-binding motifs. In contrast, the inward-folded substrate recognition site 1 region of CYP6SN2vA compresses the ligand-binding cavity, precluding abamectin interaction. These findings suggest neofunctionalization of CYP6SN2vB following duplication and provide valuable insights into the molecular basis of abamectin resistance in <em>C. suppressalis</em>, with implications for resistance management.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104357"},"PeriodicalIF":3.2,"publicationDate":"2025-07-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144566873","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-06-26DOI: 10.1016/j.ibmb.2025.104355
Tianxiang Xiao , Xiaodan Huang , Menqing Deng , Yingjie Jiang , Wenxiu Wang , Xiyue Xu , Jun Li , Xinyu Zhao , Bo Pan , Ziyu He , Zhiming Yang , Zhongxiang Sun , Kai Lu
Glutathione S-transferases (GSTs) play pivotal roles in insect metabolic adaptation to xenobiotic challenges; however, the mechanistic basis of GST-mediated insecticide detoxification, particularly the interaction between GSTs and pyrethroids remains poorly characterized. This study demonstrates that exposure to three pyrethroids (β-cypermethrin, λ-cyhalothrin, and fenvalerate) induces significant elevation in GST activity in the tobacco cutworm Spodoptera litura. The synergistic effects of the GST-specific inhibitor diethyl maleate dramatically potentiated pyrethroid toxicity, indicating a critical role of GST-mediated detoxification. Transcriptional profiling revealed selective induction of GSTe11 and GSTe16 under pyrethroid challenge, with RNA interference-mediated GSTe16 knockdown substantially increasing larval susceptibility. In vivo validation through CRISPR/Cas9 mutagenesis and transgenic Drosophila melanogaster models established GSTe16 as a critical determinant of pyrethroid detoxification. In vitro analyses uncovered the bifunctional capacity of GSTe16: direct metabolic processing of pyrethroids via conjugation and secondary antioxidant defense through reactive oxygen species neutralization. Molecular docking and site-directed mutagenesis identified Arg111 and Asn122 as substrate-specificity determinants in the binding and catalytic subsites, with catalytic mutants retaining full antioxidant activity. This functional specialization reflects evolutionary adaptation of GST architecture, coordinating xenobiotic metabolism with oxidative stress responses. Collectively, these results establish an evolutionary-driven functional compartmentalization within GST architecture, proposing a dual-defense model that synergizes pyrethroid metabolism with oxidative stress resilience.
{"title":"Unveiling the functional contribution of GSTe16 to pyrethroid detoxification in Spodoptera litura","authors":"Tianxiang Xiao , Xiaodan Huang , Menqing Deng , Yingjie Jiang , Wenxiu Wang , Xiyue Xu , Jun Li , Xinyu Zhao , Bo Pan , Ziyu He , Zhiming Yang , Zhongxiang Sun , Kai Lu","doi":"10.1016/j.ibmb.2025.104355","DOIUrl":"10.1016/j.ibmb.2025.104355","url":null,"abstract":"<div><div>Glutathione <em>S</em>-transferases (GSTs) play pivotal roles in insect metabolic adaptation to xenobiotic challenges; however, the mechanistic basis of GST-mediated insecticide detoxification, particularly the interaction between GSTs and pyrethroids remains poorly characterized. This study demonstrates that exposure to three pyrethroids (<em>β</em>-cypermethrin, <em>λ</em>-cyhalothrin, and fenvalerate) induces significant elevation in GST activity in the tobacco cutworm <em>Spodoptera litura</em>. The synergistic effects of the GST-specific inhibitor diethyl maleate dramatically potentiated pyrethroid toxicity, indicating a critical role of GST-mediated detoxification. Transcriptional profiling revealed selective induction of <em>GSTe11</em> and <em>GSTe16</em> under pyrethroid challenge, with RNA interference-mediated <em>GSTe16</em> knockdown substantially increasing larval susceptibility. <em>In vivo</em> validation through CRISPR/Cas9 mutagenesis and transgenic <em>Drosophila melanogaster</em> models established GSTe16 as a critical determinant of pyrethroid detoxification. <em>In vitro</em> analyses uncovered the bifunctional capacity of GSTe16: direct metabolic processing of pyrethroids via conjugation and secondary antioxidant defense through reactive oxygen species neutralization. Molecular docking and site-directed mutagenesis identified Arg111 and Asn122 as substrate-specificity determinants in the binding and catalytic subsites, with catalytic mutants retaining full antioxidant activity. This functional specialization reflects evolutionary adaptation of GST architecture, coordinating xenobiotic metabolism with oxidative stress responses. Collectively, these results establish an evolutionary-driven functional compartmentalization within GST architecture, proposing a dual-defense model that synergizes pyrethroid metabolism with oxidative stress resilience.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104355"},"PeriodicalIF":3.2,"publicationDate":"2025-06-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144501663","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-06-25DOI: 10.1016/j.ibmb.2025.104349
Ludwig Dersch , Ole Bohlken , Jonas Krämer , Lennart Schulte , Kornelia Hardes , Michael Marner , Till F. Schäberle , Andreas Vilcinskas , Tim Lüddecke
Venoms are complex cocktails containing novel biomolecules. Many venomous animal groups are understudied, including Horseflies of the family Tabanidae. Their larvae utilize venom to overpower prey and to defend against predators. Recent studies uncovered short linear peptides in their venoms but their bioactivity spectrum and biological function remain unknown. Here, we employ a bioactivity profiling of eight synthesized toxins of the large marsh horsefly (Tabanus autumnalis) larval venom via in silico, in vitro, and in vivo experiments. Bioinformatic analysis suggest, that the selected peptides fold into cationic and amphipathic alpha-helices. Assays against microbes reveal antimicrobial activity potentially via membrane interaction caused by some of the linear Tabanus toxins, while tests on vertebrate cells reveal cytotoxic and hemolytic effects. Lastly, injections of the peptides in Drosophila melanogaster flies recovered insecticidal and paralytic activity. Based on our analysis, we propose that some of the linear T. autumnalis peptides facilitate prey capture via their rapid paralytic activity in insects. A second line of functional utilization may be present in defense against predators and microbial colonization. From a translational perspective, it appears unlikely that the tested peptides could efficiently be translated into anti-infectives, while an evaluation for agricultural exploitation demands further research. Our study provides functional insights into the toxin repertoire of one of the least studied venom systems on earth and serves as starting point to further investigate dipteran venom biology.
{"title":"Linear peptides from the venom of large marsh horsefly (Tabanus autumnalis) larvae facilitate prey capture and defense","authors":"Ludwig Dersch , Ole Bohlken , Jonas Krämer , Lennart Schulte , Kornelia Hardes , Michael Marner , Till F. Schäberle , Andreas Vilcinskas , Tim Lüddecke","doi":"10.1016/j.ibmb.2025.104349","DOIUrl":"10.1016/j.ibmb.2025.104349","url":null,"abstract":"<div><div>Venoms are complex cocktails containing novel biomolecules. Many venomous animal groups are understudied, including Horseflies of the family <em>Tabanidae</em>. Their larvae utilize venom to overpower prey and to defend against predators. Recent studies uncovered short linear peptides in their venoms but their bioactivity spectrum and biological function remain unknown. Here, we employ a bioactivity profiling of eight synthesized toxins of the large marsh horsefly (<em>Tabanus autumnalis</em>) larval venom via <em>in silico</em>, <em>in vitro</em>, and <em>in vivo</em> experiments. Bioinformatic analysis suggest, that the selected peptides fold into cationic and amphipathic alpha-helices. Assays against microbes reveal antimicrobial activity potentially via membrane interaction caused by some of the linear <em>Tabanus</em> toxins, while tests on vertebrate cells reveal cytotoxic and hemolytic effects. Lastly, injections of the peptides in <em>Drosophila melanogaster</em> flies recovered insecticidal and paralytic activity. Based on our analysis, we propose that some of the linear <em>T. autumnalis</em> peptides facilitate prey capture via their rapid paralytic activity in insects. A second line of functional utilization may be present in defense against predators and microbial colonization. From a translational perspective, it appears unlikely that the tested peptides could efficiently be translated into anti-infectives, while an evaluation for agricultural exploitation demands further research. Our study provides functional insights into the toxin repertoire of one of the least studied venom systems on earth and serves as starting point to further investigate dipteran venom biology.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104349"},"PeriodicalIF":3.2,"publicationDate":"2025-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144511309","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-06-25DOI: 10.1016/j.ibmb.2025.104354
Dongbiao Lü, Dandan Pan, Donghan Wang, Jiacheng Ye, Xiaofang Sha, Xiaoyan Zhang, Yanshen Fu, Xiangqun Yuan, Yiping Li
Grapholita molesta is an important pest of fruit trees worldwide. Its larvae feed on a wide range of hosts, including fruits, as well as tender shoots, causing significant damage and making effective control challenging. Trypsin is an important digestive enzyme in the midgut of insects, playing a crucial role in digestion, growth, and development. We identified and cloned the trypsin genes, GmolTrypsin1 and GmolTrypsin10, both of which showed the highest expression levels in 4th instar larvae and the midgut. We utilized RNA interference to silence GmolTrypsin1 and GmolTrypsin10 in G. molesta larvae feeding on artificial diets, peaches, pears, plums, apples, and peach shoots. The expression level of the target genes decreased by 64–73 %, and western blot analysis confirmed a significant reduction in protein expression in the treatment group.
Silencing GmolTrypsin1 led to significant changes in the relative activity of trypsin at the larval stage, in pupal weight, and in the emergence rate of G. molesta larvae fed on peach shoots. Silencing GmolTrypsin10 caused significant effects in larvae fed peach, apple, pear, and plum. We clarified the expression sites of GmolTrypsin1 and GmolTrypsin10, and their proteins in the midgut of G. molesta larvae and Sf9 cells. Our study demonstrated the roles of GmolTrypsin1 and GmolTrypsin10 in regulating G molesta larval responses to different host plants and clarified their localization in the larval midgut, providing an important theoretical basis for the development of pest control strategies targeting midgut digestive enzymes.
{"title":"Study on the functional role and tissue localization of GmolTrypsin in larvae of Grapholita molesta feeding on different host plants","authors":"Dongbiao Lü, Dandan Pan, Donghan Wang, Jiacheng Ye, Xiaofang Sha, Xiaoyan Zhang, Yanshen Fu, Xiangqun Yuan, Yiping Li","doi":"10.1016/j.ibmb.2025.104354","DOIUrl":"10.1016/j.ibmb.2025.104354","url":null,"abstract":"<div><div><em>Grapholita molesta</em> is an important pest of fruit trees worldwide. Its larvae feed on a wide range of hosts, including fruits, as well as tender shoots, causing significant damage and making effective control challenging. Trypsin is an important digestive enzyme in the midgut of insects, playing a crucial role in digestion, growth, and development. We identified and cloned the trypsin genes, <em>GmolTrypsin1</em> and <em>GmolTrypsin10</em>, both of which showed the highest expression levels in 4th instar larvae and the midgut. We utilized RNA interference to silence <em>GmolTrypsin1</em> and <em>GmolTrypsin10</em> in <em>G</em>. <em>molesta</em> larvae feeding on artificial diets, peaches, pears, plums, apples, and peach shoots. The expression level of the target genes decreased by 64–73 %, and western blot analysis confirmed a significant reduction in protein expression in the treatment group.</div><div>Silencing <em>GmolTrypsin1</em> led to significant changes in the relative activity of trypsin at the larval stage, in pupal weight, and in the emergence rate of <em>G. molesta</em> larvae fed on peach shoots. Silencing <em>GmolTrypsin10</em> caused significant effects in larvae fed peach, apple, pear, and plum. We clarified the expression sites of <em>GmolTrypsin1</em> and <em>GmolTrypsin10</em>, and their proteins in the midgut of <em>G. molesta</em> larvae and Sf9 cells. Our study demonstrated the roles of <em>GmolTrypsin1</em> and <em>GmolTrypsin10</em> in regulating <em>G molesta</em> larval responses to different host plants and clarified their localization in the larval midgut, providing an important theoretical basis for the development of pest control strategies targeting midgut digestive enzymes.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104354"},"PeriodicalIF":3.2,"publicationDate":"2025-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144501662","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Chromosome ends of most eukaryotes are composed of simple telomeric repeats. For arthropods, TTAGG pentanucleotide repeats, (TTAGG)n has been considered as the ancestral telomeric repeat. However, in the order Odonata, the earliest diverged group in insects that contains dragonflies and damselflies, (TTAGG)n signal has been almost undetectable in most examined species. Here, we report the pentanucleotide repeat (TTGGG)n as the typical telomeric repeat sequence for Odonata (dragonflies and damselflies). Based on genomic information from ten Odonata species, (TTGGG)n was considered the most likely candidate for telomeric repeat sequences. By fluorescence in situ hybridization (FISH) using 12 Odonata species, clear (TTGGG)n signals were detected at the chromosome ends in both dragonflies and damselflies. By Southern hybridization using 63 Odonata species, strong (TTGGG)n signals were detected from the majority of species covering all three suborders of Odonata, indicating that the telomeric repeat of the common ancestor of extant Odonata is (TTGGG)n. Notably, there were a few Odonata species in which (TTGGG)n signals were faint or absent, suggesting that the telomeric repeat sequence has repeatedly diverged in Odonata, even within genera such as Sympetrum. We also identified telomerase genes in both dragonflies and damselflies, and in some species, more than two telomerase genes are suggested to be present. Overall, this study demonstrates the ancestral acquisition of novel telomeric repeats and their repeated alteration in Odonata.
{"title":"Acquisition and repeated alteration of (TTGGG)n telomeric repeats in Odonata (dragonflies and damselflies)","authors":"Tatsuhiro Gotoh , Haruka Suzuki , Minoru Moriyama , Ryo Futahashi , Mizuko Osanai-Futahashi","doi":"10.1016/j.ibmb.2025.104353","DOIUrl":"10.1016/j.ibmb.2025.104353","url":null,"abstract":"<div><div>Chromosome ends of most eukaryotes are composed of simple telomeric repeats. For arthropods, TTAGG pentanucleotide repeats, (TTAGG)<sub>n</sub> has been considered as the ancestral telomeric repeat. However, in the order Odonata, the earliest diverged group in insects that contains dragonflies and damselflies, (TTAGG)<sub>n</sub> signal has been almost undetectable in most examined species. Here, we report the pentanucleotide repeat (TTGGG)<sub>n</sub> as the typical telomeric repeat sequence for Odonata (dragonflies and damselflies). Based on genomic information from ten Odonata species, (TTGGG)<sub>n</sub> was considered the most likely candidate for telomeric repeat sequences. By fluorescence <em>in situ</em> hybridization (FISH) using 12 Odonata species, clear (TTGGG)<sub>n</sub> signals were detected at the chromosome ends in both dragonflies and damselflies. By Southern hybridization using 63 Odonata species, strong (TTGGG)<sub>n</sub> signals were detected from the majority of species covering all three suborders of Odonata, indicating that the telomeric repeat of the common ancestor of extant Odonata is (TTGGG)<sub>n</sub>. Notably, there were a few Odonata species in which (TTGGG)<sub>n</sub> signals were faint or absent, suggesting that the telomeric repeat sequence has repeatedly diverged in Odonata, even within genera such as <em>Sympetrum</em>. We also identified telomerase genes in both dragonflies and damselflies, and in some species, more than two telomerase genes are suggested to be present. Overall, this study demonstrates the ancestral acquisition of novel telomeric repeats and their repeated alteration in Odonata.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"182 ","pages":"Article 104353"},"PeriodicalIF":3.2,"publicationDate":"2025-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144504344","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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