Pub Date : 2025-04-05DOI: 10.1016/j.ibmb.2025.104310
Qian Wang, Xin Chen, Yu-Feng Wang
sec61β encodes a subunit of the Sec61 translocon which is a highly conserved heterotrimer responsible for translocating the nascent polypeptides into the lumen of the endoplasmic reticulum (ER) or onto the ER membrane. In this study, we show that knockdown of sec61β in the early germline leads to male sterility in Drosophila melanogaster. These males exhibit testes that are dramatically reduced in size and devoid of germ cells. However, the somatic cells with hub markers extend abnormally beyond the stem cell niche region. Stat92E-positive cells are also expanded into the posterior region of the small testes and primarily in the nuclei. Through tracking the developmental processes of germ cells, we find that the loss of germ cells occurs during the 3rd instar larval stage. Additionally, studies in Drosophila S2 cells reveal that Sec61β can directly interact with Ocnus (Ocn), likely at the nuclear membrane. Genetically, we show that overexpression of ocn partially restores fertility in sec61β knockdown males, while overexpression of sec61β fails to compensate for the defects in male fertility induced by ocn knockdown. These findings suggest that Sec61β might play a critical role in testis development and spermatogenesis, potentially coordinating with Ocn and involving in the JAK/STAT pathway.
{"title":"Sec61β, a subunit of the Sec61 complex at the endoplasmic reticulum, coordinates with Ocnus in regulating Drosophila spermatogenesis","authors":"Qian Wang, Xin Chen, Yu-Feng Wang","doi":"10.1016/j.ibmb.2025.104310","DOIUrl":"10.1016/j.ibmb.2025.104310","url":null,"abstract":"<div><div><em>sec61β</em> encodes a subunit of the Sec61 translocon which is a highly conserved heterotrimer responsible for translocating the nascent polypeptides into the lumen of the endoplasmic reticulum (ER) or onto the ER membrane. In this study, we show that knockdown of <em>sec61β</em> in the early germline leads to male sterility in <em>Drosophila melanogaster.</em> These males exhibit testes that are dramatically reduced in size and devoid of germ cells. However, the somatic cells with hub markers extend abnormally beyond the stem cell niche region. Stat92E-positive cells are also expanded into the posterior region of the small testes and primarily in the nuclei. Through tracking the developmental processes of germ cells, we find that the loss of germ cells occurs during the 3rd instar larval stage. Additionally, studies in <em>Drosophila</em> S2 cells reveal that Sec61β can directly interact with Ocnus (Ocn), likely at the nuclear membrane. Genetically, we show that overexpression of <em>ocn</em> partially restores fertility in <em>sec61β</em> knockdown males, while overexpression of <em>sec61β</em> fails to compensate for the defects in male fertility induced by <em>ocn</em> knockdown. These findings suggest that <em>Sec61β</em> might play a critical role in testis development and spermatogenesis, potentially coordinating with Ocn and involving in the JAK/STAT pathway.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104310"},"PeriodicalIF":3.2,"publicationDate":"2025-04-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143799807","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 : 2025-03-31DOI: 10.1016/j.ibmb.2025.104309
Katherine Nevard , Rajdeep Kaur , Tim Harvey-Samuel
Culex quinquefasciatus Say is a mosquito which acts as a vector for numerous diseases including West Nile virus, lymphatic filariasis and avian malaria, over a broad geographical range. As the effectiveness of insecticidal mosquito control methods declines, the need has grown to develop genetic control methods to curb the spread of disease. The piggyBac transposon system - the most widely used genetic transformation tool in insects, including mosquitoes - generates quasi-random insertions of donor DNA into the host genome. However, despite the broad reported species range of piggyBac, previous attempts to use this tool to transform Culex quinquefasciatus mosquitoes have failed. Here we report the first successful transformation of Culex quinquefasciatus with the piggyBac transposon system. Using commercially synthesised piggyBac mRNA as a transposase source, we were able to generate three independent insertions of a ZsGreen fluorescent marker gene, with transformation efficiencies of up to 5 %. Through this work, we have expanded the genetic toolkit available for the genetic manipulation of Culex mosquitoes and thus removed a barrier to developing novel genetic control methods in this important disease vector.
{"title":"Germline transformation of the West Nile virus and avian malaria vector Culex quinquefasciatus Say using the piggyBac transposon system","authors":"Katherine Nevard , Rajdeep Kaur , Tim Harvey-Samuel","doi":"10.1016/j.ibmb.2025.104309","DOIUrl":"10.1016/j.ibmb.2025.104309","url":null,"abstract":"<div><div><em>Culex quinquefasciatus</em> Say is a mosquito which acts as a vector for numerous diseases including West Nile virus, lymphatic filariasis and avian malaria, over a broad geographical range. As the effectiveness of insecticidal mosquito control methods declines, the need has grown to develop genetic control methods to curb the spread of disease. The piggyBac transposon system - the most widely used genetic transformation tool in insects, including mosquitoes - generates quasi-random insertions of donor DNA into the host genome. However, despite the broad reported species range of piggyBac, previous attempts to use this tool to transform <em>Culex quinquefasciatus</em> mosquitoes have failed. Here we report the first successful transformation of <em>Culex quinquefasciatus</em> with the piggyBac transposon system. Using commercially synthesised piggyBac mRNA as a transposase source, we were able to generate three independent insertions of a <em>ZsGreen</em> fluorescent marker gene, with transformation efficiencies of up to 5 %. Through this work, we have expanded the genetic toolkit available for the genetic manipulation of <em>Culex</em> mosquitoes and thus removed a barrier to developing novel genetic control methods in this important disease vector.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104309"},"PeriodicalIF":3.2,"publicationDate":"2025-03-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143770992","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 : 2025-03-31DOI: 10.1016/j.ibmb.2025.104308
Gözde Güney , Doga Cedden , Stefan Scholten , Michael Rostás
The cabbage stem flea beetle (Psylliodes chrysocephala, CSFB) is a significant pest of winter oilseed rape crops in northern Europe. CSFB adults aestivate during the summer to protect themselves from heat and desiccation stress. Trehalose, the primary hemolymph sugar, has been linked to energy homeostasis and stress resilience, but its regulation and function during aestivation remain poorly understood. Here, we investigated the roles of two trehalose transporters, Tret-1 and Tret-2, in modulating trehalose dynamics across different adult stages in CSFB. Through spatiotemporal transcript profiling, we found that Tret-1 was predominantly expressed in the fat body, where it facilitates trehalose export to the hemolymph, whereas Tret-2 expression was higher in the Malpighian tubules, mediating trehalose uptake from the hemolymph. RNA interference experiments revealed that Tret-1 is involved in transporting trehalose from the fat body into the hemolymph, while Tret-2 works reciprocally to transport trehalose from the hemolymph into the Malpighian tubules. The disruption of trehalose transportation resulted in excess glucose, glycogen, and triglyceride levels, mainly in pre-aestivation beetles. Furthermore, the knockdown of either trehalose transporter caused a compensatory increase in feeding activity in pre-aestivation beetles, while the knockdown of Tret-2 compromised resilience to heat stress. Our findings uncover the reciprocal functions of Tret-1 and Tret-2 in regulating trehalose distribution and maintaining metabolic stability during aestivation, offering insights into the physiological strategies underpinning insect survival during aestivation.
{"title":"Reciprocal roles of two trehalose transporters in aestivating cabbage stem flea beetle (Psylliodes chrysocephala)","authors":"Gözde Güney , Doga Cedden , Stefan Scholten , Michael Rostás","doi":"10.1016/j.ibmb.2025.104308","DOIUrl":"10.1016/j.ibmb.2025.104308","url":null,"abstract":"<div><div>The cabbage stem flea beetle (<em>Psylliodes chrysocephala</em>, CSFB) is a significant pest of winter oilseed rape crops in northern Europe. CSFB adults aestivate during the summer to protect themselves from heat and desiccation stress. Trehalose, the primary hemolymph sugar, has been linked to energy homeostasis and stress resilience, but its regulation and function during aestivation remain poorly understood. Here, we investigated the roles of two trehalose transporters, <em>Tret-1</em> and <em>Tret-2</em>, in modulating trehalose dynamics across different adult stages in CSFB. Through spatiotemporal transcript profiling, we found that <em>Tret-1</em> was predominantly expressed in the fat body, where it facilitates trehalose export to the hemolymph, whereas <em>Tret-2</em> expression was higher in the Malpighian tubules, mediating trehalose uptake from the hemolymph. RNA interference experiments revealed that <em>Tret-1</em> is involved in transporting trehalose from the fat body into the hemolymph, while <em>Tret-2</em> works reciprocally to transport trehalose from the hemolymph into the Malpighian tubules. The disruption of trehalose transportation resulted in excess glucose, glycogen, and triglyceride levels, mainly in pre-aestivation beetles. Furthermore, the knockdown of either trehalose transporter caused a compensatory increase in feeding activity in pre-aestivation beetles, while the knockdown of <em>Tret-2</em> compromised resilience to heat stress. Our findings uncover the reciprocal functions of <em>Tret-1</em> and <em>Tret-2</em> in regulating trehalose distribution and maintaining metabolic stability during aestivation, offering insights into the physiological strategies underpinning insect survival during aestivation.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104308"},"PeriodicalIF":3.2,"publicationDate":"2025-03-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143747685","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-03-30DOI: 10.1016/j.ibmb.2025.104307
Marie Inger Dam , Bao-Jian Ding , Kristina Brauburger , Hong-Lei Wang , Daniel Powell , Astrid T. Groot , David G. Heckel , Christer Löfstedt
The Oriental fruit moth Grapholita molesta is distributed throughout temperate regions and considered to be a pest in peach production and other high-value fruit crops in the rose family. Insecticide treatment has led to resistance development, but the use of sex pheromones in pest management has shown great promise. We investigated the pheromone biosynthesis pathway in G. molesta with the aim of elucidating pheromone evolution in the Olethreutinae subfamily of moths and harnessing pathway genes in biotechnological production of sex pheromone for use in pest management. In vivo labelling experiments suggested that an uncommon Δ8 fatty acyl desaturase is involved in sex pheromone biosynthesis. CRISPR/Cas9 knock-out of the highly expressed candidate desaturase gene Gmol_CPRQ almost completely blocked the production of Δ8 pheromone components in vivo. Heterologous expression of Gmol_CPRQ protein in yeast- or Sf9 insect cells, however, failed to demonstrate the expected Δ8 desaturase activity. Instead, Δ9 desaturase activity was observed. Co-expression in the yeast system of the electron donor, cytochrome b5, from G. molesta still produced only Δ9 desaturase activity. We suggest that Gmol_CPRQ is intimately involved in pheromone production in vivo, via an unknown reaction mechanism that may possibly involve another co-factor that is absent in the yeast and Sf9 expression systems, or depend on its subcellular site of activity. Solving this puzzle will shed further light on pheromone biosynthesis in the family Tortricidae and will be required for successful biotechnological production of fatty acids and pheromones requiring Δ8 desaturation.
{"title":"Sex pheromone biosynthesis in the Oriental fruit moth Grapholita molesta involves Δ8 desaturation","authors":"Marie Inger Dam , Bao-Jian Ding , Kristina Brauburger , Hong-Lei Wang , Daniel Powell , Astrid T. Groot , David G. Heckel , Christer Löfstedt","doi":"10.1016/j.ibmb.2025.104307","DOIUrl":"10.1016/j.ibmb.2025.104307","url":null,"abstract":"<div><div>The Oriental fruit moth <em>Grapholita molesta</em> is distributed throughout temperate regions and considered to be a pest in peach production and other high-value fruit crops in the rose family. Insecticide treatment has led to resistance development, but the use of sex pheromones in pest management has shown great promise. We investigated the pheromone biosynthesis pathway in <em>G. molesta</em> with the aim of elucidating pheromone evolution in the Olethreutinae subfamily of moths and harnessing pathway genes in biotechnological production of sex pheromone for use in pest management. <em>In vivo</em> labelling experiments suggested that an uncommon Δ8 fatty acyl desaturase is involved in sex pheromone biosynthesis. CRISPR/Cas9 knock-out of the highly expressed candidate desaturase gene <em>Gmol_CPRQ</em> almost completely blocked the production of Δ8 pheromone components <em>in vivo</em>. Heterologous expression of Gmol_CPRQ protein in yeast- or Sf9 insect cells, however, failed to demonstrate the expected Δ8 desaturase activity. Instead, Δ9 desaturase activity was observed. Co-expression in the yeast system of the electron donor, cytochrome <em>b</em>5, from <em>G. molesta</em> still produced only Δ9 desaturase activity. We suggest that <em>Gmol_CPRQ</em> is intimately involved in pheromone production <em>in vivo</em>, via an unknown reaction mechanism that may possibly involve another co-factor that is absent in the yeast and Sf9 expression systems, or depend on its subcellular site of activity. Solving this puzzle will shed further light on pheromone biosynthesis in the family Tortricidae and will be required for successful biotechnological production of fatty acids and pheromones requiring Δ8 desaturation.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104307"},"PeriodicalIF":3.2,"publicationDate":"2025-03-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143762675","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-03-29DOI: 10.1016/j.ibmb.2025.104305
Xiao Wang , Minghui Xu , Xue Kong , Shaofeng Zhong , Jeremiah Joe Kabissa , Dandan Li , Zhiwei Kang , Yongyu Xu , Zhenzhen Chen
Insects usually diapause, a process regulated by hormonal signals as an adaptive mechanism developed through long-term evolution to survive unfavorable environmental conditions. Chrysoperla nipponensis is classified as a photoperiod-sensitive insect. Treatments with short-day (SD) and long-day (LD) conditions have distinct effects on ovarian development and lipid accumulation in adults, with SD condition inducing diapause. Injecting bovine insulin promoted ovarian development and egg formation in diapause females, while injecting insulin receptor induced diapause-like traits in reproductive females. This study investigate the biological function of insulin signaling in the reproductive diapause of females of C. nipponensis. Under SD treatment the mRNA expression level of InR1 and InR2, as well as the protein expression level of InR1 were significantly reduced. This reduction led to stagnant ovarian development, increased adipose tissue mass, and a significant rise in triglyceride (TG) content. Silencing InR1 under LD conditions resulted in halted ovarian development and enhanced lipid accumulation, with the expression levels of Akt, Kr-h1, and Vg significantly decreased mirroring those observed under SD conditions. Interestingly, silencing InR2 under LD condition did not affect ovarian development. Furthermore, transcriptome analysis identified six genes (Akt, PkN, Skp2, CycB3, BTrC, and AurkA) associated with reproductive regulation and eight genes (FadΔ11, EchA, EcI, Ugts (2A3, 1–9), AR, Gpdh and Cbr) linked to lipid metabolism, all of which are involved in InR1 mediated regulation of C. nipponensis reproduction.
{"title":"The role of insulin receptor InR in photoperiod-regulated reproductive diapause of Chrysoperla nipponensis","authors":"Xiao Wang , Minghui Xu , Xue Kong , Shaofeng Zhong , Jeremiah Joe Kabissa , Dandan Li , Zhiwei Kang , Yongyu Xu , Zhenzhen Chen","doi":"10.1016/j.ibmb.2025.104305","DOIUrl":"10.1016/j.ibmb.2025.104305","url":null,"abstract":"<div><div>Insects usually diapause, a process regulated by hormonal signals as an adaptive mechanism developed through long-term evolution to survive unfavorable environmental conditions. <em>Chrysoperla nipponensis</em> is classified as a photoperiod-sensitive insect. Treatments with short-day (SD) and long-day (LD) conditions have distinct effects on ovarian development and lipid accumulation in adults, with SD condition inducing diapause. Injecting bovine insulin promoted ovarian development and egg formation in diapause females, while injecting insulin receptor induced diapause-like traits in reproductive females. This study investigate the biological function of insulin signaling in the reproductive diapause of females of <em>C. nipponensis.</em> Under SD treatment the mRNA expression level of <em>InR1</em> and <em>InR2</em>, as well as the protein expression level of <em>InR1</em> were significantly reduced. This reduction led to stagnant ovarian development, increased adipose tissue mass, and a significant rise in triglyceride (TG) content. Silencing <em>InR1</em> under LD conditions resulted in halted ovarian development and enhanced lipid accumulation, with the expression levels of <em>Akt</em>, <em>Kr-h1,</em> and <em>Vg</em> significantly decreased mirroring those observed under SD conditions. Interestingly, silencing <em>InR2</em> under LD condition did not affect ovarian development. Furthermore, transcriptome analysis identified six genes (<em>Akt</em>, <em>PkN</em>, <em>Skp2</em>, <em>CycB3</em>, <em>BTrC, and AurkA</em>) associated with reproductive regulation and eight genes (<em>FadΔ11</em>, <em>EchA</em>, <em>EcI</em>, <em>Ugts</em> (2A3, 1–9), <em>AR</em>, <em>Gpdh</em> and <em>Cbr</em>) linked to lipid metabolism, all of which are involved in <em>InR1</em> mediated regulation of <em>C. nipponensis</em> reproduction.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104305"},"PeriodicalIF":3.2,"publicationDate":"2025-03-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143738211","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 : 2025-03-28DOI: 10.1016/j.ibmb.2025.104306
Anna Christina Böhringer, Carmen Constanze Sievers, Maximilian Burghaus, Hans Merzendorfer
Activation of the dual oxidase (DUOX) pathway is an important intestinal defense mechanism against enteric infection triggering the formation of radical oxygen species by stimulating DUOX enzyme activity and/or gene expression. In insects, several studies have suggested that uracil released by pathogenic bacteria functions as a major trigger molecule for the activation of DUOX, which leads to the formation of antimicrobial hypochlorous acid (HOCl). While the recognition of pathogen-associated molecular patterns of microbes by pattern recognition receptors is well understood, the detection of uracil is still elusive. It has been postulated that a G-protein coupled receptor (GPCR) binds the pyrimidine uracil, which activates PLCβ signalling and further downstream events. So far, no pyrimidinergic receptor has been identified in insects, particularly none that binds uracil nucleotides or sugar derivatives. To identify potential candidates for insect pyrimidine receptors, we used a human P2Y4 receptor as a template to screen the Tribolium castaneum reference proteome. Four promising receptor candidates were identified, of which two were analyzed using RNA interference to determine their influence on uracil-induced TcDUOX expression, HOCl formation and development in control larvae and larvae that were challenged with the enteric pathogen Bacillus thuringiensis. Silencing TcGPCR41 resulted in a loss of uracil-induced TcDUOX expression and HOCl formation. Furthermore, the development of challenged larvae was affected in a manner like that observed in a TcDUOX knockdown. We conclude that the identified receptor may play a role in the uracil-dependent activation of the DUOX-pathways.
{"title":"A G-protein coupled receptor is involved in the DUOX pathway in Tribolium castaneum","authors":"Anna Christina Böhringer, Carmen Constanze Sievers, Maximilian Burghaus, Hans Merzendorfer","doi":"10.1016/j.ibmb.2025.104306","DOIUrl":"10.1016/j.ibmb.2025.104306","url":null,"abstract":"<div><div>Activation of the dual oxidase (DUOX) pathway is an important intestinal defense mechanism against enteric infection triggering the formation of radical oxygen species by stimulating DUOX enzyme activity and/or gene expression. In insects, several studies have suggested that uracil released by pathogenic bacteria functions as a major trigger molecule for the activation of DUOX, which leads to the formation of antimicrobial hypochlorous acid (HOCl). While the recognition of pathogen-associated molecular patterns of microbes by pattern recognition receptors is well understood, the detection of uracil is still elusive. It has been postulated that a G-protein coupled receptor (GPCR) binds the pyrimidine uracil, which activates PLCβ signalling and further downstream events. So far, no pyrimidinergic receptor has been identified in insects, particularly none that binds uracil nucleotides or sugar derivatives. To identify potential candidates for insect pyrimidine receptors, we used a human P2Y<sub>4</sub> receptor as a template to screen the <em>Tribolium castaneum</em> reference proteome. Four promising receptor candidates were identified, of which two were analyzed using RNA interference to determine their influence on uracil-induced <em>TcDUOX</em> expression, HOCl formation and development in control larvae and larvae that were challenged with the enteric pathogen <em>Bacillus thuringiensis</em>. Silencing <em>TcGPCR41</em> resulted in a loss of uracil-induced <em>TcDUOX</em> expression and HOCl formation. Furthermore, the development of challenged larvae was affected in a manner like that observed in a <em>TcDUOX</em> knockdown. We conclude that the identified receptor may play a role in the uracil-dependent activation of the DUOX-pathways.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104306"},"PeriodicalIF":3.2,"publicationDate":"2025-03-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143738212","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 : 2025-03-23DOI: 10.1016/j.ibmb.2025.104304
Zhaoyang Li , June-Sun Yoon , Zexin Zhong , Yalin Ruan , Chunxiao Yang , Xuguo Zhou , Youjun Zhang , Huipeng Pan
RNA interference (RNAi) has been shown to be relatively effective in coleopteran insects, with limited exploration into the molecular mechanisms that underlie this effectiveness. This study specifically examines the 28-spotted ladybeetle, Henosepilachna vigintioctopunctata (Hvig), known for its high RNAi efficiency. Here, we utilized RNAi and CRISPR/Cas9 techniques to identify and validate the genes involved in the RNAi pathway that enhance RNAi efficacy in Hvig. We identified a total of 15 potential genes within the RNAi pathway that may impact RNAi efficiency. The bioassay results showed that only knockdown of HvStaufenC in the 3rd instar larvae could block the abnormal body color phenotype and lethality induced by the subsequent silencing of the two marker genes, HvTH (tyrosine hydroxylase) and HvABCH1 (ATP-binding cassette H transporter gene), respectively. Additionally, successful CRISPR/Cas9-mediated knockout of HvStaufenC led to the generation of stable, heritable mutants that exhibited insensitivity to RNAi, displaying no response to RNAi targeting HvTH and HvABCH1. Compared to the wild-type strain, the HvStaufenC knockout (HvStaufenCKO) mutant females demonstrated a 42 % decrease in oviposition rate and a 41.3 % reduction in egg hatchability. This study demonstrates that HvStaufenC gene is crucial for the RNAi efficiency of Hvig and offers new evidence into the RNAi mechanisms in coleopteran species.
{"title":"HvStaufenC contributes to the high RNAi efficiency in the 28-spotted ladybeetle, Henosepilachna vigintioctopunctata","authors":"Zhaoyang Li , June-Sun Yoon , Zexin Zhong , Yalin Ruan , Chunxiao Yang , Xuguo Zhou , Youjun Zhang , Huipeng Pan","doi":"10.1016/j.ibmb.2025.104304","DOIUrl":"10.1016/j.ibmb.2025.104304","url":null,"abstract":"<div><div>RNA interference (RNAi) has been shown to be relatively effective in coleopteran insects, with limited exploration into the molecular mechanisms that underlie this effectiveness. This study specifically examines the 28-spotted ladybeetle, <em>Henosepilachna vigintioctopunctata</em> (Hvig), known for its high RNAi efficiency. Here, we utilized RNAi and CRISPR/Cas9 techniques to identify and validate the genes involved in the RNAi pathway that enhance RNAi efficacy in Hvig. We identified a total of 15 potential genes within the RNAi pathway that may impact RNAi efficiency. The bioassay results showed that only knockdown of <em>HvStaufenC</em> in the 3rd instar larvae could block the abnormal body color phenotype and lethality induced by the subsequent silencing of the two marker genes, <em>HvTH</em> (tyrosine hydroxylase) and <em>HvABCH1</em> (ATP-binding cassette H transporter gene), respectively. Additionally, successful CRISPR/Cas9-mediated knockout of <em>HvStaufenC</em> led to the generation of stable, heritable mutants that exhibited insensitivity to RNAi, displaying no response to RNAi targeting <em>HvTH</em> and <em>HvABCH1</em>. Compared to the wild-type strain, the HvStaufenC knockout (HvStaufenCKO) mutant females demonstrated a 42 % decrease in oviposition rate and a 41.3 % reduction in egg hatchability. This study demonstrates that <em>HvStaufenC</em> gene is crucial for the RNAi efficiency of Hvig and offers new evidence into the RNAi mechanisms in coleopteran species.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104304"},"PeriodicalIF":3.2,"publicationDate":"2025-03-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143704586","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 : 2025-03-22DOI: 10.1016/j.ibmb.2025.104303
Qianqian Jiao , Jiashuang Li , Yunhui Kong , Xia Sun , Sheng Qin , Muwang Li
Ultrabithorax (Ubx) is a famous gene for insect body segment determination and c-myc plays a crucial role in cell fate and is well-known as proto-oncogene. As two pivotal transcription factors, whether there is a regulatory relationship between them has not yet been reported. In our previous research, we found that excess Ubx can strongly repress the expression of Myc in the posterior silk gland of Bombyx mori (B. mori). One unresolved question is whether Ubx inhibits the Myc expression directly or indirectly. In our current study, multiple Ubx binding sites were identified from the upstream sequence and 5′-UTR of Myc. Meanwhile, the expression profile data in silkDB 3.0 showed that Ubx and Myc were expressed simultaneously in multiple tissues and developmental stages of B. mori. Luciferase reporter assay results indicated that overexpressed Ubx down-regulated the activity of luciferase which was activated by the upstream sequence and 5′-UTR of Myc in BmN cells. Electrophoretic Mobility Shift Assay (EMSA) confirmed that Ubx bound to the 5′-UTR of Myc directly. Prediction results and ChIP-Seq data suggested that this interaction might be widespread in insects. In conclusion, our study uncovered the relationship between two famous gene, Ubx and Myc, which play critical regulatory roles in insect growth and development.
{"title":"Ultrabithorax inhibits the expression of myc by directly binding to its 5′-UTR in the posterior silk gland of Bombyx mori","authors":"Qianqian Jiao , Jiashuang Li , Yunhui Kong , Xia Sun , Sheng Qin , Muwang Li","doi":"10.1016/j.ibmb.2025.104303","DOIUrl":"10.1016/j.ibmb.2025.104303","url":null,"abstract":"<div><div>Ultrabithorax (<em>Ubx</em>) is a famous gene for insect body segment determination and <em>c-myc</em> plays a crucial role in cell fate and is well-known as proto-oncogene. As two pivotal transcription factors, whether there is a regulatory relationship between them has not yet been reported. In our previous research, we found that excess Ubx can strongly repress the expression of <em>Myc</em> in the posterior silk gland of <em>Bombyx mori</em> (<em>B. mori</em>). One unresolved question is whether Ubx inhibits the <em>Myc</em> expression directly or indirectly. In our current study, multiple Ubx binding sites were identified from the upstream sequence and 5′-UTR of <em>Myc</em>. Meanwhile, the expression profile data in silkDB 3.0 showed that <em>Ubx</em> and <em>Myc</em> were expressed simultaneously in multiple tissues and developmental stages of <em>B. mori</em>. Luciferase reporter assay results indicated that overexpressed <em>Ubx</em> down-regulated the activity of luciferase which was activated by the upstream sequence and 5′-UTR of <em>Myc</em> in BmN cells. Electrophoretic Mobility Shift Assay (EMSA) confirmed that Ubx bound to the 5′-UTR of <em>Myc</em> directly. Prediction results and ChIP-Seq data suggested that this interaction might be widespread in insects. In conclusion, our study uncovered the relationship between two famous gene, <em>Ubx</em> and <em>Myc</em>, which play critical regulatory roles in insect growth and development.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104303"},"PeriodicalIF":3.2,"publicationDate":"2025-03-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143699167","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 : 2025-03-18DOI: 10.1016/j.ibmb.2025.104302
Muhammad Atif, Youngseok Lee
Flonicamid, a widely used insecticide, presents an intriguing question: does it function as an antifeedant by directly activating bitter-sensing gustatory receptor neurons (GRNs) in Drosophila melanogaster. Here, we found that electrophysiological recordings revealed that S-type labellar sensilla exhibited strong neuronal responses to flonicamid, while inhibition of bitter-sensing GRNs nullified this response. Genetic screening identified Gr28b, Gr93a, and Gr98b as essential gustatory receptors for flonicamid detection. Isoform-specific rescue experiments confirmed that Gr28b.a is responsible for restoring sensory responses in Gr28b mutants. Proboscis extension response assays demonstrated that wild-type flies avoided flonicamid, whereas Gr28b, Gr93a, and Gr98b mutants failed to. Functional rescue of these mutants restored the behavioral response, confirming the involvement of these receptors in mediating gustatory aversion. Our findings uncover a novel sensory mechanism for detecting flonicamid through specific gustatory receptors and highlight their potential as molecular targets for insect control strategies.
{"title":"Taste detection of flonicamid in Drosophila melanogaster","authors":"Muhammad Atif, Youngseok Lee","doi":"10.1016/j.ibmb.2025.104302","DOIUrl":"10.1016/j.ibmb.2025.104302","url":null,"abstract":"<div><div>Flonicamid, a widely used insecticide, presents an intriguing question: does it function as an antifeedant by directly activating bitter-sensing gustatory receptor neurons (GRNs) in <em>Drosophila melanogaster</em>. Here, we found that electrophysiological recordings revealed that S-type labellar sensilla exhibited strong neuronal responses to flonicamid, while inhibition of bitter-sensing GRNs nullified this response. Genetic screening identified <em>Gr28b</em>, <em>Gr93a</em>, and <em>Gr98b</em> as essential gustatory receptors for flonicamid detection. Isoform-specific rescue experiments confirmed that <em>Gr28b.a</em> is responsible for restoring sensory responses in <em>Gr28b</em> mutants. Proboscis extension response assays demonstrated that wild-type flies avoided flonicamid, whereas <em>Gr28b</em>, <em>Gr93a</em>, and <em>Gr98b</em> mutants failed to. Functional rescue of these mutants restored the behavioral response, confirming the involvement of these receptors in mediating gustatory aversion. Our findings uncover a novel sensory mechanism for detecting flonicamid through specific gustatory receptors and highlight their potential as molecular targets for insect control strategies.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104302"},"PeriodicalIF":3.2,"publicationDate":"2025-03-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143668585","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 : 2025-03-13DOI: 10.1016/j.ibmb.2025.104301
Jéssica Pereira , Thamara Rios , Juliana Amorim , Allana Faria-Reis , Elisa de Almeida , Matheus Neves , Samara Santos-Araújo , Lukas Selim , Felipe Bertuci , Marcyellen B. Silva , Raquel Onofre , Mellisia Brandão , Bruno Moraes , Ana Beatriz Walter-Nuno , Carlos Logullo , Gabriela O. Paiva-Silva , Katia C. Gondim , Isabela Ramos
In insects, vitellogenesis plays a critical role in providing the energy reserves needed for embryonic development as it ensures the accumulation of yolk in the oocytes. Vitellogenin (Vg), the precursor to vitellin (Vt), is primarily synthesized in the fat body of females and transported to the oocytes via receptor-mediated endocytosis. In Rhodnius prolixus, a key vector of Chagas disease, two Vg genes, Vg1 and Vg2, were characterized. These genes share 65 % amino acid identity and present the conserved Vitellogenin_N, DUF1943, and VWD domains typical of Vg proteins across various species. We found that Vg1 is expressed at significantly higher levels than Vg2 in adult females. Still, the expression of both isoforms was also detected in organs such as the flight muscle, midgut, and ovary, as well as in males and nymphs. RNAi-mediated knockdown of Vg1 and Vg2 in adult females resulted in the production of yolk-depleted eggs with drastically reduced levels of Vg and RHBP, the second most import yolk protein in this species. Despite regular oviposition rates, most of these eggs were inviable, highlighting the essential role of Vg and RHBP in embryo development. Although Vg expression was detected in adult males, the mating of Vg-knockdown males with wild-type females did not impact oviposition or egg viability, indicating that male Vg is not crucial for oogenesis in this species. Interestingly, Vg knockdown increased lifespan for both males and females, suggesting additional physiological functions beyond reproduction. These findings reveal the importance of Vg in oogenesis and embryonic development in R. prolixus while also suggesting potential non-reproductive roles of Vg in adult insect physiology.
{"title":"Functional characterization of vitellogenin unveils novel roles in RHBP uptake and lifespan regulation in the insect vector Rhodnius prolixus","authors":"Jéssica Pereira , Thamara Rios , Juliana Amorim , Allana Faria-Reis , Elisa de Almeida , Matheus Neves , Samara Santos-Araújo , Lukas Selim , Felipe Bertuci , Marcyellen B. Silva , Raquel Onofre , Mellisia Brandão , Bruno Moraes , Ana Beatriz Walter-Nuno , Carlos Logullo , Gabriela O. Paiva-Silva , Katia C. Gondim , Isabela Ramos","doi":"10.1016/j.ibmb.2025.104301","DOIUrl":"10.1016/j.ibmb.2025.104301","url":null,"abstract":"<div><div>In insects, vitellogenesis plays a critical role in providing the energy reserves needed for embryonic development as it ensures the accumulation of yolk in the oocytes. Vitellogenin (Vg), the precursor to vitellin (Vt), is primarily synthesized in the fat body of females and transported to the oocytes via receptor-mediated endocytosis. In <em>Rhodnius prolixus</em>, a key vector of Chagas disease, two Vg genes, Vg1 and Vg2, were characterized. These genes share 65 % amino acid identity and present the conserved Vitellogenin_N, DUF1943, and VWD domains typical of Vg proteins across various species. We found that Vg1 is expressed at significantly higher levels than Vg2 in adult females. Still, the expression of both isoforms was also detected in organs such as the flight muscle, midgut, and ovary, as well as in males and nymphs. RNAi-mediated knockdown of Vg1 and Vg2 in adult females resulted in the production of yolk-depleted eggs with drastically reduced levels of Vg and RHBP, the second most import yolk protein in this species. Despite regular oviposition rates, most of these eggs were inviable, highlighting the essential role of Vg and RHBP in embryo development. Although Vg expression was detected in adult males, the mating of Vg-knockdown males with wild-type females did not impact oviposition or egg viability, indicating that male Vg is not crucial for oogenesis in this species. Interestingly, Vg knockdown increased lifespan for both males and females, suggesting additional physiological functions beyond reproduction. These findings reveal the importance of Vg in oogenesis and embryonic development in <em>R. prolixus</em> while also suggesting potential non-reproductive roles of Vg in adult insect physiology.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"180 ","pages":"Article 104301"},"PeriodicalIF":3.2,"publicationDate":"2025-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143633184","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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