Pub Date : 2023-09-22DOI: 10.1016/j.ibmb.2023.104012
Alessio Iannucci , Jiao Zhu , Livio Antonielli , Anas Ayari , Karima Nasri-Ammar , Wolfgang Knoll , Paolo Pelosi , Francesca Romana Dani
The order Isopoda contains both aquatic and terrestrial species, among which Hemilepistus reaumurii, which lives in arid environments and is the most adapted to terrestrial life. Olfaction has been deeply investigated in insects while it has received very limited attention in other arthropods, particularly in terrestrial crustaceans. In insects, soluble proteins belonging to two main families, Odorant Binding Proteins (OBPs) and Chemosensory Proteins (CSPs), are contained in the olfactory sensillar lymph and are suggested to act as carriers of hydrophobic semiochemicals to or from membrane-bound olfactory receptors. Other protein families, namely Nieman-Pick type 2 (NPC2) and Lipocalins (LCNs) have been also reported as putative odorant carriers in insects and other arthropod clades. In this study, we have sequenced and analysed the transcriptomes of antennae and of the first pair of legs of H. reaumurii focusing on soluble olfactory proteins. Interestingly, we have found 13 genes encoding CSPs, whose sequences differ from those of the other arthropod clades, including non-isopod crustaceans, for the presence of two additional cysteine residues, besides the four conserved ones. Binding assays on two of these proteins showed strong affinities for fatty acids and long-chain unsaturated esters and aldehydes, putative semiochemicals for this species.
{"title":"Chemosensory proteins as putative semiochemical carriers in the desert isopod Hemilepistus reaumurii","authors":"Alessio Iannucci , Jiao Zhu , Livio Antonielli , Anas Ayari , Karima Nasri-Ammar , Wolfgang Knoll , Paolo Pelosi , Francesca Romana Dani","doi":"10.1016/j.ibmb.2023.104012","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.104012","url":null,"abstract":"<div><p>The order Isopoda contains both aquatic and terrestrial species, among which <em>Hemilepistus reaumurii</em>, which lives in arid environments and is the most adapted to terrestrial life. Olfaction has been deeply investigated in insects while it has received very limited attention in other arthropods, particularly in terrestrial crustaceans. In insects, soluble proteins belonging to two main families, Odorant Binding Proteins (OBPs) and Chemosensory Proteins (CSPs), are contained in the olfactory sensillar lymph and are suggested to act as carriers of hydrophobic semiochemicals to or from membrane-bound olfactory receptors. Other protein families, namely Nieman-Pick type 2 (NPC2) and Lipocalins (LCNs) have been also reported as putative odorant carriers in insects and other arthropod clades. In this study, we have sequenced and analysed the transcriptomes of antennae and of the first pair of legs of <em>H</em>. <em>reaumurii</em> focusing on soluble olfactory proteins<em>.</em> Interestingly, we have found 13 genes encoding CSPs<em>,</em> whose sequences differ from those of the other arthropod clades, including non-isopod crustaceans, for the presence of two additional cysteine residues, besides the four conserved ones. Binding assays on two of these proteins showed strong affinities for fatty acids and long-chain unsaturated esters and aldehydes, putative semiochemicals for this species.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-09-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"41087238","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 : 2023-09-01DOI: 10.1016/j.ibmb.2023.103986
Renato Martins da Silva , Carlos Renato de Oliveira Daumas Filho , Christiano Calixto , Jhenifer Nascimento da Silva , Cintia Lopes , Itabajara da Silva Vaz Jr , Carlos Logullo
The fat body is responsible for a variety of functions related to energy metabolism in arthropods, by controlling the processes of de novo glucose production (gluconeogenesis) and glycogen metabolism. The rate-limiting factor of gluconeogenesis is the enzyme phosphoenolpyruvate carboxykinase (PEPCK), generally considered to be the first committed step in this pathway. Although the study of PEPCK and gluconeogenesis has been for decades restricted to mammalian models, especially focusing on muscle and liver tissue, current research has demonstrated particularities about the regulation of this enzyme in arthropods, and described new functions. This review will focus on arthropod PEPCK, discuss different aspects to PEPCK regulation and function, its general role in the regulation of gluconeogenesis and other pathways. The text also presents our views on potentially important new directions for research involving this enzyme in a variety of metabolic adaptations (e.g. diapause), discussing enzyme isoforms, roles during arthropod embryogenesis, as well as involvement in vector-pathogen interactions, contributing to a better understanding of insect vectors of diseases and their control.
{"title":"PEPCK and glucose metabolism homeostasis in arthropods","authors":"Renato Martins da Silva , Carlos Renato de Oliveira Daumas Filho , Christiano Calixto , Jhenifer Nascimento da Silva , Cintia Lopes , Itabajara da Silva Vaz Jr , Carlos Logullo","doi":"10.1016/j.ibmb.2023.103986","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.103986","url":null,"abstract":"<div><p>The fat body is responsible for a variety of functions related to energy metabolism in arthropods, by controlling the processes of <em>de novo</em><span><span> glucose production<span> (gluconeogenesis) and glycogen metabolism<span>. The rate-limiting factor of gluconeogenesis is the enzyme<span> phosphoenolpyruvate carboxykinase (PEPCK), generally considered to be the first committed step in this pathway. Although the study of PEPCK and gluconeogenesis has been for decades restricted to </span></span></span></span>mammalian models<span><span>, especially focusing on muscle and liver tissue, current research has demonstrated particularities about the regulation of this enzyme in arthropods, and described new functions. This review will focus on arthropod PEPCK, discuss different aspects to PEPCK regulation and function, its general role in the regulation of gluconeogenesis and other pathways. The text also presents our views on potentially important new directions for research involving this enzyme in a variety of metabolic adaptations (e.g. diapause), discussing enzyme isoforms, roles during arthropod </span>embryogenesis, as well as involvement in vector-pathogen interactions, contributing to a better understanding of insect vectors of diseases and their control.</span></span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3036723","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 : 2023-09-01DOI: 10.1016/j.ibmb.2023.103991
Kristýna Pospíšilová , Arjen E. Van't Hof , Atsuo Yoshido , Renata Kružíková , Sander Visser , Magda Zrzavá , Kseniya Bobryshava , Martina Dalíková , František Marec
The molecular mechanisms of sex determination in moths and butterflies (Lepidoptera) with female heterogamety (WZ/ZZ) are poorly understood, except in the silkworm Bombyx mori. However, the Masculinizer (Masc) gene that controls male development and dosage compensation in B. mori, appears to be conserved in Lepidoptera, as its masculinizing function was recently confirmed in several moth species. In this work, we investigated the role of the Masc gene in sex determination of the codling moth Cydia pomonella (Tortricidae), a globally important pest of pome fruits and walnuts. The gene structure of the C. pomonella Masc ortholog, CpMasc, is similar to B. mori Masc. However, unlike B. mori, we identified 14 splice variants of CpMasc in the available transcriptomes. Subsequent screening for sex specificity and genetic variation using publicly available data and RT-PCR revealed three male-specific splice variants. Then qPCR analysis of these variants revealed sex-biased expression showing a peak only in early male embryos. Knockdown of CpMasc by RNAi during early embryogenesis resulted in a shift from male-to female-specific splicing of the C. pomonella doublesex (Cpdsx) gene, its downstream effector, in ZZ embryos, leading to a strongly female-biased sex ratio. These data clearly demonstrate that CpMasc functions as a masculinizing gene in the sex-determining cascade of C. pomonella. Our study also showed that CpMasc transcripts are provided maternally, as they were detected in unfertilized eggs after oviposition and in mature eggs dissected from virgin females. This finding is unique, as maternal provision of mRNA has rarely been studied in Lepidoptera.
{"title":"Masculinizer gene controls male sex determination in the codling moth, Cydia pomonella","authors":"Kristýna Pospíšilová , Arjen E. Van't Hof , Atsuo Yoshido , Renata Kružíková , Sander Visser , Magda Zrzavá , Kseniya Bobryshava , Martina Dalíková , František Marec","doi":"10.1016/j.ibmb.2023.103991","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.103991","url":null,"abstract":"<div><p><span>The molecular mechanisms of sex determination in moths and butterflies<span> (Lepidoptera) with female heterogamety (WZ/ZZ) are poorly understood, except in the silkworm </span></span><span><em>Bombyx </em><em>mori</em></span>. However, the <em>Masculinizer</em> (<em>Masc</em><span>) gene that controls male development and dosage compensation in </span><em>B. mori</em><span>, appears to be conserved in Lepidoptera, as its masculinizing function was recently confirmed in several moth species. In this work, we investigated the role of the </span><em>Masc</em><span> gene in sex determination of the codling moth </span><em>Cydia pomonella</em><span> (Tortricidae), a globally important pest of pome fruits and walnuts. The gene structure of the </span><em>C. pomonella Masc</em> ortholog, <em>CpMasc</em>, is similar to <em>B. mori Masc</em>. However, unlike <em>B. mori</em>, we identified 14 splice variants of <em>CpMasc</em><span><span> in the available transcriptomes. Subsequent screening for sex specificity and </span>genetic variation using publicly available data and RT-PCR revealed three male-specific splice variants. Then qPCR analysis of these variants revealed sex-biased expression showing a peak only in early male embryos. Knockdown of </span><em>CpMasc</em><span> by RNAi<span> during early embryogenesis resulted in a shift from male-to female-specific splicing of the </span></span><span><em>C. pomonella </em><em>doublesex</em></span> (<em>Cpdsx</em>) gene, its downstream effector, in ZZ embryos, leading to a strongly female-biased sex ratio. These data clearly demonstrate that <em>CpMasc</em> functions as a masculinizing gene in the sex-determining cascade of <em>C. pomonella</em>. Our study also showed that <em>CpMasc</em><span> transcripts are provided maternally, as they were detected in unfertilized eggs after oviposition and in mature eggs dissected from virgin females. This finding is unique, as maternal provision of mRNA has rarely been studied in Lepidoptera.</span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3034974","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 : 2023-08-30DOI: 10.1016/j.ibmb.2023.104003
Ping Zhou, Xin Zong, Shuo Yan, Junzheng Zhang, Dan Wang, Jie Shen
Wing dimorphism occurs in insects as a survival strategy to adapt to environmental changes. In response to environmental cues, mother aphids transmit signals to their offspring, and the offspring either emerge as winged adults or develop as wingless adults with degeneration of the wing primordia in the early instar stage. However, how the wing morph is determined in the early instar stage is still unclear. Here, we established a surgical sampling method to obtain precise wing primordium tissues for transcriptome analysis. We identified Wnt as a regulator of wing determination in the early second instar stage in the pea aphid. Inhibiting Wnt signaling via knockdown of Wnt2, Wnt11b, the Wnt receptor-encoding gene fz2 or the downstream targets vg and omb resulted in a decreased proportion of winged aphids. Activation of Wnt signaling via knockdown of miR-8, an inhibitor of the Wnt/Wg pathway, led to an increased proportion of winged aphids. Furthermore, the wing primordia of wingless nymphs underwent apoptosis in the early second instar, and cell death was activated by knockdown of fz2 under the wing-inducing condition. These results indicate that the developmental plasticity of aphid wings is modulated by the intrinsic Wnt pathway in response to environmental challenges.
{"title":"The Wnt pathway regulates wing morph determination in Acyrthosiphon pisum","authors":"Ping Zhou, Xin Zong, Shuo Yan, Junzheng Zhang, Dan Wang, Jie Shen","doi":"10.1016/j.ibmb.2023.104003","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.104003","url":null,"abstract":"<div><p>Wing dimorphism occurs in insects as a survival strategy to adapt to environmental changes. In response to environmental cues, mother aphids transmit signals to their offspring, and the offspring either emerge as winged adults or develop as wingless adults with degeneration of the wing primordia in the early instar stage. However, how the wing morph is determined in the early instar stage is still unclear. Here, we established a surgical sampling method to obtain precise wing primordium tissues for transcriptome analysis. We identified Wnt as a regulator of wing determination in the early second instar stage in the pea aphid. Inhibiting Wnt signaling via knockdown of <em>Wnt2</em>, <em>Wnt11b</em>, the Wnt receptor-encoding gene <em>fz</em>2 or the downstream targets <em>vg</em> and <em>omb</em> resulted in a decreased proportion of winged aphids. Activation of Wnt signaling via knockdown of <em>miR-8</em>, an inhibitor of the Wnt/Wg pathway, led to an increased proportion of winged aphids. Furthermore, the wing primordia of wingless nymphs underwent apoptosis in the early second instar, and cell death was activated by knockdown of <em>fz2</em> under the wing-inducing condition. These results indicate that the developmental plasticity of aphid wings is modulated by the intrinsic Wnt pathway in response to environmental challenges.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3035015","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}
The exceptional quality of silkworm silk is attributed to the amino acid sequence of its fibroin heavy chain (Fib-H) protein. The large central domain of Fib-H, which consists of glycine- and alanine-rich crystalline regions interspersed with amorphous motifs of approximately 30 amino acid residues, is considered crucial for fibrilization and determines the properties of the silk fiber. We established a technical platform to modify the Fib-H core region systematically using transcription activator-like effector nuclease-mediated homologous recombination through a somatic and germline gene knockin assay along with PCR-based screening. This efficient knockin system was used to generate a silkworm strain carrying a mutant Fib-H allele, in which the core region was replaced with a highly ordered synthetic repeat sequence of a length comparable with native Fib-H core. Heterozygous knockin mutants produced seemingly normal cocoons, whereas homozygotes did not and exhibited considerable degradation in their posterior silk glands (PSGs). Cross-sectional examination of the PSG lumen and tensile tests conducted on reeled silk threads indicated that the mutant Fib-H, which exhibited reduced stability in the PSG cells and lumen, affected the mechanical properties of the fiber. Thus, sequence manipulation of the Fib-H core domain was identified as a crucial step in successfully creating artificial silk using knockin technology.
{"title":"Fibroin heavy chain gene replacement with a highly ordered synthetic repeat sequence in Bombyx mori","authors":"Yoko Takasu, Nobuto Yamada, Katsura Kojima, Masatoshi Iga, Fumiko Yukuhiro, Tetsuya Iizuka, Taiyo Yoshioka","doi":"10.1016/j.ibmb.2023.104002","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.104002","url":null,"abstract":"<div><p>The exceptional quality of silkworm silk is attributed to the amino acid sequence of its fibroin heavy chain (Fib-H) protein. The large central domain of Fib-H, which consists of glycine- and alanine-rich crystalline regions interspersed with amorphous motifs of approximately 30 amino acid residues, is considered crucial for fibrilization and determines the properties of the silk fiber. We established a technical platform to modify the <em>Fib-H</em> core region systematically using transcription activator-like effector nuclease-mediated homologous recombination through a somatic and germline gene knockin assay along with PCR-based screening. This efficient knockin system was used to generate a silkworm strain carrying a mutant <em>Fib-H</em> allele, in which the core region was replaced with a highly ordered synthetic repeat sequence of a length comparable with native <em>Fib-H</em> core. Heterozygous knockin mutants produced seemingly normal cocoons, whereas homozygotes did not and exhibited considerable degradation in their posterior silk glands (PSGs). Cross-sectional examination of the PSG lumen and tensile tests conducted on reeled silk threads indicated that the mutant Fib-H, which exhibited reduced stability in the PSG cells and lumen, affected the mechanical properties of the fiber. Thus, sequence manipulation of the Fib-H core domain was identified as a crucial step in successfully creating artificial silk using knockin technology.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3212816","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 : 2023-08-22DOI: 10.1016/j.ibmb.2023.104001
Charles Ducker , Stanley French , Monika Pathak , Harry Taylor , Adam Sainter , William Askem , Ingrid Dreveny , Antônio Euzébio Goulart Santana , John A. Pickett , Neil J. Oldham
Leishmaniasis is a debilitating and often fatal neglected tropical disease. Males from sub-populations of the Leishmania-harbouring sandfly, Lutzomyia longipalpis, produce the diterpene sex and aggregation pheromone, sobralene, for which geranylgeranyl diphosphate (GGPP) is the likely isoprenoid precursor. We have identified a GGPP synthase (lzGGPPS) from L. longipalpis, which was recombinantly expressed in bacteria and purified for functional and kinetic analysis. In vitro enzymatic assays using LC-MS showed that lzGGPPS is an active enzyme, capable of converting substrates dimethylallyl diphosphate (DMAPP), (E)-geranyl diphosphate (GPP), (E,E)-farnesyl diphosphate (FPP) with co-substrate isopentenyl diphosphate (IPP) into (E,E,E)-GGPP, while (Z,E)-FPP was also accepted with low efficacy. Comparison of metal cofactors for lzGGPPS highlighted Mg2+ as most efficient, giving increased GGPP output when compared against other divalent metal ions tested. In line with previously characterised GGPPS enzymes, GGPP acted as an inhibitor of lzGGPPS activity. The molecular weight in solution of lzGGPPS was determined to be ∼221 kDa by analytical SEC, suggesting a hexameric assembly, as seen in the human enzyme, and representing the first assessment of GGPPS quaternary structure in insects.
{"title":"Characterisation of geranylgeranyl diphosphate synthase from the sandfly Lutzomyia longipalpis","authors":"Charles Ducker , Stanley French , Monika Pathak , Harry Taylor , Adam Sainter , William Askem , Ingrid Dreveny , Antônio Euzébio Goulart Santana , John A. Pickett , Neil J. Oldham","doi":"10.1016/j.ibmb.2023.104001","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.104001","url":null,"abstract":"<div><p>Leishmaniasis is a debilitating and often fatal neglected tropical disease. Males from sub-populations of the <em>Leishmania</em>-harbouring sandfly, <em>Lutzomyia longipalpis</em>, produce the diterpene sex and aggregation pheromone, sobralene, for which geranylgeranyl diphosphate (GGPP) is the likely isoprenoid precursor. We have identified a GGPP synthase (<em>lz</em>GGPPS) from <em>L. longipalpis</em>, which was recombinantly expressed in bacteria and purified for functional and kinetic analysis. <em>In vitro</em> enzymatic assays using LC-MS showed that <em>lz</em>GGPPS is an active enzyme, capable of converting substrates dimethylallyl diphosphate (DMAPP), (<em>E</em>)-geranyl diphosphate (GPP), (<em>E,E</em>)-farnesyl diphosphate (FPP) with co-substrate isopentenyl diphosphate (IPP) into (<em>E,E,E</em>)-GGPP, while (<em>Z,E</em>)-FPP was also accepted with low efficacy. Comparison of metal cofactors for <em>lz</em>GGPPS highlighted Mg<sup>2+</sup> as most efficient, giving increased GGPP output when compared against other divalent metal ions tested. In line with previously characterised GGPPS enzymes, GGPP acted as an inhibitor of <em>lz</em>GGPPS activity. The molecular weight in solution of <em>lz</em>GGPPS was determined to be ∼221 kDa by analytical SEC, suggesting a hexameric assembly, as seen in the human enzyme, and representing the first assessment of GGPPS quaternary structure in insects.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3403444","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 : 2023-08-01DOI: 10.1016/j.ibmb.2023.103983
Charles Grant , Kumar Saurabh Singh , Angela Hayward , Benjamin J. Hunt , Bartlomiej J. Troczka , Adam Pym , Seung-Joon Ahn , Bin Zeng , Cong-Fen Gao , Alicia Leroux , Eve Daum , Philip Süess , Dariane Souza , Jan Elias , Richard H. ffrench-Constant , John Vontas , Emmanouil Roditakis , Pablo Bielza , Christoph T. Zimmer , Chris Bass
The tomato leafminer, Tuta absoluta, is an invasive crop pest that has evolved resistance to many of the insecticides used for its control. To facilitate the investigation of the underpinning mechanisms of resistance in this species we generated a contiguous genome assembly using long-read sequencing data. We leveraged this genomic resource to investigate the genetic basis of resistance to the diamide insecticide chlorantraniliprole in Spanish strains of T. absoluta that exhibit high levels of resistance to this insecticide. Transcriptomic analyses revealed that, in these strains, resistance is not associated with previously reported target-site mutations in the diamide target-site, the ryanodine receptor, but rather is associated with the marked overexpression (20- to >100-fold) of a gene encoding a UDP-glycosyltransferase (UGT). Functional expression of this UGT, UGT34A23, via ectopic expression in Drosophila melanogaster demonstrated that it confers strong and significant resistance in vivo. The genomic resources generated in this study provide a powerful resource for further research on T. absoluta. Our findings on the mechanisms underpinning resistance to chlorantraniliprole will inform the development of sustainable management strategies for this important pest.
{"title":"Overexpression of the UDP-glycosyltransferase UGT34A23 confers resistance to the diamide insecticide chlorantraniliprole in the tomato leafminer, Tuta absoluta","authors":"Charles Grant , Kumar Saurabh Singh , Angela Hayward , Benjamin J. Hunt , Bartlomiej J. Troczka , Adam Pym , Seung-Joon Ahn , Bin Zeng , Cong-Fen Gao , Alicia Leroux , Eve Daum , Philip Süess , Dariane Souza , Jan Elias , Richard H. ffrench-Constant , John Vontas , Emmanouil Roditakis , Pablo Bielza , Christoph T. Zimmer , Chris Bass","doi":"10.1016/j.ibmb.2023.103983","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.103983","url":null,"abstract":"<div><p>The tomato leafminer, <em>Tuta absoluta</em>, is an invasive crop pest that has evolved resistance to many of the insecticides used for its control. To facilitate the investigation of the underpinning mechanisms of resistance in this species we generated a contiguous genome assembly using long-read sequencing data. We leveraged this genomic resource to investigate the genetic basis of resistance to the diamide insecticide chlorantraniliprole in Spanish strains of <em>T. absoluta</em> that exhibit high levels of resistance to this insecticide. Transcriptomic analyses revealed that, in these strains, resistance is not associated with previously reported target-site mutations in the diamide target-site, the ryanodine receptor, but rather is associated with the marked overexpression (20- to >100-fold) of a gene encoding a UDP-glycosyltransferase (UGT). Functional expression of this UGT, <em>UGT34A23</em>, via ectopic expression in <em>Drosophila melanogaster</em> demonstrated that it confers strong and significant resistance <em>in vivo</em>. The genomic resources generated in this study provide a powerful resource for further research on <em>T. absoluta</em>. Our findings on the mechanisms underpinning resistance to chlorantraniliprole will inform the development of sustainable management strategies for this important pest.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3458278","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 : 2023-08-01DOI: 10.1016/j.ibmb.2023.103984
Mi Young Noh , Karl J. Kramer , Subbaratnam Muthukrishnan , Yasuyuki Arakane
Most insects reproduce by laying eggs that have an eggshell/chorion secreted by follicle cells, which serves as a protective barrier for developing embryos. Thus, eggshell formation is vital for reproduction. Insect yellow family genes encode for secreted extracellular proteins that perform different, context-dependent functions in different tissues at various stages of development involving, for example, cuticle/eggshell coloration and morphology, molting, courtship behavior and embryo hatching. In this study we investigated the function of two of this family's genes, yellow-g (TcY-g) and yellow-g2 (TcY-g2), on the formation and morphology of the eggshell of the red flour beetle, Tribolium castaneum. Real-time PCR analysis revealed that both TcY-g and TcY-g2 were specifically expressed in the ovarioles of adult females. Loss of function produced by injection of double-stranded RNA (dsRNA) for either TcY-g or TcY-g2 gene resulted in failure of oviposition. There was no effect on maternal survival. Ovaries dissected from those dsRNA-treated females exhibited ovarioles containing not only developing oocytes but also mature eggs in their egg chambers. However, the ovulated eggs were collapsed and ruptured, resulting in swollen lateral oviducts and calyxes. TEM analysis showed that lateral oviducts were filled with electron-dense material, presumably from some cellular content leakage out of the collapsed eggs. In addition, morphological abnormalities in lateral oviduct epithelial cells and the tubular muscle sheath were evident. These results support the hypothesis that both TcY-g and TcY-g2 proteins are required for maintaining the rigidity and integrity of the chorion, which is critical for resistance to mechanical stress and/or rehydration during ovulation and egg activation in the oviducts of T. castaneum. Because Yellow-g and Yellow-g2 are highly conserved among insect species, both genes are potential targets for development of gene-based insect pest population control methods.
{"title":"Ovariole-specific Yellow-g and Yellow-g2 proteins are required for fecundity and egg chorion rigidity in the red flour beetle, Tribolium castaneum","authors":"Mi Young Noh , Karl J. Kramer , Subbaratnam Muthukrishnan , Yasuyuki Arakane","doi":"10.1016/j.ibmb.2023.103984","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.103984","url":null,"abstract":"<div><p>Most insects reproduce by laying eggs that have an eggshell/chorion secreted by follicle cells, which serves as a protective barrier for developing embryos. Thus, eggshell formation is vital for reproduction. Insect <em>yellow</em> family genes encode for secreted extracellular proteins that perform different, context-dependent functions in different tissues at various stages of development involving, for example, cuticle/eggshell coloration and morphology, molting, courtship behavior and embryo hatching. In this study we investigated the function of two of this family's genes, <em>yellow-g</em> (<em>TcY-g</em>) and <em>yellow-g2</em> (<em>TcY-g2</em>), on the formation and morphology of the eggshell of the red flour beetle, <em>Tribolium castaneum</em>. Real-time PCR analysis revealed that both <em>TcY-g</em> and <em>TcY-g2</em> were specifically expressed in the ovarioles of adult females. Loss of function produced by injection of double-stranded RNA (dsRNA) for either <em>TcY-g</em> or <em>TcY-g2</em> gene resulted in failure of oviposition. There was no effect on maternal survival. Ovaries dissected from those dsRNA-treated females exhibited ovarioles containing not only developing oocytes but also mature eggs in their egg chambers. However, the ovulated eggs were collapsed and ruptured, resulting in swollen lateral oviducts and calyxes. TEM analysis showed that lateral oviducts were filled with electron-dense material, presumably from some cellular content leakage out of the collapsed eggs. In addition, morphological abnormalities in lateral oviduct epithelial cells and the tubular muscle sheath were evident. These results support the hypothesis that both TcY-g and TcY-g2 proteins are required for maintaining the rigidity and integrity of the chorion, which is critical for resistance to mechanical stress and/or rehydration during ovulation and egg activation in the oviducts of <em>T. castaneum</em>. Because <em>Yellow-g</em> and <em>Yellow-g2</em> are highly conserved among insect species, both genes are potential targets for development of gene-based insect pest population control methods.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3458279","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 : 2023-08-01DOI: 10.1016/j.ibmb.2023.103982
Kenta Tomihara , Takashi Kiuchi
The dilute black (bd) of the silkworm Bombyx mori is a recessive mutant that produces a grayish-black color in the larval integument, instead of the characteristic white color found in wild-type larvae. In addition, eggs produced by bd females are sterile due to a deficiency in the micropylar apparatus. We identified candidate genes responsible for the bd phenotype using publicly available RNA-seq data. One of these candidate genes was homologous to the maternal gene required for meiosis (mamo) of Drosophila melanogaster, which encodes a broad-complex, tramtrack, and bric-à-brac-zinc finger (BTB-ZF) transcription factor essential for female fertility. In three independent bd strains, the expression of the B. mori mamo (Bmmamo) was downregulated in the larval integument. Using a CRISPR/Cas9-mediated knockout strategy, we found that Bmmamo knockout mutants exhibit a grayish-black color in the larval integument and female infertility. Moreover, larvae obtained from the complementation cross between bd/+ mutants and heterozygous knockouts for the Bmmamo also exhibited a grayish-black color, indicating that Bmmamo is responsible for the bd phenotype. Gene expression analysis using Bmmamo knockout mutants suggested that the BmMamo protein suppresses the expression of melanin synthesis genes. Previous comparative genome analysis revealed that the Bmmamo was selected during silkworm domestication, and we found that Bmmamo expression in the larval integument is higher in B. mori than in the wild silkworm B. mandarina, suggesting that the Bmmamo is involved in domestication-associated pigmentation changes of the silkworm.
{"title":"Disruption of a BTB-ZF transcription factor causes female sterility and melanization in the larval body of the silkworm, Bombyx mori","authors":"Kenta Tomihara , Takashi Kiuchi","doi":"10.1016/j.ibmb.2023.103982","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.103982","url":null,"abstract":"<div><p>The <em>dilute black</em> (<em>bd</em><span>) of the silkworm </span><span><em>Bombyx </em><em>mori</em></span><span> is a recessive mutant that produces a grayish-black color in the larval integument, instead of the characteristic white color found in wild-type larvae. In addition, eggs produced by </span><em>bd</em> females are sterile due to a deficiency in the micropylar apparatus. We identified candidate genes responsible for the <em>bd</em> phenotype using publicly available RNA-seq data. One of these candidate genes was homologous to the <em>maternal gene required for meiosis</em> (<span><em>mamo</em></span>) of <span><em>Drosophila melanogaster</em></span>, which encodes a broad-complex, tramtrack, and bric-à-brac-zinc finger (BTB-ZF) transcription factor essential for female fertility. In three independent <em>bd</em> strains, the expression of the <em>B. mori mamo</em> (<em>Bmmamo</em>) was downregulated in the larval integument. Using a CRISPR/Cas9-mediated knockout strategy, we found that <em>Bmmamo</em><span> knockout mutants exhibit a grayish-black color in the larval integument and female infertility. Moreover, larvae obtained from the complementation cross between </span><em>bd/+</em> mutants and heterozygous knockouts for the <em>Bmmamo</em> also exhibited a grayish-black color, indicating that <em>Bmmamo</em> is responsible for the <em>bd</em><span> phenotype. Gene expression analysis using </span><em>Bmmamo</em><span> knockout mutants suggested that the BmMamo protein suppresses the expression of melanin synthesis genes. Previous comparative genome analysis revealed that the </span><em>Bmmamo</em> was selected during silkworm domestication, and we found that <em>Bmmamo</em> expression in the larval integument is higher in <em>B. mori</em><span> than in the wild silkworm </span><em>B. mandarina</em>, suggesting that the <em>Bmmamo</em><span> is involved in domestication-associated pigmentation changes of the silkworm.</span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3343829","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 : 2023-08-01DOI: 10.1016/j.ibmb.2023.103985
Yaoyu Jiao, Subba Reddy Palli
Gene expression is regulated at various levels, including post-transcriptional mRNA modifications, where m6A methylation is the most common modification of mRNA. The m6A methylation regulates multiple stages of mRNA processing, including splicing, export, decay, and translation. How m6A modification is involved in insect development is not well known. We used the red flour beetle, Tribolium castaneum, as a model insect to identify the role of m6A modification in insect development. RNA interference (RNAi)-mediated knockdown of genes coding for m6A writers (m6A methyltransferase complex, depositing m6A to mRNA) and readers (YTH-domain proteins, recognizing and executing the function of m6A) was conducted. Knockdown of most writers during the larval stage caused a failure of ecdysis during eclosion. The loss of m6A machinery sterilized both females and males by interfering with the functioning of reproductive systems. Females treated with dsMettl3, the main m6A methyltransferase, laid significantly fewer and reduced-size eggs than the control insects. In addition, the embryonic development in eggs laid by dsMettl3 injected females was terminated in the early stages. Knockdown studies also showed that the cytosol m6A reader, YTHDF, is likely responsible for executing the function of m6A modifications during insect development. These data suggest that m6A modifications are critical for T. castaneum development and reproduction.
{"title":"N6-adenosine (m6A) mRNA methylation is required for Tribolium castaneum development and reproduction","authors":"Yaoyu Jiao, Subba Reddy Palli","doi":"10.1016/j.ibmb.2023.103985","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.103985","url":null,"abstract":"<div><p>Gene expression is regulated at various levels, including post-transcriptional mRNA modifications, where m<sup>6</sup><span>A methylation is the most common modification of mRNA. The m</span><sup>6</sup><span>A methylation regulates multiple stages of mRNA processing, including splicing, export, decay, and translation. How m</span><sup>6</sup><span>A modification is involved in insect development is not well known. We used the red flour beetle, </span><span><em>Tribolium </em><em>castaneum</em><em>,</em></span> as a model insect to identify the role of m<sup>6</sup><span>A modification in insect development. RNA interference (RNAi)-mediated knockdown of genes coding for m</span><sup>6</sup>A writers (m<sup>6</sup><span>A methyltransferase complex, depositing m</span><sup>6</sup>A to mRNA) and readers (YTH-domain proteins, recognizing and executing the function of m<sup>6</sup><span><span>A) was conducted. Knockdown of most writers during the larval stage caused a failure of ecdysis during </span>eclosion. The loss of m</span><sup>6</sup>A machinery sterilized both females and males by interfering with the functioning of reproductive systems. Females treated with <em>dsMettl3</em>, the main m<sup>6</sup><span>A methyltransferase, laid significantly fewer and reduced-size eggs than the control insects. In addition, the embryonic development in eggs laid by </span><em>dsMettl3</em> injected females was terminated in the early stages. Knockdown studies also showed that the cytosol m<sup>6</sup>A reader, YTHDF, is likely responsible for executing the function of m<sup>6</sup>A modifications during insect development. These data suggest that m<sup>6</sup>A modifications are critical for <em>T. castaneum</em> development and reproduction.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":null,"pages":null},"PeriodicalIF":3.8,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"3343830","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}