Insect Biochemistry and Molecular Biology最新文献

{"title":"Identification of salivary proteins of the cowpea aphid Aphis craccivora by transcriptome and LC-MS/MS analyses","authors":"Shanmugasundram Pavithran ,&nbsp;Marimuthu Murugan ,&nbsp;Jayakanthan Mannu ,&nbsp;Kalenahalli Yogendra ,&nbsp;Venkatasamy Balasubramani ,&nbsp;Hemalatha Sanivarapu ,&nbsp;Sankarasubramanian Harish ,&nbsp;Senthil Natesan","doi":"10.1016/j.ibmb.2023.104060","DOIUrl":"10.1016/j.ibmb.2023.104060","url":null,"abstract":"<div><p><span><span>Aphid salivary proteins mediate the interaction between aphids and their host plants. Moreover, these proteins facilitate digestion, detoxification of secondary metabolites, as well as activation and suppression of plant defenses. The </span>cowpea aphid, </span><em>Aphis craccivora</em><span><span><span>, is an important sucking pest of leguminous crops worldwide. Although aphid saliva plays an important role in aphid plant interactions, knowledge of the cowpea aphid salivary proteins is limited. In this study, we performed </span>transcriptomic and LC-MS/MS analyses to identify the proteins present in the </span>salivary glands and saliva of </span><em>A. craccivora</em><span><span>. A total of 1,08,275 assembled transcripts were identified in the salivary glands of aphids. Of all these assembled transcripts, 53,714 (49.11%) and 53,577 (49.48%) transcripts showed high similarity to known proteins in the Nr and UniProt databases, respectively. A total of 2159 proteins were predicted as secretory proteins<span> from the salivary gland transcriptome dataset, which contain </span></span>digestive enzymes<span><span>, detoxification enzymes, previously known effectors and </span>elicitors<span>, and potential proteins whose functions have yet to be determined. The proteomic analysis of aphid saliva resulted in the identification of 171 proteins. Tissue-specific expression of selected genes using RT-PCR showed that three genes were expressed only in the salivary glands. Overall, our results provide a comprehensive repertoire of cowpea aphid salivary proteins from the salivary gland and saliva, which will be a good resource for future effector functional studies and might also be useful for sustainable aphid management.</span></span></span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"165 ","pages":"Article 104060"},"PeriodicalIF":3.8,"publicationDate":"2023-12-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138715110","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}

{"title":"Evidence that hematophagous triatomine bugs may eat plants in the wild","authors":"Jean-Luc Da Lage ,&nbsp;Alice Fontenelle ,&nbsp;Jonathan Filée ,&nbsp;Marie Merle ,&nbsp;Jean-Michel Béranger ,&nbsp;Carlos Eduardo Almeida ,&nbsp;Elaine Folly Ramos ,&nbsp;Myriam Harry","doi":"10.1016/j.ibmb.2023.104059","DOIUrl":"10.1016/j.ibmb.2023.104059","url":null,"abstract":"<div><p><span><span>Blood feeding is a secondary adaptation in hematophagous bugs. Many proteins are secreted in the saliva that are devoted to coping with the host's defense and to process the blood meal. Digestive enzymes<span><span> that are no longer required for a blood meal would be expected to be eventually lost. Yet, in many strictly hematophagous arthropods, α-amylase genes, which encode the </span>enzymes that digest starch from plants, are still present and transcribed, including in the </span></span>kissing bug </span><span><em>Rhodnius</em><em> prolixus</em></span><span> (Hemiptera, Reduviidae) and its related species, which transmit the Chagas disease. We hypothesized that retaining α-amylase could be advantageous if the bugs occasionally consume plant tissues. We first checked that the α-amylase protein of </span><em>Rhodnius robustus</em><span> retains normal amylolytic activity. Then we surveyed hundreds of gut DNA extracts from the sylvatic </span><em>R. robustus</em> to detect traces of plants. We found plant DNA in 8% of the samples, mainly identified as <span><em>Attalea</em></span><span> palm trees, where </span><em>R. robustus</em> are usually found. We suggest that although of secondary importance in the blood-sucking bugs, α-amylase may be needed during occasional plant feeding and thus has been retained.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"165 ","pages":"Article 104059"},"PeriodicalIF":3.8,"publicationDate":"2023-12-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138631515","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}

{"title":"Advances in understanding insect chitin biosynthesis","authors":"Ailing Yu ,&nbsp;Marius Beck ,&nbsp;Hans Merzendorfer ,&nbsp;Qing Yang","doi":"10.1016/j.ibmb.2023.104058","DOIUrl":"10.1016/j.ibmb.2023.104058","url":null,"abstract":"<div><p><span>Chitin, a natural polymer of N-acetylglucosamine chains, is a principal component of the apical </span>extracellular matrix<span> in arthropods. Chitin microfibrils<span><span> serve as structural components of natural biocomposites<span> present in the extracellular matrix of a variety of invertebrates including sponges, molluscs, nematodes, fungi and arthropods. In this review, we summarize the frontier advances of insect chitin synthesis. More specifically, we focus on the chitin synthase (CHS), which catalyzes the key </span></span>biosynthesis<span><span><span> step. CHS is also known as an attractive insecticidal target in that this enzyme is absent in mammals, birds or plants. As no insect chitin synthase structure have been reported so far, we review recent studies on </span>glycosyltransferase domain structures derived from fungi and </span>oomycetes, which are conserved in CHS from all species containing chitin. Auxiliary proteins, which coordinate with CHS in chitin biosynthesis and assembly, are also discussed.</span></span></span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104058"},"PeriodicalIF":3.8,"publicationDate":"2023-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138566099","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}

{"title":"Expression of the non-neuronal cholinergic system components in Malpighian tubules of Mythimna separata and evidence for non-neuronal acetylcholine synthesis","authors":"Baolei Zhan,&nbsp;Ting Zhou,&nbsp;Di Tong,&nbsp;Zile Xu,&nbsp;Xiaoyu Liang,&nbsp;Anqi Ma,&nbsp;Shumin Lü","doi":"10.1016/j.ibmb.2023.104047","DOIUrl":"10.1016/j.ibmb.2023.104047","url":null,"abstract":"<div><p><span>The non-neuronal cholinergic system, widely distributed in nature, is an ancient system that has not been well studied in insects. This study aims to investigate the key components of the cholinergic system and to identify the non-neuronal acetylcholine<span> (ACh)-producing cells and the acting sites of ACh in the Malpighian tubules (MTs) of </span></span><span><em>Mythimna separata</em></span><span><span>. We found that non-neuronal ACh in MTs is synthesized by carnitine acetyltransferase (CarAT), rather than </span>choline acetyltransferase<span> (ChAT), as confirmed by using enzyme inhibitors<span> and high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS/MS). Fluorescence </span></span></span><em>in situ</em><span> hybridization revealed the presence of CarAT mRNA within MTs, specifically localized in the principal cells. Immunohistochemistry<span> showed strong staining for A-mAChR, a muscarinic acetylcholine receptor, in the principal cells. Pharmacological analysis further demonstrated that ACh acts through A-mAChR in the principal cells to increase the intracellular Ca</span></span>²⁺ concentration. These findings provide compelling evidence for the existence of a non-neuronal cholinergic system in the MTs of <em>M. separata</em>, and the principal cells play a crucial role in ACh synthesis via CarAT.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104047"},"PeriodicalIF":3.8,"publicationDate":"2023-12-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138566350","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}

{"title":"Serine protease homolog pairs CLIPA4-A6, A4-A7Δ, and A4-A12 act as cofactors for proteolytic activation of prophenoloxidase-2 and -7 in Anopheles gambiae","authors":"Qiao Jin,&nbsp;Yang Wang,&nbsp;Yingxia Hu,&nbsp;Yan He,&nbsp;Chao Xiong,&nbsp;Haobo Jiang","doi":"10.1016/j.ibmb.2023.104048","DOIUrl":"https://doi.org/10.1016/j.ibmb.2023.104048","url":null,"abstract":"<div><p><span><span>Phenoloxidase (PO) catalyzed melanization and other insect immune responses are mediated by </span>serine proteases (SPs) and their noncatalytic homologs (SPHs). Many of these SP-like proteins have a regulatory clip domain and are called CLIPs. In most insects studied so far, PO precursors are activated by a PAP (</span><em>i.e</em>., PPO activating protease) and its cofactor of clip-domain SPHs. Although melanotic encapsulation is a well-known refractory mechanism of mosquitoes against malaria parasites, it is unclear if a cofactor is required for PPO activation. In <span><em>Anopheles gambiae</em></span>, CLIPA4 is 1:1 orthologous to <span><em>Manduca sexta</em></span> SPH2; CLIPs A5−7, A12–14, A26, A31, A32, E6, and E7 are 11:4 orthologous to <em>M. sexta</em> SPH1a, 1b, 4, and 101, SPH2 partners in the cofactors. Here we produced proCLIPs A4, A6, A7Δ, A12, and activated them with CLIPB9 or <em>M. sexta</em> PAP3. <em>A. gambiae</em> PPO2 and PPO7 were expressed in <em>Escherichia coli</em> for use as PAP substrates. CLIPB9 was mutated to CLIPB9_Xa by including a Factor X_a<span> cleavage site. CLIPA7Δ was a deletion mutant with a low complexity region removed. After PAP3 or CLIPB9</span>_Xa processing, CLIPA4 formed a high <em>M</em>_r complex with CLIPA6, A7Δ or A12, which assisted PPO2 and PPO7 activation. High levels of specific PO activity (55−85 U/μg for PO2 and 1131−1630 U/μg for PO7) were detected <em>in vitro</em>, indicating that cofactor-assisted PPO activation also occurs in this species. The cleavage sites and mechanisms for complex formation and cofactor function are like those reported in <em>M. sexta</em> and <span><em>Drosophila melanogaster</em></span>. In conclusion, these data suggest that the three (and perhaps more) SPHI-II pairs may form cofactors for CLIPB9-mediated activation of PPOs for melanotic encapsulation in <em>A. gambiae</em>.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104048"},"PeriodicalIF":3.8,"publicationDate":"2023-12-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138490726","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}

{"title":"Evidence for a role of SNMP2 and antennal support cells in sensillum lymph clearance processes of moth pheromone-responsive sensilla","authors":"Sina Cassau,&nbsp;Jürgen Krieger","doi":"10.1016/j.ibmb.2023.104046","DOIUrl":"10.1016/j.ibmb.2023.104046","url":null,"abstract":"<div><p>In insect antenna, following the activation of olfactory sensory neurons, odorant molecules are inactivated by enzymes in the sensillum lymph. How the inactivation products are cleared from the sensillum lymph is presently unknown. Here we studied the role of support cells (SCs) and the so-called sensory neuron membrane protein 2 (SNMP2), a member of the CD36 family of lipid transporters abundantly expressed in SCs, in sensillum lymph clearance processes in the moths <em>Heliothis virescens</em> and <em>Bombyx mori</em>. In these species, the sex pheromone components are inactivated to long-chain fatty acids. To approach a role of SNMP2 in the removal of such inactivation products, we analyzed the uptake of a fluorescent long-chain fatty acid analog into a newly generated HvirSNMP2-expressing cell line. We found an increased uptake of the analog into SNMP2-cells compared to control cells, which could be blocked by the CD36 protein inhibitor, SSO. Furthermore, analyses of sensilla from antenna treated with the fatty acid analog indicated that SNMP2-expressing SCs are able to take up fatty acids from the sensillum lymph. In addition, sensilla from SSO-pretreated antenna of <em>B. mori</em> showed reduced removal of the fluorescent analog from the sensillum lymph. Finally, we revealed that SSO pretreatment of male silkmoth antenna significantly prolonged the duration of the female pheromone-induced wing-fluttering behavior, possibly as a result of impaired lymph clearance processes. Together our findings in <em>H. virescens</em> and <em>B. mori</em> support a pivotal role of olfactory SCs in sensillum lymph maintenance processes and suggest an integral role of SNMP2 in the removal of lipophilic “waste products” such as fatty acids resulting from sex pheromone inactivation.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104046"},"PeriodicalIF":3.8,"publicationDate":"2023-12-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0965174823001406/pdfft?md5=1b627524123b913872620830ed0e2d5d&pid=1-s2.0-S0965174823001406-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138476345","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}

{"title":"Overexpression of the F116V allele of CYP9A186 in transgenic Helicoverpa armigera confers high-level resistance to emamectin benzoate","authors":"Lin Li ,&nbsp;Yayun Zuo ,&nbsp;Yu Shi ,&nbsp;Yihua Yang ,&nbsp;Yidong Wu","doi":"10.1016/j.ibmb.2023.104042","DOIUrl":"10.1016/j.ibmb.2023.104042","url":null,"abstract":"<div><p><span>Insect cytochrome P450s play important roles in the detoxification of xenobiotics and the metabolic resistance to insecticides. However, the approach for </span><em>in vivo</em><span> validation of the contribution of specific candidate P450s to resistance is still limited in most non-model insect species. Previous studies with heterologous expression and </span><em>in vitro</em> functional assays have confirmed that a natural substitution (F116V) in the substrate recognition site 1 (SRS1) of the CYP9A186 of <span><em>Spodoptera exigua</em></span><span> is a gain-of-function mutation, which results in detoxification capability of and thus high-level resistance to both emamectin<span> benzoate (EB) and abamectin. In this study, we established an effective </span></span><em>piggyBac</em>-based transformation system in the serious agricultural pest <span><em>Helicoverpa armigera</em></span> and overexpressed <em>in vivo</em> a resistance P450 allele, <em>CYP9A186-F116V</em><span>, from another lepidopteran pest </span><em>Spodoptera exigua</em><span>. Bioassays showed that transgenic </span><em>H. armigera</em> larvae expressing CYP9A186-F116V obtained 358-fold and 38.6-fold resistance to EB and abamectin, respectively. In contrast, a transgenic line of <span><em>Drosophila melanogaster</em></span><span> overexpressing this P450 variant only confers ∼20-fold resistance to the two insecticides. This bias towards the resistance level revealed that closely related species might provide a more appropriate cellular environment for gene expression and subsequent toxicokinetics of insecticides. These results not only present an alternative method for </span><em>in vivo</em> functional characterization of P450s in <em>H. armigera</em> and other phylogenetically close species but also provide a valuable genetic engineering toolkit for the genetic manipulation of <em>H. armigera.</em></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"163 ","pages":"Article 104042"},"PeriodicalIF":3.8,"publicationDate":"2023-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138456830","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}

{"title":"The crystal structure of insecticidal protein Txp40 from Xenorhabdus nematophila reveals a two-domain unique binary toxin with homology to the toxin-antitoxin (TA) system","authors":"Omkar U. Kinkar ,&nbsp;Ashwani Kumar ,&nbsp;Arpit Prashar ,&nbsp;Beena Yadav ,&nbsp;Ashok B. Hadapad ,&nbsp;Ramesh S. Hire ,&nbsp;Ravindra D. Makde","doi":"10.1016/j.ibmb.2023.104045","DOIUrl":"10.1016/j.ibmb.2023.104045","url":null,"abstract":"<div><p>Txp40 is a ubiquitous, conserved, and novel toxin from <span><em>Xenorhabdus</em></span> and <span><em>Photorhabdus</em></span><span> bacteria, toxic to a wide range of insect pests<span>. However, the three-dimensional structure and toxicity mechanism for Txp40 or any of its sequence homologs are not yet known. Here, we are reporting the crystal structure of the insecticidal protein Txp40 from </span></span><span><em>Xenorhabdus nematophila</em></span><span> at 2.08 Å resolution. The Txp40 was structurally distinct from currently known insecticidal proteins. Txp40 consists of two structurally different domains, an N-terminal domain (NTD) and a C-terminal domain (CTD), primarily joined by a 33-residue long linker peptide. Txp40 displayed proteolytic propensity. Txp40 gets proteolyzed, removing the linker peptide, which is essential for proper crystal packing. NTD adopts a novel fold composed of nine amphipathic helices and has no shared sequence or structural homology to any known proteins. CTD has structural homology with RNases of type II toxin-antitoxin (TA) complex belonging to the RelE/ParE toxin domain superfamily. NTD and CTD were individually toxic to </span><span><em>Galleria mellonella</em></span> larvae. However, maximal toxicity was observed when both domains were present. Our results suggested that the Txp40 acts as a two-domain binary toxin, which is unique and different from any known binary toxins and insecticidal proteins. Txp40 is also unique because it belongs to the prokaryotic RelE/ParE toxin family with a toxic effect on eukaryotic organisms, in contrast to other members of the same family. Broad insect specificity and unique binary toxin complex formation make Txp40 a viable candidate to overcome the development of resistance in insect pests.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104045"},"PeriodicalIF":3.8,"publicationDate":"2023-11-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138469618","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}

{"title":"Culex quinquefasciatus membrane-bound alkaline phosphatase is a putative receptor for Lysinibacillus sphaericus Tpp49Aa1 toxin","authors":"Qingyun Guo,&nbsp;Wei Li,&nbsp;Yingchao Niu,&nbsp;Xiaohua Dai,&nbsp;Lin Chen","doi":"10.1016/j.ibmb.2023.104044","DOIUrl":"10.1016/j.ibmb.2023.104044","url":null,"abstract":"<div><p>The binary toxin Cry48Aa1/Tpp49Aa1 produced by <span><em>Lysinibacillus sphaericus</em></span><span> exhibits potent toxicity against Culicidae larvae. Both Cry48Aa1 and Tpp49Aa1 toxins are crucial for binding to the toxin receptor in </span><span><em>Culex quinquefasciatus</em></span><span> larvae, albeit with different binding sites. Previous studies have identified Glu71, a membrane-bound α-glucosidase, as a putative binding protein for the Cry48Aa1 toxin, involved in the Cry48Aa1/Tpp49Aa1 toxicity. In this study, we employed pulldown assays to identify a group of Tpp49Aa1-binding proteins from </span><em>C. quinquefasciatus</em><span><span><span> solubilized midgut brush-border membrane proteins (BBMFs). </span>RNA interference assays revealed that the silencing of an </span>alkaline phosphatase gene (referred to as ALP1263) in </span><em>C. quinquefasciatus</em><span> resulted in a significant reduction in larval mortality upon exposure to Cry48Aa1/Tpp49Aa1 toxin in vivo. Furthermore, the ALP1263 protein exhibited specific and high-affinity binding to the Tpp49Aa1 toxin, with a dissociation constant (Kd) of approximately 57.3 nM. The dot blot analysis demonstrated that Tpp49Aa1 C-terminal region was essential for its interaction with the ALP1263 protein. In summary, our findings establish ALP1263 as a functional receptor for Tpp49Aa1 and emphasize its role in the toxicity of Cry48Aa1/Tpp49Aa1.</span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104044"},"PeriodicalIF":3.8,"publicationDate":"2023-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138456829","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}

{"title":"Insights into unique features of Drosophila CYP4G enzymes","authors":"Mary Kefi ,&nbsp;Parasyris Konstantinos ,&nbsp;Vasileia Balabanidou ,&nbsp;Chara Sarafoglou ,&nbsp;Dimitra Tsakireli ,&nbsp;Vassilis Douris ,&nbsp;Maria Monastirioti ,&nbsp;Jean-Didier Maréchal ,&nbsp;René Feyereisen ,&nbsp;John Vontas","doi":"10.1016/j.ibmb.2023.104041","DOIUrl":"10.1016/j.ibmb.2023.104041","url":null,"abstract":"<div><p><span>The cytochrome P450 enzymes of the CYP4G subfamily are some of the most intriguing insect P450s in terms of structure and function. In </span><em>Drosophila</em><span><span><span>, CYP4G1 is highly expressed in the oenocytes and is the last enzyme in the </span>biosynthesis<span> of cuticular hydrocarbons, while CYP4G15 is expressed in the brain and is of unknown function. Both proteins have a CYP4G-specific and characteristic </span></span>amino acid sequence insertion corresponding to a loop between the G and H helices whose function is unclear. Here we address these enigmatic structural and functional features of </span><em>Drosophila</em><span> CYP4Gs. First, we used reverse genetics to generate </span><span><em>D. </em><em>melanogaster</em></span><span><span><span> strains in which all or part of the CYP4G-specific loop was removed from CYP4G1. We showed that the full loop was not needed for proper folding of the P450, but it is essential for function, and that just a short stretch of six amino acids is required for the enzyme’s ability to make hydrocarbons. Second, we confirmed by </span>immunocytochemistry that CYP4G15 is expressed in the brain and showed that it is specifically associated with the cortex glia cell subtype. We then expressed CYP4G15 ectopically in oenocytes, revealing that it can produce of a blend of hydrocarbons, albeit to quantitatively lower levels resulting in only a partial rescue of CYP4G1 knockdown flies. The CYP4G1 structural variants studied here should facilitate the biochemical characterization of CYP4G enzymes. Our results also raise the question of the putative role of hydrocarbons and their synthesis by cortex </span>glial cells.</span></p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"164 ","pages":"Article 104041"},"PeriodicalIF":3.8,"publicationDate":"2023-11-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138439936","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}