Insect Biochemistry and Molecular Biology最新文献

{"title":"Multi-omics analysis reveal the fall armyworm Spodoptera frugiperda tolerate high temperature by mediating chitin-related genes","authors":"Xiaorui Yan ,&nbsp;Zihua Zhao ,&nbsp;Shiqian Feng ,&nbsp;Yongjun Zhang ,&nbsp;Zhenying Wang ,&nbsp;Zhihong Li","doi":"10.1016/j.ibmb.2024.104192","DOIUrl":"10.1016/j.ibmb.2024.104192","url":null,"abstract":"<div><div>Climate change facilitates the rapid invasion of agricultural pests, threatening global food security. The fall armyworm <em>Spodoptera frugiperda</em> is a highly polyphagous migratory pest tolerant to high temperatures, allowing its proliferation in harsh thermal environments. We aimed to demonstrate mechanisms of its high-temperature tolerance, particularly transcriptional and metabolic regulation, which are poorly understood. To achieve the aim, we examined the impact and mechanism of heat events on <em>S. frugiperda</em> by using multiple approaches: ecological measurements, transcriptomics, metabolomics, RNAi, and CRISPR/Cas9 technology. We observed that several physiological indices (larval survival rate, larval period, pupation rate, pupal weight, eclosion rate, and average fecundity) decreased as the temperature increased, with the 32 °C treatment displaying a significant difference from the control group at 26 °C. Significantly upregulated expression of genes encoding endochitinase and chitin deacetylase was observed in the chitin-binding, extracellular region, and carbohydrate metabolic process GO terms of hemolymph, fat body, and brain, exhibiting a tissue-specific pattern. Significantly enriched pathways (e.g., cutin, suberin, and wax biosynthesis; oxidative phosphorylation and cofactor biosynthesis; diverse amino acid biosynthesis and degradation; carbon metabolism; and energy metabolism), all of which are essential for <em>S. frugiperda</em> larvae to tolerate temperature, were found in metabolites that were expressed differently. Successful RNA interference targeting of the three chitin-related genes reduced gene expression levels and larval survival rate. Knockout of the endochitinase gene by using the CRISPR/Cas9 system significantly reduced the relative gene expression and increased sensitivity to high-temperature exposure. On the basis of our findings, theoretical foundations for understanding the high-temperature tolerance of <em>S. frugiperda</em> populations and latent genetic control strategies were established.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"174 ","pages":"Article 104192"},"PeriodicalIF":3.2,"publicationDate":"2024-10-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142442780","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":"Identification of salivary proteins in the rice leaf folder Cnaphalocrocis medinalis by transcriptome and LC-MS/MS analyses","authors":"Jiarong Cui ,&nbsp;Xianjing Yao ,&nbsp;Zhihan Ni ,&nbsp;Hongfeng Zhao ,&nbsp;Yajun Yang ,&nbsp;Hongxing Xu ,&nbsp;Zhongxian Lu ,&nbsp;Pingyang Zhu","doi":"10.1016/j.ibmb.2024.104191","DOIUrl":"10.1016/j.ibmb.2024.104191","url":null,"abstract":"<div><div>Salivary proteins in the oral secretion (OS) of chewing insects play a crucial role in insect-plant interactions during feeding. The rice leaf folder <em>Cnaphalocrocis medinalis</em>, a notorious pest in global rice production, triggers defense responses during feeding, but little is known about its salivary proteins. In this study, we confirmed that <em>C. medinalis</em> releases OS during feeding. By employing transcriptomic analysis and liquid chromatography-tandem mass spectroscopy (LC-MS/MS), we examined the salivary proteins from labial salivary glands and OS from <em>C. medinalis</em>. A total of 14,397 genes were expressed at the RNA level and 229 salivary proteins were identified. Comparative analysis with other 25 arthropod species revealed that 43 proteins were unique to <em>C. medinalis</em>. Expression pattern analysis revealed that most of the selected genes were highly expressed in the gut and the larval stages (4th–5th instar). These findings provide a comprehensive resource for future functional studies of salivary proteins, offering new insights into the molecular mechanisms by which <em>C. medinalis</em> modulates plant defenses and potential applications in pest management.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"174 ","pages":"Article 104191"},"PeriodicalIF":3.2,"publicationDate":"2024-10-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142405800","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":"Bipartite nuclear localization sequence is indispensable for nuclear import and stability of self-dimerization of ADARa in Bombyx mori","authors":"Song Jiang ,&nbsp;Junzhe Peng ,&nbsp;Syeda Saneela ,&nbsp;Ruoyun Shi ,&nbsp;Daoming Wang ,&nbsp;Qingheng Tang ,&nbsp;Xiaming Shi ,&nbsp;Yan Meng","doi":"10.1016/j.ibmb.2024.104190","DOIUrl":"10.1016/j.ibmb.2024.104190","url":null,"abstract":"<div><div>The conservative post-transcriptional modification in mammals and <em>Drosophila</em> is adenosine-to-inosine (A-to-I) deamination in double-stranded RNA, catalyzed by RNA-editing enzymes known as adenosine deaminases acting on RNA (ADARs). The traditional nuclear import pathway for ADARs involves the recognition of a putative classical nuclear localization sequence (NLS) by importin α4 and α5. In our previous research, ADAR in silkworm, <em>Bombyx mori</em> (BmADARa) was confirmed predominantly located in the nucleus. However, the location of the NLS in BmADARa and its impact on nuclear import and self-dimerization remained unclear. Utilizing NLS prediction software, we predicted the presence of a bipartite NLS within the amino-terminal, 85 amino acids of BmADARa (N85). This prediction was validated through point mutation, which demonstrated that the bipartite NLS could directly mediate nuclear import of BmADARa. Co-immunoprecipitation analysis revealed that BmADARa is mainly dependent on BmKaryopherin α3 (homologous to mammalian importin α4) for nuclear import, although both BmKaryopherin α3 and BmImportin α5 could recognize bipartite NLS. The N-terminal truncated mutants and the bipartite NLS mutants of BmADARa suggest that the bipartite NLS is the major nuclear import site and a crucial structure for self-dimerization of BmADARa. In conclusion, the N-terminal bipartite NLS of BmADARa is recognized by BmKaryopherin α3 and BmImportin α5, facilitating its nuclear import. This promotes BmADARa self-dimerization and maintains the stability of dimerization, thereby enhancing its editing efficiency on target substrates. The results of this research demonstrate the role of bipartite NLS in BmADARa editing and laying a foundation for further research on the regulation of BmADARa in the growth and development in <em>B. mori</em>.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"174 ","pages":"Article 104190"},"PeriodicalIF":3.2,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142398829","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":"Genome editing: A novel approach to manage insect vectors of plant viruses","authors":"Sumit Jangra ,&nbsp;Jesse Potts ,&nbsp;Amalendu Ghosh ,&nbsp;Dakshina R. Seal","doi":"10.1016/j.ibmb.2024.104189","DOIUrl":"10.1016/j.ibmb.2024.104189","url":null,"abstract":"<div><div>Insect vectors significantly threaten global agriculture by transmitting numerous plant viruses. Various measures, from conventional insecticides to genetic engineering, are used to mitigate this threat. However, none provide complete resistance. Therefore, researchers are looking for novel control options. In recent years with the advancements in genomic technologies, genomes and transcriptomes of various insect vectors have been generated. However, the lack of knowledge about gene functions hinders the development of novel strategies to restrict virus spread. RNA interference (RNAi) is widely used to elucidate gene functions, but its variable efficacy hampers its use in managing insect vectors and plant viruses. Genome editing has the potential to overcome these challenges and has been extensively used in various insect pest species. This review summarizes the progress and potential of genome editing in plant virus vectors and its application as a functional genomic tool to elucidate virus-vector interactions. We also discuss the major challenges associated with editing genes of interest in insect vectors.</div></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"174 ","pages":"Article 104189"},"PeriodicalIF":3.2,"publicationDate":"2024-09-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142338236","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 role of OR5, which is highly expressed in the winged grain aphid Sitobion miscanthi, in specific recognition of EBF","authors":"Jia Fan ,&nbsp;Bing Wang ,&nbsp;Tianjiao Zhang ,&nbsp;Miaomiao Yu ,&nbsp;Mengyu Wang ,&nbsp;Siyu Zhang ,&nbsp;Changqing Su ,&nbsp;Jingyang Xu ,&nbsp;Xin Jiang ,&nbsp;Qian Zhang ,&nbsp;Guirong Wang ,&nbsp;Julian Chen","doi":"10.1016/j.ibmb.2024.104180","DOIUrl":"10.1016/j.ibmb.2024.104180","url":null,"abstract":"<div><p>Winged parthenogenetic aphids are mainly responsible for migration and dispersal. Aphid alarm pheromone (<em>E</em>)-β-Farnesene (EBF) has dual effects on repelling and stimulating wing differentiation in aphids. Previous studies have shown that the odorant coreceptor SmisOrco is involved in the perception of EBF by <em>S. miscanthi</em>; however, its EBF-specific odorant receptor (OR) and the difference between winged and wingless aphids remain unclear. In this study, the <em>Xenopus oocyte</em> expression system and RNAi technology were used to detect the transmission of EBF signals, and it was found that the olfactory receptor <em>SmisOR5</em> is an EBF-specific OR in <em>S. miscanthi</em> and is specifically highly expressed in the antennae of winged aphids. Furthermore, when <em>OR5</em> was silenced with dsRNA, the repellent effect of EBF was weakened, and aphids showed more active aimless movements. Therefore, as a specific OR for EBF, the high expression level of <em>SmisOR5</em> in winged aphids suggests a molecular basis for its high sensitivity to EBF. This study advances our understanding of the molecular mechanisms of aphid EBF perception and provides novel ideas for effective management and prevention of the migration of winged aphids.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"173 ","pages":"Article 104180"},"PeriodicalIF":3.2,"publicationDate":"2024-08-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142095391","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":"Lipogenesis in Nasonia vitripennis: Influence of sugar chemistry, preferential production of triacylglycerides, and comparison of fatty acid biosynthetic capacity with Drosophila melanogaster","authors":"Joachim Ruther,&nbsp;Julian Hoheneder,&nbsp;Vera Koschany","doi":"10.1016/j.ibmb.2024.104179","DOIUrl":"10.1016/j.ibmb.2024.104179","url":null,"abstract":"<div><p>Sugar consumption increases the fecundity and longevity in many species of parasitic wasps (parasitoids) but whether these insects use sugars to synthesize significant amounts of fatty acids and storage fat de novo (lipogenesis) is discussed controversially. It has long been assumed that parasitic wasps lost this ability during evolution, mainly because in several species wasps with ad libitum access to sugar did not increase teneral lipid levels. Recent studies demonstrated that many species are nonetheless capable of synthesizing fatty acids de novo from glucose. It is unclear, however, whether also other sugars are used for fatty acid biosynthesis and whether an increase of sugar concentration to levels occurring in natural sugar sources translates into higher fatty acid production. Furthermore, it has been suggested that fatty acid production in parasitoids is negligible compared to species increasing teneral fat reserves such as <em>Drosophila melanogaster.</em> Here we show by stable isotope labeling experiments that females of <em>Nasonia vitripennis</em> convert D-glucose, D-fructose, sucrose, and α,α-trehalose, major sugars consumed by adult parasitoids in nature, equally well to palmitic, stearic, oleic, and linoleic acid. Lipogenesis from D-galactose occurs as well albeit to a lesser extent. Sugar concentration is crucial for lipogenic activity, and almost 80% of de novo synthesized fatty acids were incorporated into storage fat (triacylglycerides). Comparison of fatty acid biosynthesis within a 48-h feeding period with <em>D. melanogaster</em> revealed that <em>N. vitripennis</em> produced approximately half as many fatty acids per body mass unit. Both species fed equal amounts of the glucose offered. We conclude that lipogenesis is far from negligible in <em>N. vitripennis</em> and plays an important role for the energy balance when teneral lipid reserves deplete.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"173 ","pages":"Article 104179"},"PeriodicalIF":3.2,"publicationDate":"2024-08-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0965174824001103/pdfft?md5=d910e65536a49cc9d870ed05ed81e8d5&pid=1-s2.0-S0965174824001103-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142095390","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":"Ion transport peptide and ion transport peptide-like regulate ecdysis behavior and water transport during ecdysis in Gryllus bimaculatus","authors":"Zhen Zhu,&nbsp;Shinji Nagata","doi":"10.1016/j.ibmb.2024.104178","DOIUrl":"10.1016/j.ibmb.2024.104178","url":null,"abstract":"<div><p>Ion transport peptide (ITP) and ITP-like (ITPLs) are pleiotropic bioactive peptides in insects. Although the contribution of these peptides to ecdysis has been studied, the precise regulatory mechanisms remain poorly understood. Here, we characterized the functions of <em>itp</em> and <em>itpl</em> variants in the two-spotted cricket, <em>Gryllus bimaculatus</em>. Reverse transcription-quantitative PCR and whole-mount <em>in situ</em> hybridization revealed that <em>itp</em> was expressed in the brain and terminal abdominal ganglion, whereas <em>itpl</em> variants were expressed in all ganglia of the central nervous system. Simultaneous knockdown of <em>itp</em> and <em>itpls</em> disrupted ecdysis behavior and water transport from the gut into the hemolymph during molting. Nevertheless, knockdown of <em>itpls</em> without influencing <em>itp</em> expression did not significantly affect ecdysis behavior but caused a reduction in hemolymph mass. Although water transport into the hemolymph is considered necessary for the swelling required to split the old cuticle layers during molting, a rescue experiment by injection of water or cricket Ringer's solution into the hemolymph of knockdown crickets did not recover the normal phenotype. Therefore, we propose that ITP/ITPL control ecdysis behavior probably not by regulating water transport from the gut into the hemolymph in crickets.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"173 ","pages":"Article 104178"},"PeriodicalIF":3.2,"publicationDate":"2024-08-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0965174824001097/pdfft?md5=05eb373a08d09f2ceb6cce49d1146f88&pid=1-s2.0-S0965174824001097-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142071589","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":"Notch signaling promotes differentiation, cell death and autophagy in Drosophila hematopoietic system","authors":"Fangzhou Luo,&nbsp;Chengcheng Zhang,&nbsp;Zhengqi Shi,&nbsp;Tong Mao,&nbsp;Li Hua Jin","doi":"10.1016/j.ibmb.2024.104176","DOIUrl":"10.1016/j.ibmb.2024.104176","url":null,"abstract":"<div><p>Notch signaling is a highly conserved pathway between mammals and <em>Drosophila</em> and plays a key role in various biological processes. <em>Drosophila</em> has emerged as a powerful model for studying hematopoiesis and leukemia. In exception to crystal cells, the strength of Notch signaling in <em>Drosophila</em> lymph gland cortical zone (CZ)/intermediate zone (IZ) cells is weak. However, the influence of Notch activation in the lymph gland CZ/IZ cells and circulating hemocytes on hematopoietic homeostasis maintenance is unclear. Here, we showed that Notch activation in lymph gland CZ/IZ cells induced overdifferentiation of progenitors. Moreover, Notch activation promoted lamellocyte generation via NFκB/Toll signaling activation and increased reactive oxygen species (ROS). In addition, we found that Notch activation in lymph gland CZ/IZ cells and circulating hemocytes caused caspase-independent and nonautophagic cell death. However, crystal cell autophagy was activated by upregulation of the expression of the target gene of the Hippo/Yki pathway Diap1. Moreover, we showed that Notch activation could alleviate cytokine storms and improve the survival of Ras^v12 leukemia model flies. Our study revealed the various mechanisms of hematopoietic dysregulation induced by Notch activation in healthy flies and the therapeutic effect of Notch activation on leukemia model flies.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"173 ","pages":"Article 104176"},"PeriodicalIF":3.2,"publicationDate":"2024-08-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142015902","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":"Binding characteristics and structural dynamics of two general odorant-binding proteins with plant volatiles in the olfactory recognition of Glyphodes pyloalis","authors":"Yijiangcheng Li ,&nbsp;Wenmiao Song ,&nbsp;Shanshan Wang ,&nbsp;Wanglong Miao ,&nbsp;Zhixiang Liu ,&nbsp;Fuan Wu ,&nbsp;Jun Wang ,&nbsp;Sheng Sheng","doi":"10.1016/j.ibmb.2024.104177","DOIUrl":"10.1016/j.ibmb.2024.104177","url":null,"abstract":"<div><p><em>Glyphodes pyloalis</em> Walker (Lepidoptera: Pyralidae) is the most destructive pest, causing severe damage to mulberry production in China's sericulture industry. The insecticide application in mulberry orchards poses a significant risk of poisoning to <em>Bombyx mori</em>. Shifting from insecticides to odor attractants is a beneficial alternative, but not much data is available on the olfactory system of <em>G</em>. <em>pyloalis</em>. We identified 114 chemosensory genes from the antennal transcriptome database of <em>G</em>. <em>pyloalis</em>, with 18 <em>odorant-binding protein</em> (<em>OBP</em>) and 17 <em>chemosensory protein</em> (<em>CSP</em>) genes significantly expressed in the antennae. Ligand-binding assays for two antennae-biased expressed general odorant-binding proteins (GOBPs) showed high binding affinities of GOBP1 to hexadecanal, <em>β</em>-ionone, and 2-ethylhexyl acrylate, while GOBP2 exhibited binding to 4-tert-octylphenol, benzyl benzoate, <em>β</em>-ionone, and farnesol. Computational simulations indicated that van der Waal forces predominantly contributed to the binding free energy in the binding processes of complexes. Among them, Phe12 of GOBP1 and Phe19 of GOBP2 were demonstrated to play crucial roles in their bindings to plant volatiles using site-directed mutagenesis experiments. Moreover, hexadecanal and <em>β</em>-ionone attracted <em>G</em>. <em>pyloalis</em> male moths in the behavioral assays, while none of the candidate plant volatiles significantly affected female moths. Our findings provide a comprehensive understanding of the molecular mechanisms underlying olfactory recognition in <em>G</em>. <em>pyloalis</em>, setting the groundwork for novel mulberry pests control strategies based on insect olfaction.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"173 ","pages":"Article 104177"},"PeriodicalIF":3.2,"publicationDate":"2024-08-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142034790","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 the activation mechanism of Bm-CPA: Implications for insect molting regulation","authors":"Lingzhen Yang ,&nbsp;Yuejing Cheng ,&nbsp;Qinglang Wang,&nbsp;Jianing Hou,&nbsp;Qingyu Rong,&nbsp;Chunxia Xiao,&nbsp;Yuhao Zhang,&nbsp;Jiamin Yan,&nbsp;Qingyou Xia,&nbsp;Yong Hou","doi":"10.1016/j.ibmb.2024.104175","DOIUrl":"10.1016/j.ibmb.2024.104175","url":null,"abstract":"<div><p>Carboxypeptidase A has been found across various animal species, yet its activation mechanism during the insect molting process remains elusive. Our study specifically delved into the activation mechanism of carboxypeptidase A (Bm-CPA), identified in <em>Bombyx mori</em>'s molting fluid during metamorphosis. Initially, western blotting identified two forms of Bm-CPA, 65 kDa and 54 kDa, in the epidermis of silkworms during the molting stage. Expressing the complete Bm-CPA sequence in <em>Pichia pastoris</em> allowed the identification, via mass spectrometry analysis, of a 75-amino-acid propeptide for the initial hydrolysis process. Subsequently, a 35 kDa form of Bm-CPA emerged in the molting fluid, confirmed as the active form through <em>in vitro</em> assays, demonstrating potent carboxypeptidase A activity and faint carboxypeptidase B activity. Four potential activation sites (including Lys158/Arg159 and Arg177/Arg178) were identified through mass spectrometry and amino acid mutation analysis. RNAi of Bm-CPA indicates its critical role in molting. Finally, the carboxypeptidase inhibitor (Bm-CPI) from silkworm molting fluid was expressed to explore its role in regulating Bm-CPA activity, demonstrating a direct interaction with the 35 kDa Bm-CPA. Our research implies Bm-CPA's potential involvement in the silkworm molting process, suggesting diverse regulatory roles. These findings highlight intricate protein regulation patterns during insect metamorphosis and development.</p></div>","PeriodicalId":330,"journal":{"name":"Insect Biochemistry and Molecular Biology","volume":"173 ","pages":"Article 104175"},"PeriodicalIF":3.2,"publicationDate":"2024-08-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141970301","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}