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昆虫分子生物与功能基因Insect molecular biology

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For Selected: Download Citations EndNote Ris BibTeX Toggle Thumbnails Select Vesicular transport-related genes in Diaphorina citri are involved in the process of Candidatus Liberibacter asiaticus infection Yingzhe Yuan, Tao Peng, Aijun Huang, Jun He, Chenyang Yuan, Tianyuan Liu, Long Yi, Xuejin Cui, Xuefeng Wang, Changyong Zhou 2024, 23 (12): 4136-4146.   DOI: 10.1016/j.jia.2024.03.063 Abstract （81）      PDF in ScienceDirect       Asian citrus psyllid (ACP, Diaphorina citri) is the major vector of Candidatus Liberibacter asiaticus (CLas), which is a bacterial pathogen causing the devastating citrus Huanglongbing (HLB) disease.  Diaphorina citri is known to carry CLas in a persistent and propagative manner.  Some studies have suggested that CLas may use the vesicular structures of D. citri cells as its propagation organelles.  However, the mechanisms by which CLas enters the D. citri cells and how vesicle-mediated trafficking is involved remain unclear.  In this study, we monitored the titer change of CLas in D. citri nymphs during the process of CLas acquisition from feeding on infected citrus plants.  We found that the titer of CLas increased with the acquisition access period.  After infection, there was a significant upregulation in the expression of several vesicular transport-related genes in D. citri.  The titer of CLas was significantly reduced in the midgut and whole insect body when endocytosis and the endosome network in D. citri were inhibited.  Furthermore, silencing the D. citri clathrin-heavy chain gene also led to a reduction in the CLas titer in D. citri.  These results suggest that CLas infection upregulates the genes related to vesicular transport in D. citri, which facilitates the invasion of endocytosis-dependent pathogens. Reference | Related Articles | Metrics Select Functional characterization of sensory neuron membrane protein 1a involved in sex pheromone detection of Apolygus lucorum (Hemiptera: Miridae) Yan Li, Xingkui An, Shuang Shan, Xiaoqian Pang, Xiaohe Liu, Yang Sun, Adel Khashaveh, Yongjun Zhang 2024, 23 (12): 4120-4135.   DOI: 10.1016/j.jia.2024.03.043 Abstract （116）      PDF in ScienceDirect       The mirid bug Apolygus lucorum (Hemiptera: Miridae) is a polyphagous pest that affects a wide range of host plants.  Its control remains challenging mainly due to its rapid reproduction, necessitating an understanding of sex pheromone communication.  The recognition of sex pheromones is vital for courtship and mating behaviors, and is mediated by various chemosensory-associated proteins.  Among these, sensory neuron membrane protein (SNMP), a CD36-related protein, is suggested to play crucial roles in detecting sex pheromones.  In this study, we employed transcriptomic and genomic data from A. lucorum and phylogenetic approaches, and identified four putative SNMP genes (AlucSNMP1a, AlucSNMP1b, AlucSNMP2a, and AlucSNMP2b) with full open reading frames.  Expression analysis revealed the ubiquitous presence of AlucSNMP transcripts in multiple tissues, with only AlucSNMP1a exhibiting male-biased expression in the antennae, suggesting its potential role in male chemosensation.  Functional analysis using the Xenopus oocyte expression system, coupled with two-electrode voltage clamp recording, demonstrated that the co-expression of AlucSNMP1a with specific pheromone receptors (PRs) and the Odorant receptor co-receptor (Orco) significantly enhanced electrophysiological responses to sex pheromones compared to the co-expression of PRs and Orco alone.  Moreover, the results indicated that the presence of AlucSNMP1a not only affected the responsiveness to sex pheromones but also influenced the kinetics (activation and inactivation) of the induced signals.  In contrast, the co-expression of AlucSNMP1b with AlucPR/Orco complexes had no impact on the inward currents induced by two pheromone compounds.  An examination of the selective pressures on SNMP1 genes across 20 species indicated strong purifying selection, implying potential functional conservation in various insects.  These findings highlight the crucial role of AlucSNMP1a in the response to sex pheromones. Reference | Related Articles | Metrics Select Glomerular organization in the antennal lobe of the oriental armyworm Mythimna separata Baiwei Ma, Qi Chen, Xi Chu, Yidong Zuo, Jiayu Wang, Yi Yang, Guirong Wang, Bingzhong Ren 2024, 23 (11): 3812-3829.   DOI: 10.1016/j.jia.2024.04.030 Abstract （86）      PDF in ScienceDirect       In insects, the number and specificity of antennal lobe glomeruli often reflect the number and diversity of expressed chemosensory genes, which are linked to its ecological niche and specific olfactory needs.  The oriental armyworm, Mythimna separata, is an important and common lepidopteran pest of cereal crops.  Given its reliance on the olfactory system for crucial behaviors, understanding the evolutionary potential of this system requires a thorough characterization of the anatomical structure of the primary olfactory center.  Here, we systematically identified all antennal lobe glomeruli of M. separata based on synaptic antibody immunostaining and mass staining of the olfactory sensory neurons.  A total of 69 glomeruli were identified in females and 65 in males, and an intuitive nomenclature based on glomerular positions was applied.  Our findings uncovered some sex-specific glomeruli in this species.  There were ten female-specific glomeruli and three male-specific glomeruli, except for the macroglomerular complex (MGC) units, with a notable observation that the female labial pit organ glomerulus was larger than its male counterpart.  Additionally, we identified four antennal-lobe tracts (ALTs) and retrograde labeling from the calyx revealed that all glomeruli were innervated by the medial ALT projection neurons.  The comparison of the olfactory system structures between M. separata and sympatric moths supports their evolutionary convergence in noctuid moths.  These results collectively lay the foundation for future studies on olfactory processing in M. separata. Reference | Related Articles | Metrics Select Knock-in of exogenous sequences based on CRISPR/Cas9 targeting autosomal genes and sex chromosomes in the diamondback moth, Plutella xylostella Shanyu Li, Guifang Lin, Haoqi Wen, Haiyan Lu, Anyuan Yin, Chanqin Zheng, Feifei Li, Qingxuan Qiao, Lu Jiao, Ling Lin, Yi Yan, Xiujuan Xiang, Huang Liao, Huiting Feng, Yussuf Mohamed Salum, Minsheng You, Wei Chen, Weiyi He 2024, 23 (9): 3089-3103.   DOI: 10.1016/j.jia.2024.04.029 Abstract （98）      PDF in ScienceDirect       Genetic pest control strategies based on precise sex separation and only releasing sterile males can be accomplished by site-specific genome editing.  In the current study, we showed that the mutation of single-allele Pxfl(2)d can significantly impair the normal mating behavior and testis development in male adults of the notorious cruciferous insect pest Plutella xylostella, in addition to its known functions in the ovarian development in female adults and egg hatching.  Subsequent CRISPR/Cas9-based knock-in experiments revealed that site-specific integration of an exogenous green fluorescent protein (GFP) gene into autosomal Pxfl(2)d for labelling mutants could be achieved.  However, this gene is not a suitable target for GFP insertion to establish a genetically stable knock-in strain because of the severe decline in reproductive capacity.  We further screened for the W-chromosome-linked and Z-chromosome-linked regions to test the knock-in efficiency mediated by CRISPR/Cas9.  The results verified that both types of chromosomes can be targeted for the site-specific insertion of exogenous sequences.  We ultimately obtained a homozygous knock-in strain with the integration of both Cas9 and cyan fluorescent protein (CFP) expression cassettes on a Z-linked region in P. xylostella, which can also be used for early sex detection.  By injecting the sgRNA targeting Pxfl(2)d alone into the eggs laid by female adults of the Z-Cas9-CFP strain, the gene editing efficiency reached 29.73%, confirming the success of expressing a functional Cas9 gene.  Taken together, we demonstrated the feasibility of the knock-in of an exogenous gene to different genomic regions in P. xylostella, while the establishment of a heritable strain required the positioning of appropriate sites.  This study provides an important working basis and technical support for further developing genetic strategies for insect pest control. Reference | Related Articles | Metrics Select The Ca2+/CaN/ACC and cAMP/PKA/HK signal pathways are required for PBAN-mediated sex pheromone biosynthesis in Conogethes punctiferalis Yao Zhang, Zelong She, Ruolan He, Shuangyan Yao, Xiang Li, Xiaoguang Liu, Xinming Yin, Jizhen Wei, Mengfang Du, Shiheng An 2024, 23 (8): 2735-2751.   DOI: 10.1016/j.jia.2023.09.001 Abstract （107）      PDF in ScienceDirect       Conogethes punctiferalis is a crop and fruit pest that has caused serious economic losses to agricultural production.  This pest relies heavily on its sex pheromone to ensure sexual encounters and subsequent mating success.  However, the molecular mechanism underlying sex pheromone biosynthesis in this species remains elusive.  The present study investigated the detailed mechanism underlying PBAN-regulated sex pheromone biosynthesis in C. punctiferalis by transcriptome sequencing of the C. punctiferalis pheromone glands (PGs) and subsequent functional identification of the target genes.  The results showed that female mating started from the first scotophase, and peaked at the second to fifth scotophases in accordance with the release of sex pheromones.  PBAN regulated sex pheromone biosynthesis by employing Ca2+ and cAMP as secondary messengers, as demonstrated by RNA interference (RNAi), pharmacological inhibitors, and behavioral assays.  Further investigation revealed that calcineurin (CaN) and acetyl-CoA carboxylase (ACC) were activated by PBAN/Ca2+ signaling, and the RNAi-mediated knockdown of CaN and ACC transcripts significantly reduced sex pheromone production, ultimately leading to a significantly reduced ability of females to attract males.  Importantly, hexokinase (HK) was found to regulate sex pheromone biosynthesis in response to the PBAN/cAMP/PKA signaling pathway, as demonstrated by RNAi, enzyme activity, and pharmacological inhibitor assays.  Furthermore, Far2 and Desaturase1 were found to participate in PBAN-regulated sex pheromone biosynthesis.  Altogether, our findings revealed that PBAN regulates sex pheromone biosynthesis through the PBANR/Ca2+/CaN/ACC and PBANR/cAMP/PKA/HK pathways in C. punctiferalis, which enriches our comprehension of the details of sex pheromone biosynthesis in moths. Reference | Related Articles | Metrics Select Identification of transient receptor potential channel genes and functional characterization of TRPA1 in Spodoptera frugiperda  Yutong Zhang, Hangwei Liu, Song Cao, Bin Li, Yang Liu, Guirong Wang 2024, 23 (6): 1994-2005.   DOI: 10.1016/j.jia.2023.09.023 Abstract （126）      PDF in ScienceDirect       Spodoptera frugiperda is a highly destructive pest that has become a global problem due to its robust reproductive and migratory capabilities.  Transient receptor potential (TRP) channels, which constitute a vast ion channel family, play pivotal roles in sensing the external environment and maintaining internal homeostasis in insects.  TRP channels have been widely investigated for their critical roles in regulating various insect behaviors in recent years.  In this study, we identified 15 TRP gene loci encoding 26 transcripts in the genome of S. frugiperda and analyzed their expression profiles at different developmental stages.  The results revealed that S. frugiperda possesses four TRPC genes, six TRPA genes, one TRPM gene, two TRPV genes, one TRPN gene, and one TRPML gene, while a canonical TRPP is absent.  Moreover, the SfruTRPA1 was functionally characterized using the Xenopus oocyte expression system.  The results showed that SfruTRPA1 is activated by temperature increases from 20 to 45°C, and there is no significant desensitization after repeated stimuli within the same temperature range.  Additionally, SfruTRPA1 is activated by certain natural chemicals, including allyl isothiocyanate (AITC) and cinnamaldehyde (CA).  These findings provide valuable insights to the TRP genes in S. frugiperda. Reference | Related Articles | Metrics Select CRISPR/Cas9-mediated NlInR2 mutants: Analyses of residual mRNA and truncated proteins Jun Lü, Jingxiang Chen, Yutao Hu , Lin Chen, Shihui Li, Yibing Zhang, Wenqing Zhang 2024, 23 (6): 2006-2017.   DOI: 10.1016/j.jia.2023.06.039 Abstract （53）      PDF in ScienceDirect       CRISPR/Cas9 technology is a powerful genome manipulation tool in insects.  However, little is known about whether mRNA and protein of a target gene are completely cleared in homozygous mutants.  This study generated homozygous mutants of the insulin receptor gene 2 (NlInR2) in the brown planthopper (Nilaparvata lugens) using CRISPR/Cas9 genome editing.  Both frameshift mutants, E5_D17 and E6_I7, differentiated towards long wings, but there were differences in wing morphology, with E5_D17 showing wing deformities.  Subsequent investigations revealed the presence of residual expression of NlInR2 mRNA in both mutants, as well as the occurrence of spliceosomes featuring exon skipping splicing in E5_D17.  Additionally, the E5_D17 exhibited the detection of N-terminally truncated NlInR2 protein.  RNA interference experiments indicated that the knockdown of NlInR2 expression in the E5_D17 mutant line increased the proportion of wing deformities from 11.1 to 65.6%, suggesting that the residual NlInR2 mRNA of the E5_D17 mutant might have retained some genetic functions.  Our results imply that systematic characterization of residual protein expression or function in CRISPR/Cas9-generated mutant lines is necessary for phenotypic interpretation. Reference | Related Articles | Metrics Select Functional assessment of cadherin as a shared mechanism for cross/dual resistance to Cry1Ac and Cry2Ab in Helicoverpa zea Jizhen Wei, Min Zhang, Pin Li, Zhongyuan Deng, Xinming Yin, Shiheng An, Xianchun Li 2024, 23 (5): 1604-1617.   DOI: 10.1016/j.jia.2023.11.001 Abstract （100）      PDF in ScienceDirect       Helicoverpa zea is a major target pest of pyramided transgenic crops expressing Cry1, Cry2 and/or Vip3Aa proteins from Bacillus thuringiensis (Bt) in the United States.  Laboratory-selected Cry1Ac/Cry2Ab cross resistance and field-evolved practical dual resistance of H. zea to these two toxins have been widely reported.  Whether the widespread Cry1Ac/Cy2Ab dual resistance of H. zea has resulted from the selection of one shared or two independent resistance mechanisms by pyramided Bt crops remains unclear.  Cadherin is a well-confirmed receptor of Cry1Ac and a suggested receptor of Cry2Ab in at least three Lepidopteran species.  To test whether cadherin may serve as one shared mechanism for the cross and dual resistance of H. zea to Cry1Ac and Cry2Ab, we cloned H. zea cadherin (HzCadherin) cDNA and studied its functional roles in the mode of action of Cry1Ac and Cry2Ab by gain- and loss-of-function analyses.  Heterologous expression of HzCadherin in H. zea midgut, H. zea fat body and Sf9 cells made all three of these cell lines more susceptible to activated Cry1Ac but not activated Cry2Ab, whereas silencing HzCadherin of H. zea midgut and fat body cells significantly reduced the susceptibility to Cry1Ac but not Cry2Ab.  Likewise, suppressing HzCadherin with siRNA made H. zea larvae resistant to Cry1Ac.  These results clearly demonstrate that HzCadherin is not a receptor for Cry2Ab, and thus it is unlikely to serve as one shared mechanism for the cross and dual resistance of H. zea to Cry1Ac and Cry2Ab. Reference | Related Articles | Metrics Select Host-induced silencing of MpPar6 confers Myzus persicae resistance in transgenic rape plants Qi Zhang, Wenqin Zhan, Chao Li, Ling Chang, Yi Dong, Jiang Zhang 2024, 23 (1): 187-194.   DOI: 10.1016/j.jia.2023.05.027 Abstract （177）      PDF in ScienceDirect       Plant-mediated RNA interference (RNAi) has emerged as a promising technology for insect control.  The green peach aphid, Myzus persicae, feeds on over 400 species of host plants.  Brassica napus (rape) is the second most important oilseed crop worldwide.  Myzus persicae is highly reproductive and causes severe damage to the rape plants due to its quite flexible life cycle.  In this study, we tested the RNAi effects of transgenic rape plants on M. persicae.  By in vitro feeding M. persicae with artificial diets containing double-stranded RNAs (dsRNAs) targeting seven aphid genes, we identified a new gene encoding the partitioning-defective protein 6 (Par6) as the most potent RNAi target.  Tissue- and stage-expression analysis of Par6 suggested this gene is highly expressed in the embryo and adult stage of M. persicae.  We next generated transgenic rape plants expressing dsPar6 by Agrobacterium-mediated transformation and obtained nine independent transgenic lines.  Compared to wild-type control plants, transgenic rape lines expressing dsPar6 showed strong resistance to M. persicae.  Feeding assays revealed that feeding transgenic rape plants to M. persicae significantly decreased MpPar6 expression and survival rate and impaired fecundity.  Furthermore, we showed that the resistance levels to M. persicae are positively correlated with dsPar6 expression levels in transgenic rape plants.  Our study demonstrates that transgenic rape plants expressing dsPar6 are efficiently protected from M. persicae.  Interfering with the genes involved in embryo development could be the effective RNAi targets for controlling aphids and potentially other insect pests. Reference | Related Articles | Metrics Select Mutagenesis of odorant coreceptor Orco reveals the distinct role of olfaction between sexes in Spodoptera frugiperda CAO Song, SUN Dong-dong, LIU Yang, YANG Qing, WANG Gui-rong 2023, 22 (7): 2162-2172.   DOI: 10.1016/j.jia.2022.11.004 Abstract （192）      PDF in ScienceDirect       Odorant receptor (OR) is crucial for insects to detect and recognize external chemical cues closely related to their survival.  The insect OR forms a heteromeric complex composed of a ligand-specific receptor and a ubiquitously odorant receptor coreceptor (Orco).  This study used the CRISPR/Cas9 technique to knock out (KO) Orco and reveal its essential role in acting on OR-meditated olfactory behavior in a critical invasive agricultural pest, the fall armyworm (FAW), Spodoptera frugiperda.  Electroantennogram (EAG) results suggested that the Orco mutants of both male and female moths severely reduced their electrophysiological responses to the eight tested plant volatiles and two sex pheromones.  However, the Orco gene played distinct roles in mating behavior between sexes: the mating behavior was fully disrupted in mutated males but not in mutated females.  The oviposition result indicated that the Orco KO females displayed reduced egg laying by 24.1% compared with the mated wild type (WT) females.  Overall, these results strongly suggest that Orco is an excellent target for disrupting FAW’s normal behavior and provides a feasible pest control approach. Reference | Related Articles | Metrics Select CRISPR-based genetic control strategies for insect pests Ying YAN, Roswitha A. AUMANN, Irina HÄCKER, Marc F. SCHETELIG 2023, 22 (3): 651-668.   DOI: 10.1016/j.jia.2022.11.003 Abstract （265）      PDF in ScienceDirect       Genetic control strategies such as the sterile insect technique have successfully fought insect pests worldwide.  The CRISPR (clustered regularly interspaced short palindromic repeats) technology, together with high-quality genomic resources obtained in more and more species, greatly facilitates the development of novel genetic control insect strains that can be used in area-wide and species-specific pest control programs.  Here, we review the research progress towards state-of-art CRISPR-based genetic control strategies, including gene drive, sex ratio distortion, CRISPR-engineered genetic sexing strains, and precision-guided sterile insect technique.  These strategies’ working mechanisms, potential resistance development mechanisms, and regulations are illustrated and discussed.  In addition, recent developments such as stacked and conditional systems are introduced.  We envision that the advances in genetic technology will continue to be one of the driving forces for developing the next generation of pest control strategies.   Reference | Related Articles | Metrics Select Chromosome-level genome assembly of Cylas formicarius provides insights into its adaptation and invasion mechanisms HUA Jin-feng, ZHANG Lei, HAN Yong-hua, GOU Xiao-wan, CHEN Tian-yuan, HUANG Yong-mei, LI Yan-qing, MA Dai-fu, LI Zong-yun 2023, 22 (3): 825-843.   DOI: 10.1016/j.jia.2022.08.027 Abstract （265）      PDF in ScienceDirect       Cylas formicarius is one of the most important pests of sweet potato worldwide, causing considerable ecological and economic damage.  This study improved the effect of comprehensive management and understanding of genetic mechanisms by examining the functional genomics of C. formicarius.  Using Illumina and PacBio sequencing, this study obtained a chromosome-level genome assembly of adult weevils from lines inbred for 15 generations.  The high-quality assembly obtained was 338.84 Mb, with contig and scaffold N50 values of 14.97 and 34.23 Mb, respectively.  In total, 157.51 Mb of repeat sequences and 11 907 protein-coding genes were predicted.  A total of 337.06 Mb of genomic sequences was located on the 11 chromosomes, accounting for 99.03% of the total length of the associated chromosome.  Comparative genomic analysis showed that C. formicarius was sister to Dendroctonus ponderosae, and C. formicarius diverged from D. ponderosae approximately 138.89 million years ago (Mya).  Many important gene families expanded in the C. formicarius genome were involved in the detoxification of pesticides, tolerance to cold stress and chemosensory system.  To further study the role of odorant-binding proteins (OBPs) in olfactory recognition of C. formicarius, the binding assay results indicated that CforOBP4–6 had strong binding affinities for sex pheromones and other ligands.  The high-quality C. formicarius genome provides a valuable resource to reveal the molecular ecological basis, genetic mechanism, and evolutionary process of major agricultural pests; it also offers new ideas and new technologies for ecologically sustainable pest control. Reference | Related Articles | Metrics Select Characterization of the chemosensory protein EforCSP3 and its potential involvement in host location by Encarsia formosa WANG Ke, HE Yan-yan, ZHANG You-jun, GUO Zhao-jiang, XIE Wen, WU Qing-jun, WANG Shao-li 2023, 22 (2): 514-525.   DOI: 10.1016/j.jia.2022.08.015 Abstract （199）      PDF in ScienceDirect       Chemosensory proteins (CSPs) perform several functions in insects.  This study performed the gene expression, ligand-binding, and molecular docking assays on the EforCSP3 identified in the parasitoid wasp Encarsia formosa, to determine whether EforCSP3 functions in olfaction, especially in host location and host preference.  The results showed that EforCSP3 was highly expressed in the female head, and its relative expression was much higher in adults than in other developmental stages.  The fluorescence binding assays suggested that the EforCSP3 exhibited high binding affinities to a wide range of host-related volatiles, among which dibutyl phthalate, 1-octene, β-elemene, and tridecane had the strongest binding affinity with EforCSP3, besides α-humulene and β-myrcene, and should be assessed as potential attractants.  Protein structure modeling and molecular docking predicted the amino acid residues of EforCSP3 possibly involved in volatile binding.  α-Humulene and β-myrcene attracted E. formosa in a previous study and exhibited strong binding affinities with EforCSP3 in the current study.  In conclusion, EforCSP3 may be involved in semiochemical reception by E. formosa. 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