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Dynamic transcriptome analysis of Bombyx mori embryonic development

Insect Science 2022年第2期29卷 344-362页

作者： Guan-Feng Xu Cheng-Cheng Gong Hao Lyu Hui-Min Deng si-chun zheng Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology Institute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhou510631 China Guangzhou Key Laboratory of Insect Development Regulation and Applied Research Institute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhou510631 China

Bombyx mori has been extensively studied but the gene expression control of its embryonic development is *** this study,we performed transcriptome profiling of six stages of *** embryonic development using RNA sequencing(RNA-seq).A total of 12894 transcripts were obtained from the *** these,12456 transcripts were shared among the six stages,namely,fertilized egg,blastoderm,germ-band,organogenesis,reversal period,and youth period *** were 111,48,41,54,77,and 107 transcripts specifically expressed during the six stages,*** analyzing weighted gene correlation networks and differently expressed genes,we found that during embryonic development,many genes related to DNA replication,transcription,protein synthesis,and epigenetic modifications were upregulated in the early *** of cuticle proteins,chitin synthesis-related proteins,and neuropeptides were more abundant in the late *** pathways of juvenile hormone and the ecdysteroid 20-hydroxyecdysone,and transcription factors were expressed throughout the embryonic development stages,more regulatory pathways were highly expressed around the organogenesis stage,suggesting more gene expression for *** results of RNA-seq were confirmed by quantitative real-time polymerase chain reaction of 16 genes of different *** acid methylation and seven sites in histone H3 modifications were confirmed by dot blot and western *** study increases the understanding of the molecular mechanisms of the embryonic developmental process and information on the regulation of *** development.

关键词： Bombyx mori development embryo RNA-seq

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Genomic divergences between the two polyphagous Spodoptera relatives provide cues for successful invasion of the fall armyworm

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Insect Science 2020年第6期27卷 1257-1265页

作者： Yong Cui Yan-Dong Ren Mo Lyu si-chun zheng Qi-Li Feng Hui Xiang Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology Guangzhou Key Laboratory of Insect Development Regulation and Application ResearchInstitute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina Center for Ecological and Environmental Sciences Northwestern Polytechnical UniversityXi’anChina Guangzhou Panyu Central Hospital GuangzhouChina

The fall armyworm Spodoptera frugiperda recently invaded China,ravaging crops in many *** the possible genetic basics for its successful invasion is critical for innovative and specific control for this gluttonous *** we generated comparative genomic analyses between *** and its native relative,***,which differs in host preference,locomotivity and production *** demonstrated that *** genes are enriched in taste sensory perception and nervous system,obv iously difTerent from those of *** host adaptation genes showed generally an elevated ratio of non-synonymous substitution rate to synonymous substitution rate,suggesting a faster evolution during the divergence of the two *** on these sets of genes,we identified 23 genes being under positive selection in *** them are two notable genes involved in sensory perception,gustatory receptor(GR)and an acetaldehyde oxidase,which are important for host detection in invasion and expansion *** two genes are mitochondrial adenosine triphosphate synthase^subunit and ferritin heavy chain,which may be associated with the enhanced locomotivity and resistance,which fascinated long-distance migration needed for invasion and rapid *** interesting gene is chorion protein,in which positive selection sites in *** were found and a replacement in one site is predicted to affect the protein function,which might be associated with competent reproductivity in *** to ensure genetic resources for expansion.

关键词： chorion protein invasion positive selection sensory perception Spodoptera frugiperda Spodoptera litura

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signal transducer and activator of transcription is involved in the expression regulation of ecdysteroid-induced insulin-like growth factor-like peptide in the pupal wing disc of silkworm, Bombyx mori

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Insect Science 2020年第6期27卷 1186-1197页

作者： Xiao-Juan Zhang Dong-Dong Li Guan-Feng Xu Ya-Qing Chen si-chun zheng Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology Institute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina Guangzhou Key Laboratory of Insect Development Regulation and Applied Research Institute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina

In insects,20-hydroxyecdysone(20E)and insulin-like growth factor-like peptides(IGFLPs)regulate the development o f imaginal ***,how IGFLPs are up-regulated to impact the development of the pupal wing disc is still *** this study,we investigated the expression regulation of IGFLP in the pupal wing disc of silkworm,Bombyx *** confirmed that *** IGFLP(BmIGFLP)was mainly expressed in the pupal wing disc and the expression of BmIGFLP could be significantly induced by *** analysis of BmIGFLP promoter sequence revealed three m-regulation elements(CREs)of signal transducer and activator of transcription(STAT),which is a key component in the Janus-activated kinase/STAT *** activity assays showed that two CREs enhanced the transcriptional activity o f *** mobility shift and chromatin immunoprecipitation assays demonstrated that BmSTAT proteins in the nuclear extracts o f *** pupal wing discs and BmN cells could only bind to the STAT CRE3,indicating that STAT CRE3 activated by BmSTAT enhances BmIGFLP expression at pupal *** 20E could not enhance the expression of BmSTAT,20E enhanced the nucleus translocation o f BmSTAT to bind with the STAT CRE3 in the BmIGFLP *** increase of transcriptional activity of the STAT CRE3 by overexpression of BmSTAT and addition o f 20E in BmN cells confirmed this *** together,all data indicate that BmSTAT is one o f the transcription factors activating 20E-induced BmIGFLP expression in the pupal wing disc.

关键词： Bombyx mori expression regulation IGFLP 20-hydroxyecdysone STAT

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Detoxification of insecticides, allechemicals and heavy metals by glutathione S-transferase siGSTE1 in the gut of Spodoptera litura

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Insect Science 2015年第4期22卷 503-511页

作者： Zhi-Bin Xu Xiao-Peng Zou Ni Zhang Qi-Li Feng si-chun zheng Laboratory of Molecular and Developmental Entomology Guangdong Provincial Key Laboratory of Biotechnology for Plant Development School of Life Sciences South China Normal University Guangzhou 510631 China

Insect glutathione S-transferases （GSTs） play important roles in detoxifying toxic compounds and eliminating oxidative stress caused by these compounds. In this study, detoxification activity of the epsilon GST S1GSTE1 in Spodoptera litura was analyzed for several insecticides and heavy metals. S1GSTE1 was significantly up-regulated by chlor- pyrifos and xanthotoxin in the midgut of S. litura. The recombinant S1GSTE1 had Vmax （reaction rate of the enzyme saturated with the substrate） and Km （michaelis constant and equals to the substrate concentration at half of the maximum reaction rate of the enzyme） values of 27.95 ± 0.88μmol/min/mg and 0.87 ± 0.028 mmol/L for glutathione, respec- tively, and Vmax and Km values of 22.96 ± 0,78μmol/min/mg and 0.83 ± 0.106 mmol/L for 1-chloro-2,4-dinitrobenzene, respectively. In vitro enzyme indirect activity assay showed that the recombinant S1GSTE 1 possessed high binding activities to the insecticides chlor- pyrifos, deltamethrin, malathion, phoxim and dichloro-diphenyl-trichloroethane （DDT）. S1GSTE1 showed higher binding activity to toxic heavy metals cadmium, chromium and lead than copper and zinc that are required for insect normal growth. Western blot analysis showed that S1GSTE1 was induced in the gut of larvae fed with chlorpyrifos or cadmium. S1GSTE1 also showed high peroxidase activity. All the results together indicate that S1G- STE1 may play an important role in the gut orS. litura to protect the insect from the toxic effects of these compounds and heavy metals.

关键词： glutathioneS-transferase heavy metal insecticide midgut Spodoptera litura

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Transcriptomic analysis reveals the regulation network of BmKruppel homolog 1 in the oocyte development of Bombyx mori

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Insect Science 2021年第1期28卷 47-62页

作者： Zi-Dan Zhu Qi-Hao Hu chun-Mei Tong Hong-Guang Yang si-chun zheng Qi-Li Feng Hui-Min Deng Guangdong Key Laboratory of Insect Developmental Biology and Applied Technology Guangzhou Key Laboratory of Insect Development Regulation and Application ResearchInstitute of Insect Science and Technology&School of Life SciencesSouth China Normal UniversityGuangzhouChina

Kriippel homolog 1(Kr-hl),a zinc finger transcription factor,is involved in the metamorphosis and adult reproduction of ***,the role of Kr-hl in reproduction of holometabolic insects remains to be *** regulation network of 7-associated genes in the reproduction in Bombyx mori was investigated in this *** higher expression level of BmKr-hl in the ovaries was detected during the late pupal stage and *** interference(RNAi)-mediated depletion of BmKr-hl in the female at day 6 of pupae resulted in abnormal oocytes at 48 h post-double-stranded RNA treatment,which showed less yolk protein deposition and partially transparent ***-seq and subsequent differentially expressed transcripts analysis showed that knockdown of BmKr-hl caused a decrease in the expression of 2882 genes and an increase in the expression of 2565 genes in the oocytes at day 8 of ***,27 genes coding for transcription factors were down-regulated,while six genes coding for other transcription factors were ***-hl bound to the Kr-hl binding site of the transcription factors AP-1(activating protein-1)and FOXG1 to increase their messenger RNA tran・scripts in the BmN cells,*** Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analyses of that positively co-expressed with AP-1 and FOXG1 transcripts showed mainly enrichment in the metabolic-related pathways,the nu・tri ent absorption and the yolk protein absorption *** data suggested that BmKr-h 1 might directly regulate the metabolic-related pathways,the nutrient absorption and the yolk protein absorption processes or probably through AP-1 and/or FOXG1 to regulate oocyte development.

关键词： Kriippel homolog 1 oocyte development regulatory network RNA interference transcriptomics

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Cloning, expression and localization of a trypsin-like serine protease in the spruce budworm, Choristoneura fumiferana

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Insect Science 2009年第6期16卷 455-464页

作者： Wen-Ying He Yi-Ping zheng Lin Tang si-chun zheng Catherine Beliveau Daniel Doucet Michel Cusson Qi-Li Feng Guangdong Provincial Key Laboratory of Biotechnology for Plant Development School of Life Sciences South China Normal University Guangzhou China Great Lakes Forestry Center Sault Ste Marie Laurentian Forestry Centre Canadian Forest Service Quebec QC Canada

A trypsin-like molting-related serine protease cDNA （CfMRSP） was cloned from the spruce budworm, Choristoneurafumiferana. The full-length CfMRSP complementary DNA （cDNA） encoded a 43 kDa protein that contained a trypsin-like serine protease catalytic domain, but no clip domain. The C-terminal extension contained five cystein residues, which may allow the protein to form a homodimer through interchain disulfide bonds and regulate the activity of CfMRSE Phylogenetic tree analysis showed that CfMRSP clusters with lepidopteran homologues such as serine protease 1 of Lonomia obliqua, hemolymph proteinase 20 （HP20）, pattern recognition serine proteinase precursor （ProHP14） and a trypsin-like protein ofManduca sexta. Northern blot analysis of developmental expression of CfMRSP indicated that its transcripts were found primarily in the epidermis and were produced during all of the tested stadia, from 4th instar larvae to pupae, but increased levels of CfMRSP transcripts were always found after each molt. A high level of the protein was found in the epidermis by immunohistochemistry analysis. Altogether these data suggest that CfMRSP plays a role in the epidermis during molting and metamorphosis.

关键词： Choristoneurafumiferana cuticle metamorphosis molt serine protease

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Expression profiles of glutathione S-transferase superfamily in Spodoptera litura tolerated to sublethal doses of chlorpyrifos

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Insect Science 2016年第5期23卷 675-687页

作者： Ni Zhang Jia Liu Shu-Na Chen Li-Hua Huang Qi-Li Feng si-chun zheng Laboratory of Developmental and Molecular Entomology Guangdong Provincial Key Laboratory of Biotechnology for Plant DevelopmentSchool of Life Sciences South China Normal University Guangzhou China

Chlorpyrifos （CPF） is a broad-spectrum organophosphate insecticide. Glu- tathione S-transferases （GSTs） in insects are a family of detoxification enzymes and they play critical roles in CPF detoxification. Spodoptera litura is one of the most destructive agricultural pests in tropical and subtropical areas in the world. In this study, 37 Slgsts from 46 unique transcripts of gsts in S. litura transcriptome data, including eight pre- viously reported GSTs, were identified and their expression patterns in susceptible and 12-generation-CPF-treated strains were analyzed to understand the roles of these Slgsts in sublethal doses of CPF tolerance. The results indicate that the members of the S. litura GST superfamily could be distinguished into three major groups： one group, includ- ing six cytosolic Slgsts （siGSTel, siGSTe3, siGSTelO, siGSTe15, siGSTo2 and siGSTs5） and two microsomal Slgsts （siMGSTI-2 and siMGST1-3）, was directly responsible for CPF induction in both 12-generation-treated and susceptible strains; the second group, including three cytosolic Slgsts （SlGSTe13, siGSTtl and siGSTzl） and one microsomal Slgst （siMGSTI-1）, was induced only in the 12-generation-treated strain; the third group, including eight cytosolic Slgsts （two epsilon, three delta, one omega, one zeta and one un- classified Slgst）, was expressed 1.52-5.15-fold higher in the 12-generation-treated strain than in the susceptible strain.

关键词： chlorpyrifos expression glutathione S-transferase Spodoptera litura su-perfamily tolerance

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Identification and functional characterization of doublesex gene in the testis of Spodoptera litura

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Insect Science 2019年第6期26卷 1000-1010页

作者： Qian Du Liang Wen si-chun zheng Hong-Lun Bi Yong-Ping Huang Qi-Li Feng Lin Liu Guangzhou Key Laboratory of Insect Development Regulation and Applied Research Institute of Insect Science and Technology School of Life Sciences South China Normal University GuangzhouChina Key Laboratory of Insect Developmental and Evolutionary Biology Institute of Plant Physiology and Ecology Shanghai Institutes for Biological Science Chinese Academy of Sciences ShanghaiChina

Fusion of the testis occurs in most Lepidoptera insects, including Spodoptera litura, an important polyphagous pest. Testicular fusion in S. litura is advantageous for male reproduction, and the molecular mechanism of fusion remains unknown. Doublesex influences the formation of genitalia, the behavior of courtship, and sexually dimorphic traits in fruit-fly and silkworm, and is essential for sexual differentiation. However, its purpose in the testis of S. litura remains unknown. The doublesex gene of S. litura (Sldsx) has male-specific SldsxM and female-specific SldsF isoforms, and exhibits a higher expression level in the male testis. At the testicular fusion stage (L6D6), Sldsx attained the highest expression compared to the pre-fusion and post-fusion periods. Moreover, Sldsx had a higher expression in the peritoneal sheaths of testis than that of germ cells in the follicle. CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/Cas9) was applied to S. litura to determine the role of Sldsx. A mixture of single guide RNA messenger RNA and Cas9 protein (300 ng/μL each) was injected into eggs within 2 h following oviposition. CRISPR/Cas9 successfully induced genomic mutagenesis of Sldsx at Go generation. The mutant males had smaller testis surrounded by less tracheae. Moreover, the mutant males had abnormal external genitalia and could not finish mating with wild-type females. Additionally, testes were fused for almost all mutant males. The results showed that Sldsx was not related to testicular fusion, and is required for both testis development and the formation and function of external genitalia in S. litura. The main roles of doublesex on the male are similar to other insects.

关键词： CRISPR/Cas9 system doublesex external genitalia Spodoptera litura testis

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Homeobox protein A1-like and DNA methylation regulate embryo-specific Zinc finger protein 615 gene expression and embryonic development in the silkworm Bombyx mori

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Insect Science 2023年第4期30卷 1063-1080页

作者： Yu-Lin Tian Tong-Yu Fu Qi-En Zhong Yi-Guang Lin si-chun zheng Guan-Feng Xu Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology Guangzhou Key Laboratory of Insect Development Regulation and Applied ResearchInstitute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina

DNA methylation and transcription factors play roles in gene expression and animal *** insects,DNA methylation modifies gene bodies,but how DNA methylation and transcription factors regulate gene expression is *** this study,we investigated the mechanism that regulates the expression of Bombyx mori Zinc finger pro-tein 615(ZnF 615),which is a downstream gene of DNA methyltransferase 1(Dnmt1),and its effects on the regulation of embryonic *** progressively truncating the ZnF 615 promoter,it was found that the-223 and-190 nt region,which contains homeobox(Hox)protein cis-regulatory elements(CREs),had the greatest impact on the transcription of ZnF *** interference(RNAi)-mediated knockdown and overexpres-sion of Hox family genes showed that Hox A1-like can enhance the messenger RNA level of ZnF *** studies showed that Hox A1-like regulates ZnF 615 expression by directly binding to the-223 and-190 nt region of its *** RNAi-mediated knockdown or overexpression of Hox A1-like and Dnmtl significantly inhibited or enhanced the regulatory effect of either gene alone on ZnF 615 expression,suggesting that both DNA methylation of gene bodies and binding of transcription factors to pro-moters are essential for gene ***-mediated knockdown of Hox A1-like and Dnmtl showed that the embryonic development was retarded and the hatching rate was *** together,these data suggest that Hox A1-like and DNA methylation en-hance the expression of ZnF 615,thereby affecting the development of *** embryos.

关键词： Bombyx mori DNA methylation embryo gene expression Hox Al-like ovary

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Methyl-CpG binding domain(MBD)2/3 specifically recognizes and binds to the genomic mCpG site with a β-sheet in the MBD to affect embryonic development in Bombyx mori

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Insect Science 2023年第6期30卷 1607-1621页

作者： Tong-Yu Fu Shuang-Shun Ji Yu-Lin Tian Yi-Guang Lin Yu-Mei Chen Qi-En Zhong si-chun zheng Guan-Feng Xu Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology Guangzhou Key Laboratory of Insect Development Regulation and Application ResearchInstitute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina

Methyl-CpG(mCpG)binding domain(MBD)proteins especially bind with methylated DNA,and are involved in many important biological processes;however,the binding mechanism between insect MBD2/3 and mCpG remains *** this study,we identified 2 isoforms of the MBD2/3 gene in Bombyx mori,MBD2/3-S and MBD2/*** analysis of MBD2/3-L,MBD2/3-S,and 7 mutant MBD2/3-L proteins deficient inβ1−β6 orα1 in the MBD showed thatβ2−β3-turns in theβ-sheet of the MBD are necessary for the formation of the MBD2/3–mCpG complex;furthermore,other secondary structures,namely,β4−β6 and anα-helix,play a role in stabilizing theβ-sheet structure to ensure that the MBD is able to bind *** addition,sequence alignment and binding analyses of different insect MBD2/3s indicated that insect MBD2/3s have an intact and conserved MBD that binds to the mCpG of target ***,MBD2/3 RNA interference results showed that MBD2/3-L plays a role in regulating *** embryonic development,similar to that of DNA methylation;however,MBD2/3-S withoutβ4−β6 andα-helix does not alter embryonic *** results suggest that MBD2/3-L recognizes and binds to mCpG through the intactβ-sheet structure in its MBD,thus ensuring silkworm embryonic development.

关键词： embryonic development insects MBD2/3 methyl-CpG binding domain(MBD) β-sheet

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