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Expression, subcellular localization and protein-protein interaction of four isoforms of EcR/USP in the common cutworm

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Insect Science 2015年第1期22卷 95-105页

作者： Li-Xia Huang Yan-Jun Gong Jun Gu Bao-Juan Zeng Li-Hua Huang Qi-Li Feng laboratory of molecular and developmental entomology Guangdong Provincial Key Laboratory of Biotechnology for Plant DevelopmentSchool of Life Sciences South China Normal University Guangzhou China

Ecdysone receptor （EcR） and ultraspiracle （USP） form heterodimers to mediate ecdysteroid signaling during molting and metamorphosis. Various EcR/USP heterodimers have been reported. However, it is unclear what kind of EcR/USP combination is adopted by lepidopteran insects during the larval-pupal metamorphosis and whether the EcR/USP heterodimer varies among different tissues. To address these questions, two isoforms of each EcR and USP were cloned from the common cutworm, their messenger RNA expression patterns were examined by real-time quantitative polymerase chain reaction in different tissues during the larval-pupal metamorphosis and in the midgut in response to hormonal induction. Furthermore, their subcellular localization and protein-protein interaction were explored by transient expression and far-western blotting, respectively. All the four genes were significantly up-regulated in prepuae and/or pupae. The expression profiles of EcRB1 and USP1 were nearly identical to each other in the epidermis, fat body and midgut, and a similar situation also applied to EcRA and USP2. The three genes responded to 20-hydroxyecdysone （20E） induction except for USP2, and USP1 could be up-regulated by both 20E and juvenile hormone. The four proteins mainly localized in the nucleus and the nuclear localization was promoted by 20E. The protein-protein interaction between each EcR and USP was found in vitro. These results suggest that two types of EcR/USP heterodimer （EeRA/USP2 and EcRB 1/USP1） may exist simultaneously in the common cutworm, and the latter should play more important roles during the larval-pupal metamorphosis. In addition, the types of EcR/USP heterodimer do not vary in the tissues which undergo histolysis and regeneration during metamorphosis.

关键词： ecdysone receptor protein interaction subcellular localization ultraspiracle

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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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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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molecular Dissection of Bombyx mori Nucleopolyhedrovirus orf8 Gene

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Virologica Sinica 2009年第4期24卷 315-322页

作者： WonKyung Kang molecular entomology laboratory RIKEN Advanced Science Institute

Viruses including baculoviruses are obligatory parasites, as their genomes do not encode all the proteins required for replication. Therefore, viruses have evolved to exploit the behavior and the physiology of their hosts and olden coevolved with their hosts over millions of years. Recent comparative analyses of complete genome sequences of baculoviruses revealed the patterns of gene acquisitions and losses that have occurred during baculovirus evolution. In addition, knowledge of virus genes has also provided understanding of the mechanism of baculovirus infection including replication, species-specific virulence and host range. The Bm8 gene of Bombyx mori nucleopolyhedrovirus （NPV） and its homologues are found only in group I NPV genomes. The Autographa californica NPV Ac 16 gene is a homologue of Bm8 and, encodes a viral structural protein. It has been shown that BmS/Ac 16 interacts with baculoviral and cellular proteins. BmS/Ac 16 interacts with baculoviral IE1 that is facilitated by coiled coil domains, and the interaction with IE1 is important for Bin8 function. Acl6 also forms a complex with viral FP25 and cellular actin and associates with membranes via palmitoylation. These data suggested that this gene family encodes a multifunctional protein that accomplishes specific needs of group I NPVs.

关键词： Group I NPV Bm8 Early gene Structural protein Nuclear localization

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Genome-wide identification, characterization and evolution of cuticular protein genes in the malaria vector Anopheles sinensis （Diptera： Culicidae）

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Insect Science 2018年第5期25卷 739-750页

作者： Bai-Qi Liu Liang Qiao Qi-Yi He Yong Zhou Shuang Ren Bin Chen Chongqing Key laboratory of Vector Insects Institute of Entomology and Molecular Biology Chongqing Normal University Chongqing China

Thirteen cuticular protein （CP） families have been recognized in arthropods. In this study, 250 Anopheles sinensis CP genes were identified and named based on genome and transcriptome sequences. They were classified into 10 families based on mo- tifs and phylogenetic analyses. In 11 other insect species, nine had CP numbers 〉 150 while Apis mellifera and Tribolium castaneum had CP numbers less than 52. The CPs of eight species occupied 〉 1.4% of the total genomic gene number, whereas in three species the CPs occupied 〈 1%. The phylogenies for each CP family in An. sinensis were constructed and discussed. The 250 CPs each had 1-8 exons with 144 CPs （57.6%） having two exons. The intron length ranged from 66--3888 bp with 174 introns （54.0%） being 66--100 bp long. Except for two CPs on two contigs, 248 CPs were mapped onto 28 scaffolds with 136 genes （54.4%） restricted to five scaffolds. A total of 107 CPs were clustered and located at 27 loci. The CPR family had the conserved motif GSYS- LVEPDGTVRTV. The RR- 1 subfamily had an additional 21 amino acid （aa） motifs with the YVADENGF sequence that is common in insects. The RR-2 subfamily had an additional 50 aa motifs with two additional regions RDGDWKG and G-x（3）-VV. A comparison with 115 orthologous counterparts of An. gambiae CPs suggested purifying selection for all of these genes. This study provides basic information useful for further studies on biological functions of An. sinensis CPs as well as for comparative genomics of insect CPs.

关键词： Anopheles sinensis characterization cuticular protein genes genome-wideidentification phylogenetics Ka/Ks

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Mitogenome-based phylogeny of mosquitoes(Diptera:Culicidae)

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Insect Science 2024年第2期31卷 599-612页

作者： De-Hong Chen Shu-Lin He Wen-Bo Fu Zhen-Tian Yan Yun-Jian Hu Huan Yuan Ming-Bin Wang Bin Chen Chongqing Key laboratory of Vector Insects Institute of Entomology and Molecular BiologyChongqing Normal UniversityChongqingChina

Mosquitoes are of great medical significance as vectors of many deadly *** have been widely used in phylogenetic studies,but mitogenome knowledge within the family Culicidae is limited,and Culicidae phylogeny is far from *** this study,we surveyed the mitogenomes of 149 Culicidae species,including 7 newly sequenced *** analysis of 149 mosquito mitogenomes shows gene composition and order to be identical to that of an ancestral insect,and the AT bias,length variation,and codon usage are all consistent with that of other reported Dipteran *** analyses based on the DNA sequences of the 13 protein-coding genes from the 149 species robustly support the monophyly of the subfamily Anophelinae and the tribes Aedini,Culicini,Mansonini,Sabethini,and *** resolve ambiguous relationships between clades within the subfamily Culicinae,we performed topological tests and show that Aedini is a sister to Culicini and that Uranotaenini is a sister to(Mansonini)+(Toxorhynchitini+Sabethini).In addition,we estimated divergence times using a Bayesian relaxation clock based on the sequence data and 3 fossil calibration *** results show mosquitoes diverged during the Early Jurassic with massive Culicinae radiations during the Cretaceous,coincident with the emergence of angiosperms and the burst of mammals and ***,this study,which uses the largest number of Culicidae mitogenomes sequenced to date,comprehensively reveals the mitogenome characteristics and mitogenome-based phylogeny and divergence times of Culicidae,providing information for further studies on the mitogenome,phylogeny,evolution,and taxonomic revision of Culicidae.

关键词： characterization Culicidae evolution mitogenomes mosquitoes phylogeny

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First report of complete mitochondrial genome in the subfamily Alleculinae and mitochondrial genome-based phylogenetics in Tenebrionidae(Coleoptera:Tenebrionoidea)

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Insect Science 2022年第4期29卷 1226-1238页

作者： Chuan Wu Yong Zhou Tian Tian Ting-Jing Li Bin Chen Chongqing Key laboratory of Vector Insects Institute of Entomology and Molecular BiologyChongqing Normal UniversityChongqingChina

Despite worldwide distribution and rich diversity,the knowledge of mitochondrial genome(mtgenome)characteristics within the family Tenebrionidae(Coleoptera)is still very limited,and phylogenetics remain unresolved for the *** this study,the mtgenomes for 12 species are newly sequenced and *** of the species represent the first report of complete mtgenomes from the subfamily *** analysis of 36 tenebrionid mtgenomes finds that gene composition and order are the same as a theoretical ancestral insect mtgenome,and AT bias,length variation,and codon usage are consistent with other reported beetle *** intergenic overlap and gap sequences may contain phylogenetically informative information,whereas neither the conserved nor repeat sequences in the control region *** subfamilies Lagriinae,Pimeliinae,Stenochiinae,and Alleculinae are found to be monophyletic,and the subfamilies Diaperinae and Tenebrioninae polyphyletic in our ***,Lagriinae is sister to the rest of the *** the tribal level,the tribes Lagriini in the subfamily Lagrrinae;Strongyliini in Stenochiinae;Cteniopodini in Alleculinae;and Triboliini,Opatrini,and Amarygmini in Tenebrioninae are monophyletic;while Diaperini in Diaperinae,and Tenebrionini in Tenebrioninae are polyphyletic.

关键词： Alleculinac characteristics Colcopiera mtgenome phylogenetics Tenebrionidac

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Comparative analyses of simple sequence repeats(SSRs)in 23 mosquito species genomes:Identification,characterization and distribution(Diptera:Culicidae)

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Insect Science 2019年第4期26卷 607-619页

作者： Xiao-Ting Wang Yu-Juan Zhang Liang Qiao Bin Chen Chongqing Key laboratory of Vector Insects Chongqing Key Laboratory of Animal BiologyInstitute of Entomology and Molecular BiologyChongqing Normal UniversityChongqingChina

Simple sequence repeats (SSRs) exist in both eukaryotic and prokaryotic genomes and are the most popular genetic markers, but the SSRs of mosquito genomes are still not well understood. In this study, we identified and analyzed the SSRs in 23 mosquito species using Drosophila melanogaster as reference at the whole-genome level. The results show that SSR numbers (33 076-560 175/genome) and genome sizes (574.57-1342.21 Mb) are significantly positively correlated (R~= 0.8992, P < 0.01), but the correlation in individual species varies in these mosquito species. In six types of SSR, mono- to trinucleotide SSRs are dominant with cumulative percentages of 95.14%-99.00% and densities of 195.65/Mb-787.51/Mb, whereas tetra- to hexanucleotide SSRs are rare with 1.12%-4.22% and 3.76/Mb-40.23/Mb. The (A/T)n,(AC/GT)n and (AGC/GCT)n are the most frequent motifs in mononucleotide, dinucleotide and trinucleotide SSRs, respectively, and the motif frequencies of tetra- to hexanucleotide SSRs appear to be species-specific. The 10-20 bp length of SSRs are dominant with the number of 11() 561 ± 93 482 and the frequency of 87.25%± 5.73% on average, and the number and frequency decline with the increase oflength. Most SSRs(83.34%± 7.72%) are located in intergenic regions, followed by intron regions (11.59%± 5.59%), exon regions (3.74%± 1.95%), and untranslated regions (1.32%± 1.39%). The mono-, di- and trinucleotide SSRs are the main SSRs in both gene regions (98.55%± 0.85%) and exon regions (99.27%± 0.52%). An average of 42.52% of total genes contains SSRs, and the preference for SSR occurrenee in different gene subcategories are species-specific. The study provides useful insights into the SSR diversity, characteristics and distribution in 23 mosquito species of genomes.

关键词： characterization comparative analyses distribution mosquito simple sequence repeats(SSRs) whole-genome identification

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Identification and characterization of a gene encoding a UBX domain-containing protein in the migratory locust, Locusta migratoria manilensis

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Insect Science 2013年第4期20卷 497-504页

作者： Zheng-Bo He Yu Xie Feng-Ling Si Bin Chen Institute of entomology and molecular Biology Chongqing Key Laboratory of Animal Biology College of Life Sciences Chongqing NormalUniversity Chongqing China

Ubiquitin regulatory X （UBX） domain-containing proteins are believed to func- tion as cofactors for p97/CDC48, an adenosine triphosphatase shown to be involved in mul- tiple cellular processes. In the present study, a full-length complementary DNA （cDNA） of UBX domain-containing gene, termed LmUBX1, was cloned from Locusta migratoria manilensis and characterized, using random amplification of cDNA ends polymerase chain reaction （RACE PCR）, sequence analysis and quantitative real-time PCR. LmUBX1, 1 600 bp in length, is predicted to encode a 446-amino acid protein with a predicted molec- ular weight of 51.18 kDa that contains a central PUB domain and a carboxy-terminal UBX domain. Homology analysis revealed that LmUBX1 has higher similarity to the known UBX domain-containing proteins from insects than from other species. Moreover, based on sequence characteristics and phylogenetic relationships, it is suggested that LmUBX1 can be classified into the UBXD1 subfamily. Expression analysis founded that LmUBX1 exhibited significant expression variations at different developmental stages and in differ- ent tissues, suggesting that the expression of LmUBX1 was highly regulated. Interestingly, its messenger RNA transcript was more abundant in ovary and testis than in other tissues examined, suggesting that it may have more important roles in the reproductive system. In addition, LmUBX1 was differentially expressed in gregarious and solitary locusts and was significantly up-regulated in third and fifth instars of gregarious locusts, implying that LmUBX1 was also likely involved in the phase polyphenisms in L. migratoria manilen- sis. To our knowledge, this is the first report of cloning of a full-length cDNA of UBX domain-containing gene from L. migratoria manilensis.

关键词： cloning expression Locusta migratoria manilensis UBX domain-containing protein

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Sialotranscriptome sequencing and analysis of Anopheles sinensis and comparison with Psorophora albipes sialotranscriptome （Diptera： Culicidae）

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Insect Science 2018年第3期25卷 368-378页

作者： Ya-Jie Fang Zhen-Tian Yan Bin Chen Chongqing Key laboratory of Vector Insects Institute of Entomology and Molecular BiologyCollege of Life SciencesChongqing Normal UniversityChongqing 401331China

Most of adult female mosquitoes secrete saliva to facilitate blood sucking, digestion and nutrition, and mosquito-borne disease prevention. The knowledge of classification and characteristics of sialotranscriptome genes are still quite limited, Anopheles sinensis is a major malaria vector in China and southeast Asian countries. In this study, the An. sinensis sialotranscriptome was sequenced using Illumina sequencing technique with a total of 10 907 unigenes to be obtained and annotated in biological functions and pathways, and 10 470 tmigenes were mapped to An. sinensis reference genome with 70.46% of genes having 90%- 100% genome mapping through bioinformatics analysis. These mapped genes were classified into four categories： housekeeping （6632 genes）, secreted （1177）, protein-coding genes with function-unknown （2646） and transposable element （15）. The housekeeping genes were divided into 27 classes, and the secreted genes were divided into 11 classes and 96 families. The classification, characteristics and evolution of these classes/families of secreted genes are further described and discussed. The comparison of the 1177 secreted genes in An. sinensis in the Anophelinae subfamily with 811 in Psorophora albipes in the Culicinae subfamily show that six classes/subclasses have the gene number more than twice and two classes （uniquely found in anophelines, and Orphan proteins of unique standing） are unique in the former compared with the latter, whereas four classes/subclasses are much expanded and uniquely found in the Aedes class and is unique in the later. The An. sinensis sialotranscriptome sequence data is the most complete in mosquitoes to date, and the analyses provide a comprehensive information frame for further research of mosquito sialotranscriptome.

关键词： Anopheles sinensis comparative analysis gene classification Psorophora albipes sialotranscriptome

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