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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

Multiple microRNAs control ecdysone signaling in the midgut of Spodoptera litura

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Insect Science 2020年第6期27卷 1208-1223页

作者： Wei Luo li-Xia huang Shuang-Kang Qin Xian Zhang Qi-li Feng Jun Gu li-hua huang 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 Application Research Institute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina Hubei Key Laboratory of Application of Plant-Cancer Active Ingredients School of Chemistry and Life SciencesHubei University of EducationWuhanChina

Metamorphosis is one of the most important physiological processes in *** is regulated by a serial of ecdysone cascade ***,lots o f microRNAs(miRNAs)were investigated in insects;however,their function in metamorphosis is largely *** the present study,the dynamics of a small RNA population was investigated by RNA sequencing from the midgut of a lepidopteran pest Spodoptera litura during larval-pupal metamorphosis.A total of 101 miRNAs were identified,and 75 miRNAs were differentially expressed during the metamorphic *** relationship between these differentially expressed miRNAs and 12 ecdysone cascade genes was analyzed by four classical software programs,and a multiple-to-multiple regulatory network was found to exist between these miRNAs and their *** them,miR-14-3p and its two targets(EcR and E75)were chosen for further ***-14-3p had higher expression level in the 6 th instar larvae as compared with either the prepupae or pupae,which was opposite to that of both EcR and E75,two ecdysone cascade *** reporter assay confirmed that both EcR and E75 were regulated by ***,the 3'untranslated regions are nearly identical to each other among different transcript variants of the ecdysone cascade genes,including EcR,USP,E75,E74,E78,E93,Hr3,Hr4,Hr39,K rhl and F ***,different transcript variants of one ecdysone cascade gene could be regulated by the same mi *** above data suggest that the ecdysone signaling pathway is under the tight control of *** findings expand our understanding of the mechanism of insect metamorphosis and may also provide a novel possibility for the control of pest insects in the future.

关键词： ecdysone EcR E75 metamorphosis microRNA midgut

Cloning and structural characterization of juvenile hormone diol kinase in Spodoptera litura

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Insect Science 2016年第6期23卷 819-828页

作者： Bao-Juan Zeng Yuan Lv li-li Zhang li-hua huang Qi-li Feng Guangzhou Key Laboratory of Insect Development Regulation and Application Research School of Life Sciences South China Normal University Guangzhou 510631 China

Juvenile hormone （JH） is one of the key insect hormones that regulate metamorphosis. Juvenile hormone diol kinase （JHDK） is an enzyme involved in JH metabolism and catalyzes JH diol to form a polar end product, JH diol phosphate that has no JH activity. In this study, a JHDK complementary DNA （cDNA） was cloned from Spodoptera litura and the structure and expression of the gene was characterized. The cDNA was 714 base pairs in length and encoded a protein of 183 amino acids with a molecular mass of 21 kDa and an isoelectric point of 4.55. Based on the structure, three putative calcium binding motifs and guanosine triphosphate-binding motifs were predicted in the protein. Modeling of the 3-D structure showed that the protein consisted of eight α-helixes linked with loops, with no β-sheets. The gene was expressed in the epidermis, fat body and midgut of fifth and sixth instar larvae. The expression level in the epidermis was lower than in the fat body and midgut. The gene was expressed at higher levels at the early stages than in the later stages of fifth and sixth instar midgut and fat body. The results suggest that this gene may be involved in the regulation of the JH titer in larvae ofS. litura.

关键词： juvenile hormone juvenile hormone diol kinase metabolism Spodoptera litura

Cloning, expression and chitin-binding activity of two peritrophin-like protein genes in the common cutworm, Spodoptera fitura

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Insect Science 2014年第4期21卷 449-458页

作者： Wei-Jun Chen li-Xia huang Dan Hu li-Yan liu Jun Gu li-hua huang Qi-li Feng Guangdong Provincial Key Laboratory of Biotechnology for Plant Development School of Life Sciences South China Normal UniversityGuangzhou China

Insect midgut secretes a semi-permeable peritrophic membrane （PM）, which plays important roles in protecting the midgut and helping with food digestion. The lep- idopteran larvae produce type 1 PM, which is degraded when insects develop into the metamorphic stages. However, the underlying mechanism is unclear. In the present study, two peritrophin-like proteins （peritrophin-57 and 37） were identified from the midgut expression sequence tag library and transcriptome of the common cutworm, Spodoptera litura. The temporal and spatial expression patterns and responses to the induction of 20- hydroxyecdysone （20E） and starvation were examined by real-time quantitative polymerase chain reaction according to their common sequence region. The chitin-binding activity was also studied using a competitor, calcofluor. The open reading frames are 1 554 and 1 020 bp, respectively. They shared four highly conserved peritrophin-A domains and were ex- pressed only in the midgut rather than in the other tissues, including fat body, epidermis, Malpighian tube and hemolymph. Their transcriptional expression could only be detected at the larval stages rather than in eggs, prepupae, pupae and adults. The purified protein of peritrophin-37 bound to chitin in a dose-dependent manner. These results indicate that the two proteins are peritrophins, the structural components of PM. In addition, the messenger RNA levels of the two peritrophins were significantly down-regulated by 20E injection, whereas feeding/starvation had no effect on the expression. These findings suggest that the increase of20E titer may be an important factor which controls the degradation of PM during metamorphosis.

关键词： metamorphosis peritrophic membrane peritrophin starvation 20-hydroxyecdysone

Cytoplasmic FKBPs are involved in molting and metamorphosis through regulating the nuclear localization of EcR

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Insect Science 2024年第3期31卷 759-772页

作者： Xian Zhang Qiao-Ran Wang Qian Wu Jun Gu li-hua huang Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied Technology School of Life SciencesSouth China Normal UniversityGuangzhouChina Guangmeiyuan R&D Center Guangdong Provincial Key Laboratory of Insect Developmental Biology and Applied TechnologySouth China Normal UniversityMeizhouChina

Molting and metamorphosis are important physiological processes in insects that are tightly controlled by ecdysone receptor(EcR)through the 20-hydroxyecdysone(20E)signaling *** is a steroid nuclear receptor(SR).Several FK506-binding proteins(FKBPs)have been identified from the mammal SR complex,and are thought to be involved in the subcellular trafficking of ***,their roles in insects are poorly *** explore whether FKBPs are involved in insect molting or metamorphosis,we injected an FKBP inhibitor(FK506)into a lepidopteran insect,Spodoptera litura,and found that molting was inhibited in 61.11%of the larvae,and that the time for larvae to pupate was significantly extended.A total of 10 FKBP genes were identified from the genome of *** and were clustered into 2 distinct groups,according to their subcellular localization,with FKBP13 and FKBP14 belonging to the endoplasmic reticulum(ER)group and with the other members belonging to the cytoplasmic(Cy)*** the CyFKBPs were significantly upregulated in the prepupal or pupal stages,with the opposite being observed for the ER group ***506 completely blocked the transfer of EcR to the nucleus under 20E induction,and significantly downregulated the transcriptional expression of many 20E signaling genes.A similar phenomenon was observed after RNA interference of2 CyFKBPs(FKBP45 and FKBP12b),but not for *** together,our data indicate that the cytoplasmic FKBPs,especially FKBP45 and FKBP12b,mediate the nuclear localization of EcR,thereby regulating the 20E signaling and ultimately affecting molting and metamorphosis in insects.

关键词： EcR FKBP metamorphosis molting RNAi subcellular localization

Discovering genes responsible for silk synthesis in Bombyx moriby piggyBac-based random insertional mutagenesis

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Insect Science 2019年第5期26卷 821-830页

作者： Xing-Bao Feng Zi-Wen Zheng Xian Zhang Jun Gu Qi-li Feng li-hua huang Guangzhou Key Laboratory of Insect Development Regulation and Applied Research Institute of Insect Science and TechnologySchool of Life SciencesSouth China Normal UniversityGuangzhouChina

Silkworm mutants are valuable resources for both transgenic breeding and gene discovery. PiggyBac-based random insertional mutagenesis has been widely used in gene functional studies. In order to discover genes involved in silk synthesis, a piggyBac-based random insertional library was constructed using Bombyx mori, and the mutants with abnormal cocoon were particularly screened. By this means, a “thin cocoon” mutant was identified. This mutant revealed thinner cocoon shell and shorter posterior silk gland (PSG) compared with the wild type. The messenger RNA (mRNA) levels of all the three fibroin genes, including Fib-H, Fib-L and P25, were significantly down-regulated in the PSG of mutants. Four piggyBac insertion sites were identified in Aquaporin (AQP), Longitudinals lacking protein-like {Lola), Glutamyl aminopeptidase-like (GluAP) and Loc101744460. The mRNA levels of all the four genes were significantly altered in the silk gland of mutants. In particular, the mRNA amount of AQP, a gene responsible for the regulation of osmotic pressure, decreased dramatically immediately prior to the spinning stage in the anterior silk gland of mutants. The identification of the genes disrupted in the “thin cocoon” mutant in this study provided useful information for understanding silk production and transgenic breeding of silkworms in the future.

关键词： aquaporin Bombyx mori development mutant piggyBac silk gland

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

Postoperative body weight change and its influencing factors in patients with gastric cancer

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World Journal of Gastrointestinal Surgery 2024年第7期16卷 2242-2254页

作者： Yan li li-hua huang Hui-Di Zhu Ping He Bei-Bei li li-Jing Wen Department of Gastrointestinal Surgery The First Affiliated HospitalZhejiang University School of MedicineHangzhou 310003Zhejiang ProvinceChina Department of Nursing The First Affiliated HospitalZhejiang University School of MedicineHangzhou 310003Zhejiang ProvinceChina

BACKGROUND The high incidence and mortality of gastric cancer(GC)pose a significant threat to human life and health,and it has become an important public health challenge in *** weight loss is a common complication after surgical treatment in patients with GC and is associated with poor prognosis and GC ***,current attention to postoperative weight change in GC patients remains insufficient,and the descriptions of postoperative weight change and its influencing factors are also *** To investigate body weight changes in patients with GC within 6 mo after gastrectomy and identify factors that influence dynamic body weight *** We conducted a prospective longitudinal study of 121 patients with GC and collected data before(T0)and 1(T1),3(T2),and 6(T3)mo after gastrectomy using a general data questionnaire,psychological distress thermometer,and body weight *** general estimation equation(GEE)was used to analyze the dynamic trends of body weight changes and factors that influence body weight changes in patients with GC within 6 mo of *** The median weight loss at T1,T2,and T3 was 7.29%(2.84%,9.40%),11.11%(7.64%,14.91%),and 14.75%(8.80%,19.84%),*** GEE results showed that preoperative body mass index(BMI),significant psychological distress,religious beliefs,and sex were risk factors for weight loss in patients with GC within 6 mo after gastrectomy(Plikely to have weight loss(β=14.685,Plikely to lose weight than those without(β=2.490,Pligious patients were less likely to lose weight 6 mo after gastrectomy than those without religious beliefs(β=-6.844,P=0.001).Compared to female patients,male patients were more likely to experience weight loss 6 mo after gastrectomy(β=4.262,P=0.038).CONCLUSION Male patients wi

关键词： Gastric cancer Gastrectomy Weight loss Influencing factors Body mass index

Simultaneous Determination of Seven Alkaloids in Fufang Zhenzhu Tiaozhi Capsule by HPLC Coupled with DAD

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Chinese Herbal Medicines 2012年第3期4卷 230-236页

作者： Yuan-yuan CHEN Jiao GUO Hui FAN li-hua huang Key Unit of Modulating liver to Treat Hyperlipemia SATCM Level 3 Laboratory of Lipid Metabolism SATCMGuangdong TCM Key Laboratory for Metabolic DiseasesInstitute of Chinese Medicinal SciencesGuangdong Pharmaceutical UniversityGuangzhou 510006China Guangzhou University of Chinese Medicine Guangzhou 510006China

Objective To establish a reverse-phase liquid chromatography method for the determination of seven alkaloids(magnoflorine,columbamine,jatrorrhizine,epiberberine,coptisine,palmatine,and berberine)in Fufang Zhenzhu Tiaozhi *** Chromatography was performed on a Dionex Acclaim C_(18)column(250 mm×4.6 mm,5.0μm)at 30℃.The mobile phase was composed of acetonitrile-potassium dihydrogen phosphate solution(0.015 mol/L,40:60,including 1.7 g/L sodium dodecyl sulfate and phosphoric acid used to regulate pH value to 3.0),with a flow rate of 1.0 mL/*** detection wavelength was 270 *** The calibration curves of magnoflorine,columbamine,jatrorrhizine,epiberberine,coptisine,palmatine,and berberine were linear in the range of 1.07-10.65,0.78-7.55,0.75-7.50,1.60-15.95,2.69-26.85,2.31-23.10,and 6.04-60.40 mg/*** average recoveries of magnoflorine,columbamine,jatrorrhizine,epiberberine,coptisine,palmatine,and berberine were 101.0%,101.2%,100.1%,100.0%,100.1%,101.1%,and 99.7%,*** The method could be used for the quantitative determination of the preparation.

关键词： alkaloids diode array detection Fufang Zhenzhu Tiaozhi Capsule HPLC traditional Chinese medicines