高级检索

留言板

尊敬的读者、作者、审稿人, 关于本刊的投稿、审稿、编辑和出版的任何问题, 您可以本页添加留言。我们将尽快给您答复。谢谢您的支持!

姓名
邮箱
手机号码
标题
留言内容
验证码

舞毒蛾小分子热激蛋白基因分析及对甲萘威胁迫的响应

刘鹏 孙丽丽 张琪慧 曹传旺

刘鹏, 孙丽丽, 张琪慧, 曹传旺. 舞毒蛾小分子热激蛋白基因分析及对甲萘威胁迫的响应[J]. 北京林业大学学报, 2017, 39(1): 78-84. doi: 10.13332/j.1000-1522.20160179
引用本文: 刘鹏, 孙丽丽, 张琪慧, 曹传旺. 舞毒蛾小分子热激蛋白基因分析及对甲萘威胁迫的响应[J]. 北京林业大学学报, 2017, 39(1): 78-84. doi: 10.13332/j.1000-1522.20160179
LIU Peng, SUN Li-li, ZHANG Qi-hui, CAO Chuan-wang. Analysis of smHsp genes in Lymantria dispar and its response to carbaryl stress[J]. Journal of Beijing Forestry University, 2017, 39(1): 78-84. doi: 10.13332/j.1000-1522.20160179
Citation: LIU Peng, SUN Li-li, ZHANG Qi-hui, CAO Chuan-wang. Analysis of smHsp genes in Lymantria dispar and its response to carbaryl stress[J]. Journal of Beijing Forestry University, 2017, 39(1): 78-84. doi: 10.13332/j.1000-1522.20160179

舞毒蛾小分子热激蛋白基因分析及对甲萘威胁迫的响应

doi: 10.13332/j.1000-1522.20160179
基金项目: 

黑龙江省自然科学基金面上项目 C201409

国家自然科学基金项目 31101676

国家自然科学基金项目 31570642

详细信息
    作者简介:

    刘鹏。主要研究方向:昆虫毒理学。Email: 775627232@qq.com  地址:150040  黑龙江省哈尔滨市和兴路26号东北林业大学林学院

    责任作者:

    曹传旺,教授,博士生导师。主要研究方向:昆虫毒理学。Email: chuanwangcao@nefu.edu.cn  地址:同上

  • 中图分类号: S763.42

Analysis of smHsp genes in Lymantria dispar and its response to carbaryl stress

  • 摘要: 昆虫小分子热激蛋白(smHsp)是分子质量在12~43 kDa、具有分子伴侣活性的应激蛋白,可以在环境胁迫下帮助蛋白质折叠和转运。本文为明确舞毒蛾的smHsp基因特性及对杀虫剂甲萘威胁迫的响应,从舞毒蛾无参照全转录本文库中鉴定获得6个小分子热激蛋白家族基因,全长基因开放阅读框为444~567 bp,编码147~188个氨基酸,理论等电点为5.58~6.17。根据分子质量大小分别命名为LdHsp17.0、LdHsp18.7、LdHsp19.1、LdHsp20.3、LdHsp21.3和LdHsp21.4。亚致死剂量(LC5、LC10和LC30)的甲萘威处理可诱导舞毒蛾smHsp家族中LdHsp20.3、LdHsp19.1和LdHsp17.0上调表达,但抑制了LdHsp21.4、LdHsp21.3和LdHsp18.7的表达,且甲萘威处理对smHsp基因表达量的影响存在质量浓度和剂量效应。以上结果为进一步研究舞毒蛾小分子热激蛋白功能及其参与对杀虫剂抗性的机制提供了基础。

     

  • 图  1  舞毒蛾小分子热激蛋白氨基酸多序列比对

    相同和相似氨基酸残基分别用黑色和灰色框表示。

    Figure  1.  Alignment of the deduced amino acid sequences of six identified smHsp genes

    Identical and similar amino acid residues are indicated by black and gray boxes, respectively.

    图  2  几种昆虫小分子热激蛋白的分子进化树

    Figure  2.  Phylogenetic analysis of small heat shock proteins from some insects

    图  3  甲萘威胁迫对舞毒蛾smHsp基因表达量的影响

    A.亚致死剂量LC5处理组smHsp基因表达量;B.亚致死剂量LC10处理组smHsp基因表达量;C.亚致死剂量LC30处理组smHsp基因表达量。不同小写字母表示同一基因不同处理间的表达量差异显著(P<0.05)。

    Figure  3.  Effects of carbaryl on smHsp gene expressions in L. dispar larvae

    A, smHsp gene expression under LC5 of sublethal dose stress; B, smHsp gene expression under LC10 of sublethal dose stress; C, smHsp gene expression under LC30 of sublethal dose stress. Different lowercase letters show significant differences between treatment groups for the same gene at P < 0.05 level.

    表  1  实时荧光定量RT-PCR引物序列

    Table  1.   Primer sequences of real-time RT-PCR

    基因
    Gene
    正向引物序列(5′-3′)
    Forward primer sequence (5′-3′)
    反向引物序列(5′-3′)
    Reverse primer sequence (5′-3′)
    LdHsp20.3AGACGTCGGCTCTACTATCACATCACGGAATCGGCATCTA
    LdHsp18.7GACTCCACAGCATCAGGATTAGGGATGCCGGATCTTCAATCA
    LdHsp21.4CTCTGCTCTCCGATGACTACTACGAACTGTCGACTGATGTATCC
    LdHsp19.1CTCCGTACTGGATGCGTTATCCACCTTCTTCCCGTCGATTT
    LdHsp17.0GGAGCGTGACAAGTACGAAACCGAGGTACGGTGATAACTAGA
    LdHsp21.3GGGTGTACTGGCTAACATCAACCGAAATGAGGAAGATGGAAGA
    ActinAGAAGCACTTGCGGTGGACAATACCTGTACGCCAACACTGTCAT
    EF1αTTTGCCTTCCTTGCGCTCAACATGTAAAGCAGCTGATCGTGGGT
    TUBAATGCAAGAAAGCCTTGCGCCTATGAAGGAGGTCGACGAGCAAA
    下载: 导出CSV

    表  2  舞毒蛾smHsp基因特性

    Table  2.   Characteristics of smHsp genes in L. dispar

    基因
    Gene
    开放阅读框
    Open reading frame/bp
    编码氨基酸
    Encoding amino acids
    分子质量
    Molecular weight/kDa
    理论等电点
    Theoretical pI
    含量最多的氨基酸
    The most abundant amino acids/%
    LdHsp17.044414717.006.15Lys(9.5%)
    LdHsp18.750116618.795.58Val(9.6%)
    LdHsp19.149516419.106.13Glu(11.0%)
    LdHsp20.353417720.305.76Glu(8.5%)
    LdHsp21.456718821.426.17Leu(9.0%)
    LdHsp21.356418721.375.79Ser(11.8%)
    下载: 导出CSV
  • [1] SZABO A, KORSZUN R, HARTL F U, et al. A zinc finger-like domain of the molecular chaperone DnaJ is involved in binding to denatured protein substrates[J]. Embo Journal, 1996, 15(2): 408-417. doi: 10.1002/j.1460-2075.1996.tb00371.x
    [2] 陈华友, 张春霞, 马晓珂, 等.极端嗜热古菌的热休克蛋白[J].生物工程学报, 2008, 24(12): 2011-2021. doi: 10.3321/j.issn:1000-3061.2008.12.001

    CHEN H Y, ZHANG C X, MA X K, et al. Heat shock proteins of the hyperthermophilic archaea[J]. Chinese Journal of Biotechnology, 2008, 24(12): 2011-2021. doi: 10.3321/j.issn:1000-3061.2008.12.001
    [3] SǾRENSEN J G, KRISTENSEN T N, LOESCHCKE V. The evolutionary and ecological role of heat shock proteins[J]. Ecology Letters, 2003, 6(11): 1025-1037. doi: 10.1046/j.1461-0248.2003.00528.x
    [4] FRANK E, MADSEN O, VAN RHEELE T, et al. Evolutionary diversity of vertebrate small heat shock proteins[J]. Journal of Molecular Evolution, 2004, 59(6): 792-805. doi: 10.1007/s00239-004-0013-z
    [5] HASLBECK M, FRANZMANN T, WEINFURTNER D, et al. Some like it hot: the structure and function of small heat-shock proteins[J]. Nature Structural & Molecular Biology, 2005, 12(10): 842-846. doi: 10.1038-nsmb993/
    [6] EYLES S J, GIERASCH L M. Nature's molecular sponges: small heat shock proteins grow into their chaperone roles[J]. Proceedings of the National Academy of Sciences of the United States of America, 2010, 107(7): 2727-2728. doi: 10.1073/pnas.0915160107
    [7] NARBERHAUS F. Alpha-crystallin-type heat shock proteins: socializing mini chaperones in the context of a multi chaperone network[J]. Microbiology and Molecular Biology Reviews, 2002, 66(1): 64-93. https://www.ncbi.nlm.nih.gov/pubmed/11875128
    [8] KIM K K, KIM R, KIM S H. Crystal structure of a small heat-shock protein[J]. Nature, 1998, 394: 595-599. doi: 10.1038/29106
    [9] GARRIDO C, PAUL C, SEIGNEURIC R, et al. The small heat shock proteins family: the long forgotten chaperones[J]. International Journal of Biochemistry & Cell Biology, 2012, 44(10): 1588-1592.
    [10] WETTSTEIN G, BELLAYE P S, MICHEAU O, et al. Small heat shock proteins and the cytoskeleton: an essential interplay for cell integrity?[J]. International Journal of Biochemistry & Cell Biology, 2012, 44(10): 1680-1686. https://www.sciencedirect.com/science/article/pii/S1357272512001902
    [11] 陆明星, 徐静, 杜予州.昆虫小分子量热激蛋白的研究进展[J].应用昆虫学报, 2015, 52(6): 1326-1332. http://d.old.wanfangdata.com.cn/Periodical/kczs201506002

    LU M X, XU J, DU Y Z. Progress in research on insect small heat shock proteins[J]. Chinese Journal of Applied Entomology, 2015, 52(6): 1326-1332. http://d.old.wanfangdata.com.cn/Periodical/kczs201506002
    [12] LAZAREVI C J, ANDJELKOVIC M. Host plant effects on the genetic variation and correlations in the individual performance of the gypsy moth[J]. Functional Ecology, 1998, 12(1): 141-148. http://www.wanfangdata.com.cn/details/detail.do?_type=perio&id=10.1046/j.1365-2435.1998.00166.x
    [13] 曹传旺, 孙丽丽, 问荣荣, 等.舞毒蛾LdOA1基因克隆分析及对3种杀虫剂胁迫的响应[J].林业科学, 2014, 50(8): 102-107. http://d.old.wanfangdata.com.cn/Periodical/lykx201408015

    CAO C W, SUN L L, WEN R R, et al. Cloning and analysis of LdOA1 in Lymantria dispar and its response to the stress of three kinds of insecticides[J]. Scientia Silvae Sinicae, 2014, 50(8): 102-107. http://d.old.wanfangdata.com.cn/Periodical/lykx201408015
    [14] CAO C W, SUN L L, WEN R R, et al. Characterization of the transcriptome of the Asian gypsy moth Lymantria dispar identifies numerous transcripts associated with insecticide resistance[J]. Pesticide Biochemistry and Physiology, 2015, 119: 54-61. doi: 10.1016/j.pestbp.2015.02.005
    [15] BRODSKY L I, DRACHEV A L, LEONTOVICH A M, et al. A novel method of alignment of biopolymer sequences[J]. Biosystems, 1993, 30(1-3): 65-79. doi: 10.1016/0303-2647(93)90063-I
    [16] PFAFFL M W, HORGAN G W, DEMPFLE L. Relative expression software tool (REST) for group-wise comparison and statistical analysis of relative expression results in real-time PCR[J]. Nucleic Acids Research, 2002, 30: 36. doi: 10.1093/nar/30.9.e36
    [17] VIERLING E. The role of heat shock proteins in plants[J]. Plant Molecular Biology, 1991, 42: 579-620. http://d.old.wanfangdata.com.cn/OAPaper/oai_pubmedcentral.nih.gov_1206773
    [18] ARTS M J, SCHILL R O, KNIGGE T, et al. Stress proteins (hsp70, hsp60) induced in isopods and nematodes by field exposure to metals in a gradient near Avonmouth, UK[J]. Ecotoxicology, 2004, 13(8): 739-755. doi: 10.1007/s10646-003-4473-5
    [19] PIANO A, VALBONESI P, FABBRI E. Expression of cytoprotective proteins, heat shock protein 70 and metallothioneins, in tissues of Ostrea edulis exposed to heat and heavy metals[J]. Cell Stress & Chaperones, 2004, 9(2): 134-142. http://d.old.wanfangdata.com.cn/OAPaper/oai_pubmedcentral.nih.gov_1065293
    [20] SHARMA A, MISHRAA M, SHUKLAA A K, et al. Organochlorine pesticide, endosulfan induced cellular and organismal response in Drosophila melanogaster [J]. Journal of Hazardous Materials, 2012, 221-222(4): 275-287. http://www.wanfangdata.com.cn/details/detail.do?_type=perio&id=b291a9bde3cd280f96323a2ec52aad52
    [21] 王瑞娴, 高瑞娜, 张婷, 等.外源物质诱导家蚕小分子热激蛋白基因HSP19. 9的组织表达活性分析[J].蚕业科学, 2010, 36(5): 780-784. doi: 10.3969/j.issn.0257-4799.2010.05.008

    WANG R X, GAO R N, ZHANG T, et al. An analysis on tissue expression activities of Bombyx mori small molecule heat shock protein gene HSP19. 9 induced by exogenous substances[J]. Science of Sericulture, 2010, 36(5): 780-784. doi: 10.3969/j.issn.0257-4799.2010.05.008
    [22] 王利华, 单丹, 姚静, 等.温度和毒死蜱对灰飞虱雌成虫热激蛋白70和90基因的诱导表达特性研究[J].应用昆虫学报, 2015, 52(4): 828-836. http://d.old.wanfangdata.com.cn/Periodical/kczs201504005

    WANG L H, SHAN D, YAO J, et al. Induced expression of the heat shock protein 70 and 90 genes by exposure to high temperature and chlorpyrifosin female of Laodelphax striatellus[J]. Chinese Journal of Applied Entomology, 2015, 52(4): 828-836. http://d.old.wanfangdata.com.cn/Periodical/kczs201504005
    [23] 寇利花, 吴海花, 刘耀明, 等.镉急性染毒对中华稻蝗精卵巢小分子热休克蛋白基因表达的影响[J].农业环境科学学报, 2015, 34(1): 7-14. http://d.old.wanfangdata.com.cn/Periodical/nyhjbh201501002

    KOU L H, WU H H, LIU Y M, et al. Expression analysis of OcsHSP genes in testis and ovary of Oxya chinensis exposed to acute cadmium[J]. Journal of Agro-Environment Science, 2015, 34(1): 7-14. http://d.old.wanfangdata.com.cn/Periodical/nyhjbh201501002
  • 加载中
图(3) / 表(2)
计量
  • 文章访问数:  953
  • HTML全文浏览量:  215
  • PDF下载量:  17
  • 被引次数: 0
出版历程
  • 收稿日期:  2016-05-23
  • 修回日期:  2016-08-13
  • 刊出日期:  2017-01-01

目录

    /

    返回文章
    返回