麦红吸浆虫滞育进程中细胞色素c氧化酶活力及SmCOX1和SmCOX2的表达模式分析  

Analysis of cytochrome c oxidase activity and expression patterns of SmCOX1 and SmCOX2 during diapause of Sitodiplosis mosellana(Diptera:Cecidomyidae)

作　　者：古丽旦·白山别克 黄启通 李方向 成卫宁[1] Gulidan BAISHANBIEKE;HUANG Qi-Tong;LI Fang-Xiang;CHENG Wei-Ning(Key Laboratory of Plant Protection Resources and Pest Management of Ministry of Education,College of Plant Protection,Northwest A&F University,Yangling 712100,China;Shandong Institute of Sericulture,Yantai 264002,China;Xi′an Agricultural Technology Extension Centre,Xi′an 710061,China)

机构地区：[1]西北农林科技大学植物保护学院,植保资源与病虫害治理教育部重点实验室,杨凌712100 [2]山东省蚕业研究所,烟台264002 [3]西安市农业技术推广中心,西安710061

出　　处：《昆虫学报》2025年第1期14-22,共9页Acta Entomologica Sinica

基　　金：国家自然科学基金项目(31371933);陕西省重点研发计划项目(2020 NY-059);山东省自然科学基金项目(ZR2023QC051)。

摘　　要：【目的】本研究旨在探讨麦红吸浆虫Sitodiplosis mosellana细胞色素c氧化酶(cytochrome c oxidase,COX)酶活力及其亚基SmCOX1和SmCOX2的基因表达变化与其滞育发育的关系,为阐明麦红吸浆虫滞育的能量代谢机制提供理论依据。【方法】采用COX酶活力测试盒测定麦红吸浆虫自然滞育进程(滞育前、滞育、滞育后静息和滞育后发育)不同阶段幼虫中SmCOX活性;采用RT-PCR和RACE技术克隆SmCOX1和SmCOX2全长cDNA序列并进行生物信息分析;采用qPCR技术检测SmCOX1和SmCOX2在麦红吸浆虫滞育进程中的表达模式。【结果】麦红吸浆虫幼虫中SmCOX活性在滞育阶段较滞育前显著降低,并在整个滞阶段维持较低水平,但在滞育后静息阶段显著升高,到滞育后发育阶段再次升高,显著高于滞育和滞育后静息阶段。克隆获得了开放阅读框长度分别为1536和660 bp的SmCOX1和SmCOX2(GenBank登录号分别为PP466915和PP466916),两个基因碱基A+T含量超过75%,分别编码511和220个氨基酸,预测的蛋白分子量分别为57.70和25.76 kD。氨基酸序列分析表明,SmCOX1和SmCOX2具有线粒体COX酶催化核心典型的氧化还原铜(CuA和CuB)辅基和含血红素的金属中心heme a和heme a3,与同为瘿蚊科(Cecidomyidae)的稻瘿蚊Orseolia oryzae同源蛋白氨基酸序列一致性最高、亲缘关系最近。qPCR结果表明,SmCOX1和SmCOX2在麦红吸浆虫幼虫滞育进程中的表达量存在显著差异,在滞育前和滞育后发育阶段的表达量最高,其次为在滞育后静息期阶段的表达量,在滞育阶段的表达量最低,即变化趋势与SmCOX活性变化基本一致。【结论】麦红吸浆虫滞育进程中SmCOX活性和SmCOX1/2表达的降低与滞育阶段低水平的氧气消耗紧密相关,而其升高与滞育解除和滞育后发育阶段需要较多的能量供给紧密相关。【Aim】This study aims to explore the relationship between the changes in cytochrome c oxidase(COX)activity and gene expression of COX subunits SmCOX1 and SmCOX2 in Sitodiplosis mosellana and diapause development,so as to provide a theoretical basis for elucidating the energy metabolism mechanism of diapause of S.mosellana.【Methods】The SmCOX activities in S.mosellana larvae at different stages in the natural diapause process including pre-diapause,diapause,post-diapause quiescence and post-diapause development were measured using a COX assay kit.The full-length cDNA sequences of SmCOX1 and SmCOX2 were cloned using RT-PCR and RACE techniques,and analyzed by bioinformatics.qPCR technique was used to detect the expression pattern of SmCOX1 and SmCOX2 in the diapause process of S.mosellana.【Results】The SmCOX activity in larvae of S.mosellana in the diapause stage significantly decreased as compared to that in larvae in the pre-diapause stage,remained low level in larvae throughout the diapause stage,significantly increased in larvae in the post-diapause quiescent stage,and increased again in larvae in the post-diapause developmental stage which was significantly higher than those in larvae in the diapause and post-diapause quiescent stages.The open reading frames of SmCOX1 and SmCOX2(GenBank accession number:PP466915 and PP466916,respectively)obtained were 1536 and 660 bp in length,respectively,and contained more than 75%A+T base.SmCOX1 and SmCOX2 encoded the proteins of 511 and 220 amino acids with the predicted molecular weight of 57.70 and 25.76 kD,respectively.The amino acid sequence analysis indicated that SmCOX1 and SmCOX2 contained the classical REDOX copper coenzyme factors(CuA and CuB)and heme-containing metal centers heme a and heme a3 of mitochondria COX catalytic core.SmCOX1 and SmCOX2 displayed the highest amino acid sequence identity and the closest relationship to corresponding homologues from Orseolia oryzae(Cecidomyiidae).qPCR result showed that the expression levels of SmCOX1 and SmCOX2 ex

关 键 词：麦红吸浆虫 滞育 细胞色素C氧化酶 酶活力 基因表达 

分 类 号：Q965[生物学—昆虫学]