褐飞虱中富含丝氨酸/精氨酸的可变剪接因子特性和生物学功能研究  

Molecular Characterization and Biological Function of Serine/Arginine-rich Alternative Splicing Factors in Nilaparvata lugens(Hemiptera:Delphacidae)

作　　者：贾毅帆 王新峰 王雅宣 刘芳[1] 肖晶 魏琪 傅强[1] 万品俊 JIA Yifan;WANG Xinfeng;WANG Yaxuan;LIU Fang;XIAO Jing;WEI Qi;FU Qiang;WAN Pinjun(State Key Laboratory of Rice Biology and Breeding,China National Rice Research Institute,Hangzhou 311401)

机构地区：[1]中国水稻研究所水稻生物育种全国重点实验室,杭州311401

出　　处：《中国水稻科学》2025年第2期277-286,共10页Chinese Journal of Rice Science

基　　金：现代农业产业技术体系建设专项(CARS-01);浙江省自然科学基金资助项目(LY22C140008);中央级公益性科研院所基本科研业务费专项(CPSIBRF-CNRRI-202406)。

摘　　要：【目的】SR蛋白(serine/arginine-rich protein)是一类富含丝氨酸/精氨酸的可变剪接因子,在褐飞虱体内对基因的剪接过程起调控作用。本研究通过克隆褐飞虱(Nilaparvata lugens)的5个NlSR基因并鉴定了其分子特性、表达模式和生物学功能,为褐飞虱的防治提供新的思路。【方法】基于褐飞虱基因组数据,利用PCR技术克隆了5个NlSR基因c DNA序列,并利用生物信息学手段分析其序列特征;利用q RT-PCR技术检测它们在不同发育阶段(卵、1-5龄若虫和雌雄成虫)和组织(唾液腺、体壁、脂肪体、卵巢和中肠)中的表达特征;并测定干扰后的基因表达及褐飞虱的存活率、蜜露量和体质量变化。【结果】成功克隆了5个NlSR基因的c DNA序列,分别命名为NlSRSF1、NlSRSF2.1、NlSRSF2.2、NlSRSF7.1和NlSRSF7.2。根据进化树分类,这些基因被分为3个亚家族:SRSF1、SRSF2和SRSF7。NlSRs基因的开放阅读框长度在495bp到508bp之间,编码的氨基酸数目介于164aa到235aa,预测分子量在19.54kD到26.76kD之间,等电点在9.53到11.83之间,均为亲水性碱性蛋白,不稳定指数在63.55到131.97之间。编码的蛋白含有N端的RRM结构域(RNArecognitionmotif)和C端的RS结构域(Arginine/serine-richdomain),NlSRSF7.1和NlSRSF7.2还含有锌指(Zn F_C2HC)结构域。NlSR基因在褐飞虱的唾液腺、体壁、脂肪体、卵巢和中肠等组织中均有表达,其中NlSRSF1和NlSRSF2.1在唾液腺中的相对表达量较高;NlSRSF1主要在卵和1龄若虫中表达,其他基因主要在成虫中表达。RNAi实验结果表明,与对照组ds GFP相比,干扰NlSRSF1、NlSRSF2.1和NlSRSF7.2显著降低褐飞虱的存活率,干扰NlSRSF2.2和NlSRSF7.1对存活率无显著影响,干扰所有NlSR基因均显著降低了褐飞虱的蜜露量和体质量增加量。【结论】本研究克隆了褐飞虱的5个NlSR基因,分析了其序列和表达特征,并通过干扰确定它们对褐飞虱生命活动的影响,为深入研究褐飞虱NlSRs的【Objective】Serine/arginine-rich(SR)proteins are a class of alternative splicing factors that regulate gene splicing in the brown planthopper,Nilaparvata lugens.This study aimed to clone five NlSR genes from N.lugens and identify their molecular characteristics,expression patterns,and biological functions,providing new insights for controlling brown planthoppers.【Method】Based on the genomic data of N.lugens,the cDNA sequences of five NlSR genes were cloned using RT-PCR.Their sequence characteristics were analyzed.The expression profiles of NlSRs in different developmental stages(egg,1st-5th instar nymphs,and male and female adults)and tissues(salivary glands,integument,fat body,ovaries,and midgut)of N.lugens were detected using qRT-PCR.RNA interference(RNAi)technology was utilized to interfere with the relative expression levels of the NlSR genes.The expression of NlSRs,survival rates,honeydew production,and body weight gains of N.lugens after RNAi were measured.【Results】Five cDNA sequences of NlSR genes were cloned,which were designated as NlSRSF1,NlSRSF2.1,NlSRSF2.2,NlSRSF7.1,and NlSRSF7.2.Based on phylogenetic tree analysis,the deduced proteins were classified into three subfamilies:SRSF1,SRSF2,and SRSF7.The open reading frames of the NlSR genes ranged from 495 to 508 bp,encoding proteins with 164-235 amino acids.The predicted molecular weights ranged from 19.54 to 26.76 kD,with isoelectric points ranging from 9.53 to 11.83.These proteins were hydrophilic and alkaline,with instability indices ranging from 63.55 to 131.97.The encoded proteins contained an N-terminal RNA recognition motif(RRM)and a C-terminal arginine/serine-rich domain(RS).NlSRSF7.1 and NlSRSF7.2 also contained a ZnF_C2HC domain.NlSRs were expressed in various tissues of N.lugens,including salivary glands,integument,fat body,ovaries,and midgut.Among them,NlSRSF1 and NlSRSF2.1 exhibited high relative expression levels in salivary glands.NlSRSF1 was primarily expressed in eggs and 1st instar nymphs,while the remaining NlSR genes were p

关 键 词：褐飞虱 SR蛋白 可变剪接 RNAI 

分 类 号：S435.112.3[农业科学—农业昆虫与害虫防治]