小菜蛾活化蛋白激酶C受体1基因PxyRACK1在变态发育调控中的作用

Function of receptor for activated C kinase 1 gene PxyRACK1 in the regulation of metamorphosis in Plutella xylostella(Lepidoptera:Plutellidae)

【目的】活化蛋白激酶C受体1(receptor for activated C kinase 1,RACK1)参与了多细胞生物体中重要的信号传导,调节生物体的生长发育。本研究旨在克隆小菜蛾Plutella xylostella RACK1基因PxyRACK1,调查其表达模式,明确其对小菜蛾化蛹的影响及其机制。【方法】克隆PxyRACK1的全长cDNA,进行生物信息学分析,并利用qPCR检测其在小菜蛾各发育阶段(卵、1-4龄幼虫、预蛹、蛹和成虫)的表达;通过dsPxyRACK1显微注射小菜蛾4龄幼虫进行RNA干扰,并检测RNAi后PxyRACK1及蛹期特异表达基因PxyBr-Z2/3的表达量;统计死亡率、化蛹率、平均化蛹时间以及蛹重。【结果】小菜蛾PxyRACK1(GenBank登录号:MW160739)序列全长为1148 bp,开放阅读框长960 bp,编码319个氨基酸,具有7个WD40重复序列,每个WD40重复序列包含39~42个氨基酸。PxyRACK1在小菜蛾各发育阶段均有表达,其中4龄第2天幼虫中表达量最高,2日龄蛹中表达量最低。显微注射dsPxyRACK124 h后,处理组4龄幼虫中PxyRACK1和PxyBr-Z2/3表达量较对照组(dsGFP注射组)的分别显著降低了36.26%和83.46%,并且显微注射dsPxyRACK1导致试虫死亡率上升、化蛹推迟、化蛹率降低以及蛹重减轻。【结论】结果说明PxyRACK1参与调控小菜蛾蛹期变态发育。本研究为进一步阐明小菜蛾变态发育的信号调控通路及开发新型昆虫生长调节剂提供了思路。

【Aim】Receptor for activated C kinase 1(RACK1)is involved in the transduction of some crucial signals in multicellular organisms to regulate their growth and development.The aim of the present study is to clone the RACK1 gene PxyRACK1 in the diamondback moth,Plutella xylostella,to investigate its expression pattern and to find out its effect on pupation as well as its underlying mechanism.【Methods】The full-length cDNA of PxyRACK1 was cloned and subjected to bioinformatic analysis,and its expression profiles at different developmental stages(egg,1st-4th instar larvae,prepupa,pupa and adult)of P.xylostella were detected by qPCR.The RNA interference of PxyRACK1 was carried out by microinjecting PxyRACK1 dsRNA into the 4th instar larvae of P.xylostella.After RNAi,the expression levels of PxyRACK1 and pupal specifier PxyBr-Z2/3 were detected,and the mortality,pupation rate,average pupation time and pupal weight of P.xylostella were recorded and calculated.【Results】The full-length cDNA of PxyRACK1(GenBank accession no.:MW160739)of P.xylostella is 1148 bp in length,with an open reading frame of 960 bp encoding 319 amino acids.The predicted amino acid sequence of PxyRACK1 protein contains seven WD40 repeats,each WD40 repeat consisting of 39-42 amino acids.PxyRACK1 was ubiquitously expressed at various developmental stages of P.xylostella,with the highest expression level in the day-24th instar larva and the lowest expression level in the 2 d-old pupa.At 24 h after microinjection of dsPxyRACK1,the expression levels of PxyRACK1 and PxyBr-Z2/3 in the 4th instar larvae in the treatment group were significantly decreased by 36.26%and 83.46%,respectively,as compared to those in the control group(dsGFP injection group),and the microinjection of dsPxyRACK1 resulted in the increase in mortality,delay in pupation and decrease in pupation rate and pupal weight.【Conclusion】The results suggest that PxyRACK1 is involved in the regulation of pupal metamorphosis of P.xylostella.This study provides clues for further exploring of the signal regulation pathway of metamorphosis and developing novel insect growth regulators.