番茄潜叶蛾卵黄原蛋白受体基因TaVgR在生殖发育调控中的作用

Function of vitellogenin receptor gene TaVgR in the regulation of reproductive development in Tuta absoluta(Lepidoptera:Gelechiidae)

【目的】本研究旨在明确卵黄原蛋白受体(vitellogenin receptor,VgR)在番茄潜叶蛾Tuta absoluta生殖发育过程中的功能,为潜叶类害虫的绿色防控提供候选靶标。【方法】基于番茄潜叶蛾转录组数据,采用RT-PCR扩增TaVgR基因cDNA全序列,并进行生物信息分析;通过RT-qPCR分析TaVgR在番茄潜叶蛾不同发育阶段(1-4龄幼虫、1-7日龄雌蛹和雌成虫)和雌成虫不同组织(头、体壁、前肠、中肠、后肠、卵巢、脂肪体和马氏管)中的表达模式;进一步利用RNAi抑制番茄潜叶蛾雌蛹体内TaVgR的表达,并观测沉默TaVgR基因后番茄潜叶蛾卵巢发育及繁殖力的变化。【结果】克隆获得番茄潜叶蛾TaVgR cDNA(GenBank登录号:MZ682118)序列,其开放阅读框序列长5496 bp,编码1831个氨基酸,推测的蛋白分子量约为206 kD,等电点为5.17,信号肽包含N-端前18个氨基酸残基,并具有典型LDLR家族蛋白保守功能域。RT-qPCR结果显示,TaVgR转录水平随着番茄潜叶蛾龄期的增加逐渐上升,雌成虫羽化后达到最高水平;TaVgR在番茄潜叶蛾雌成虫的卵巢中表达量最高。TaVgR RNAi对初期雌蛹中TaVgR的表达抑制率为62.04%~72.55%,导致卵黄蛋白在卵巢中的沉积受阻,卵巢管和卵粒长度缩短,成虫10日单雌总产卵量及后代卵孵化率降低,最终引起番茄潜叶蛾繁殖力下降。【结论】TaVgR基因在番茄潜叶蛾雌成虫和卵巢中高表达,且沉默该基因严重阻碍其卵巢发育和降低繁殖力。本研究为开发以VgR基因作为靶标的鳞翅目害虫防治新技术奠定了理论基础。

【Aim】This study aims to clarify the function of vitellogenin receptor(VgR)in the reproductive development of the tomato moth,Tuta absoluta,so as to provide a potential target for the green control of leaf miner pests.【Methods】Based on the transcriptomic data of T.absoluta,the full-length cDNA sequence of TaVgR was amplified by RT-PCR and analyzed by bioinformatics methods.The expression patterns of TaVgR in different developmental stages(1st-4th instar larvae,1-7-day-old female pupae,and female adults)and female adult tissues(head,integument,foregut,midgut,hindgut,ovary,fat body,and Malpighian tubules)of T.absoluta were examined by RT-qPCR.RNAi was used to inhibit the expression of TaVgR in female pupae,and the changes in the ovarian development and fecundity of T.absoluta were observed after silencing of TaVgR.【Results】The cDNA sequence of TaVgR(GenBank accession no.:MZ682118)of T.absoluta was cloned.It contains an open reading frame of 5496 bp in length,encoding 1831 amino acids with the predicted molecular weight of 206 kD and isoelectric point of 5.17.The deduced protein of TaVgR has a signal peptide with 18 amino acid residues at the N-terminus and harbours the typical conserved functional domains of LDLR family proteins.The RT-qPCR results showed that the transcriptional level of TaVgR gradually rose with the increase of age of T.absoluta and reached the peak after female adult emergence.TaVgR exhibited the highest expression level in the ovaries of female adults of T.absoluta.RNAi of TaVgR inhibited the expression level of TaVgR in the early female pupae by 62.04%-72.55%,led to disruption of the deposition of yolk protein in the ovary,shortened the lengths of ovarioles and eggs,and reduced the total number of eggs laid per female adult in 10 d and the egg hatching rate of offspring,resulting in a decline in fertility.【Conclusion】TaVgR is highly expressed in female adults and ovaries of T.absoluta.Silencing of TaVgR severely blocks the ovarian development of T.absoluta and impairs its fecundity.This study lays a theoretical foundation for the development of new control strategies of lepidopteran pests with VgR gene as the target.