Key laboratory of Horticultural Plant Biology (MOE) and Institute of Urban and Horticultural Entomology, College of Plant Science and Technology, Huazhong Agricultural University, Wuhan, China.
Department of Agriculture, Abdul Wali Khan University Mardan, Khyber Pakhtunkhwa, Pakistan.
Insect Sci. 2019 Apr;26(2):229-239. doi: 10.1111/1744-7917.12542. Epub 2017 Dec 7.
MicroRNAs (miRNAs) regulate various biological processes during insect development; however, their role in larval-pupal development in oriental fruit fly, Bactrocera dorsalis (Hendel) remains unknown. In the current study, we address the biological function of a conserved miRNA, Bdo-Let-7 in the regulation of BdE75 gene, which belongs to the ecdysone signaling pathway and participates in the larval-pupal development in B. dorsalis. Using dual luciferase reporter assay in HEK293T cells we show that Bdo-Let-7 miRNA interacts with the 3' untranslated region of BdE75 gene and suppresses its expression. The Bdo-Let-7 and BdE75 are also co-expressed in the larval-pupal stages and in different tissues of B. dorsalis. In in vivo experiments, the injection of Bdo-Let-7 agomir and antagomir in third instar larvae down- and up-regulated the expression of BdE75, respectively. The 20-hydroxyecdysone (20E) injection assay shows that 20E up-regulated the expression of Bdo-Let-7 on the 5th day of the larvae. Moreover, abnormal pupation and eclosion were observed after larval Bdo-Let-7 antagomir injection. Based on these results, we show that Bdo-Let-7 regulates the ecdysone signaling pathway through the exact dose of BdE75 gene, and is indispensable for normal larval-pupal development in B. dorsalis.
微小 RNA（miRNA）在昆虫发育过程中调节各种生物学过程；然而，它们在东方果实蝇幼虫-蛹发育中的作用尚不清楚。在本研究中，我们研究了一个保守的 miRNA，Bdo-Let-7 在调节 BdE75 基因中的生物学功能，BdE75 基因属于蜕皮激素信号通路，参与 B. dorsalis 的幼虫-蛹发育。我们通过在 HEK293T 细胞中的双荧光素酶报告基因检测实验表明，Bdo-Let-7 miRNA 与 BdE75 基因的 3'非翻译区相互作用并抑制其表达。Bdo-Let-7 和 BdE75 也在幼虫-蛹期和 B. dorsalis 的不同组织中共同表达。在体内实验中，第三龄幼虫注射 Bdo-Let-7 agomir 和 antagomir 分别下调和上调 BdE75 的表达。20-羟基蜕皮酮（20E）注射实验表明，20E 在幼虫第 5 天上调了 Bdo-Let-7 的表达。此外，幼虫注射 Bdo-Let-7 antagomir 后会出现异常蛹化和羽化。基于这些结果，我们表明 Bdo-Let-7 通过 BdE75 基因的精确剂量调节蜕皮激素信号通路，对于 B. dorsalis 的正常幼虫-蛹发育是必不可少的。
