东方蜜蜂微孢子虫侵染意大利蜜蜂工蜂过程的nce-miR-23928及其靶基因表达谱

Expression profiles of nce-miR-23928 and its target genes in the Nosema ceranae infection of Apis mellifera ligustica workers

摘要:

【背景】东方蜜蜂微孢子虫(Nosema ceranae)专性侵染成年蜜蜂中肠上皮细胞而导致的微孢子虫病给养蜂业造成严重损失。【目的】检测东方蜜蜂微孢子虫nce-miR-23928及其靶基因在侵染意大利蜜蜂(Apis mellifera ligustica)工蜂过程的表达谱，为深入探究nce-miR-23928在东方蜜蜂微孢子虫侵染中的功能及调控机制提供依据。【方法】通过RNAhybrid、miRanda和TargetScan软件预测nce-miR-23928的靶基因。使用Blast工具将上述靶基因比对到基因本体论(gene ontology,GO)、京都基因和基因组百科全书(kyoto encyclopedia of genes and genomes,KEGG)、Nr和Swiss-Prot数据库以获得相应注释。采用实时荧光定量PCR(real time quantitative PCR,RT-qPCR)技术检测nce-miR-23928及其靶基因在东方蜜蜂微孢子虫侵染意蜂工蜂过程中的相对表达量。【结果】相较于接种后1d(1 day post infection,1dpi)，nce-miR-23928的表达量在2dpi基本不变，而在4、6、8dpi均为显著下调(P<0.05)，总体表现出下降的表达趋势。预测到nce-miR-23928的15个靶基因，分别有9、4、15和9条靶基因可注释到GO(3个条目)、KEGG(7条通路)、Nr和Swiss-Prot数据库。与1dpi相比，靶基因ABCT的表达量在2、4、6、8dpi皆显著下调；靶基因SPTK的表达量在4、6、8dpi均显著上调。【结论】明确nce-miR-23928及其靶基因ABCT和SPTK在东方蜜蜂微孢子虫侵染意蜂工蜂过程中的动态表达规律，揭示nce-miR-23928通过正调控ABCT表达和负调控SPTK表达可能在东方蜜蜂微孢子虫的侵染过程中起调节作用。

关键词:东方蜜蜂微孢子虫;蜜蜂;意大利蜜蜂;nce-miR-23928;表达谱;靶基因

Abstract:

[Background] Nosema ceranae exclusively infects midgut epithelial cells of adult bees, and the resulting microsporidiosis causes severe losses to the beekeeping industry. [Objective] This study aimed to determine the expression profiles of nce-miR-23928 and its target genes during the N. ceranae infection of Apis mellifera ligustica workers, and to provide basis for further investigation on the function and regulatory mechanism of nce-miR-23928 during the infection. [Methods] Target genes of nce-mir-23928 were predicted by RNAhybrid, miRanda and TargetScan. Blast was used to perform annotation of the aforementioned target genes in gene ontology (GO), kyoto encyclopedia of genes and genomes (KEGG), Nr and Swissprot. Real time quantitative PCR (RT-qPCR) was employed to detect relative expression of nce-miR-23928 and its target genes. [Results] As compared with the condition at 1 day post infection (1 dpi), the expression of nce-miR-23928 remained unchanged at 2 dpi, but was all down-regulated at 4 6, 8 dpi (P <0.05), presenting an overall reduced expression trend. Additionally, 15 target genes of nce-miR-23928 were predicted, among which 9, 4, 15 and 9 were annotated in GO (3 items), KEGG (7 pathways), Nr and Swiss-prot, respectively. As compared with the condition at 1 dpi, the expression of target gene ABCT was significantly down-regulated at 2, 4, 6, 8 dpi, while the expression of target gene STPK was significantly up-regulated at 4, 6, 8 dpi, displaying an overall elevation trend. [Conclusion] These results illuminated the dynamic expression rules of nce-miR-23928 and its target genes ABCT and SPTK in the N. ceranae infection of A. mellifera ligustica workers, and unraveled that nce-miR-23928 putatively modulated the infection process through positively regulating the expression of ABCT and negatively regulating the expression of STPK.

Key words:Nosema ceranae;honey bee;Apis mellifera ligustica;nce-miR-23928;expression profiles;target genes