Genome-wide identification of long non-coding RNAs and their regulatory networks involved in Apis mellifera ligustica response to Nosema ceranae infection
Long non-coding RNAs (lncRNAs) are a diverse class of transcripts that structurally resemble mRNAs but do not encode proteins, and lncRNAs have been proved to play pivotal roles in a wide range of biological processes in animals and plants. However, knowledge of expression pattern and potential role of honeybee lncRNAs response to Nosema ceranae infection is completely unknown. Here, we performed whole transcriptome strand-specific RNA sequencing of normal midguts of Apis mellifera ligustica workers (Am7CK, Am10CK) and N. ceranae-inoculated midguts (Am7T, Am10T), followed by comprehensive analyses using bioinformatic and molecular approaches. A total of 6353 A. m. ligustica lncRNAs were identified, including 4749 conserved lncRNAs and 1604 novel lncRNAs. These lncRNAs had low sequence similarities with other known lncRNAs in other species; however, their structural features were similar with counterparts in mammals and plants, including shorter exon and intron length, lower exon number, and lower expression level, compared with protein-coding transcripts. Further, 111 and 146 N. ceranae-responsive lncRNAs were identified from midguts at 7 day post inoculation (dpi) and 10 dpi compared with control midguts. 12 differentially expressed lncRNAs (DElncRNAs) were shared by Am7CK vs Am7T and Am10CK vs Am10T comparison groups, while the numbers of unique ones were 99 and 134, respectively. Functional annotation and pathway analysis showed the DElncRNAs may regulate the expression of neighboring genes by acting in cis. Moreover, we discovered 27 lncRNAs harboring eight known miRNA precursors and 513 lncRNAs harboring 2257 novel miRNA precursors. Additionally, hundreds of DElncRNAs and their target miRNAs were found to form complex competitive endogenous RNA (ceRNA) networks, suggesting these DElncRNAs may act as miRNA sponges. Furthermore, DElncRNA-miRNA-mRNA networks were constructed and investigated, the result demonstrated that part of DElncRNAs were likely to participate in regulating the material and energy metabolism as well as cellular and humoral immune during host responses to N. ceranae invasion. Finally, the expression pattern of 10 DElncRNAs was validated using RT-qPCR, confirming the reliability of our sequencing data. Our findings revealed here offer not only a rich genetic resource for further investigation of the functional roles of lncRNAs involved in A. m. ligustica response to N. ceranae infection, but also a novel insight into understanding host-pathogen interaction during microsporidiosis of honeybee.