Age and Behavior-Dependent Differential miRNAs Expression in the Hypopharyngeal Glands of Honeybees (Apis mellifera L.)

SIMPLE SUMMARY: The hypopharyngeal glands (HPGs) are a pair of aciniform glands that are located in the frontal area of the heads of worker bees (Apis mellifera L.) that exhibit age and behavior-dependent development. Little is known about whether/how miRNAs regulate the HPGs development. In this study, small RNA sequencing was employed to analyze the miRNA profiles of HPGs in newly-emerged bees (NEB), nurse bees (NB), and forager bees (FB). We found that there were a total of 31 known miRNAs differentially expressed among the three stages, which might have regulatory roles in the growth and development, protein synthesis, and carbohydrate and energy metabolism in the HPGs. Additionally, the downregulation of ame-miR-184-3p and ame-miR-252a-5p in nurse bees may be involved in royal jelly secretion, while the lower expression of ame-miR-11-3p and ame-miR-281-3p in forager bees are responsible for honey processing. ABSTRACT: This study aims to investigate the expression differences of miRNAs in the hypopharyngeal glands (HPGs) of honeybees at three developmental stages and to explore their regulation functions in the HPGs development. Small RNA sequencing was employed to analyze the miRNA profiles of HPGs in newly-emerged bees (NEB), nurse bees (NB), and forager bees (FB). Results showed that a total of 153 known miRNAs were found in the three stages, and ame-miR-276-3p, ame-miR-375-3p, ame-miR-14-3p, ame-miR-275-3p, and ame-miR-3477-5p were the top five most abundant ones. Furthermore, the expression of 11 miRNAs, 17 miRNAs, and 18 miRNAs were significantly different in NB vs. FB comparison, NB vs. NEB comparison, and in FB vs. NEB comparison, respectively, of which ame-miR-184-3p and ame-miR-252a-5p were downregulated in NB compared with that in both the FB and NEB, while ame-miR-11-3p, ame-miR-281-3p, and ame-miR-31a-5p had lower expression levels in FB compared with that in both the NB and NEB. Bioinformatic analysis showed that the potential target genes of the differentially expressed miRNAs (DEMs) were mainly enriched in several key signaling pathways, including mTOR signaling pathway, MAPK signaling pathway-fly, FoxO signaling pathway, Hippo signaling pathway-fly. Overall, our study characterized the miRNA profiles in the HPGs of honeybees at three different developmental stages and provided a basis for further study of the roles of miRNAs in HPGs development.