miR92b and miR-9a the novel candidates as regulators of the positive reproductive status in honeybees

Abstract

In a queenright colony of Apis mellifera the workers do not reproduce. However, the workers can activate their ovaries in response to the absence of the queen. However the genetic cascade behind this process is not fully understood. In view of miRNA multiple roles in gene regulation of a variety of developmental and reproductive processes, we investigated their occurrence by analyzing 35 million reads (Illumina) of a small RNA library from active ovaries of queenless A. mellifera workers. A set of 168 miRNA previously identified in honeybees were found. Using qPCR we compared the expression of let-7, miR-184, miR-92b and miR-9a (among the most differentially represented in the library and linked to reproductive and caste differentiation processes) in active and inactive worker ovaries. The miR-184 presented consistently highest expression levels in both samples. Despite its described function in Drosophila egg formation, our data suggest that miR-184 is also important in general ovary physiology.

In addition, miR-184 showed the most stable expression rates in all analyzed samples and was used as internal control to normalize the expression of other miRNAs. The involvement of let-7 in reproduction, hormonal signaling and developmental timing is known for several metazoans. Since let-7 and miR-184 showed to be expressed at similar levels in both ovary types, we postulate they govern ovary basal metabolism independent of the activation status. The expression of miR-92b and miR-9a were significantly higher in active than inactive ovaries (p <0.05). These miRNAs are related to cell cycle control in embryonic stem cells and caste differentiation, respectively. Considering that egg laying workers behave as a queen as well as their active ovaries mimics the function of a queen ovary, our results add two novel candidates as regulators of the molecular cascade leading to maintaining the positive reproductive status in honeybee female castes.