Crispr-Cas9 Knockdown Of Octopamine Beta Receptor Subtype 2 To Understand Its Role In Honey Bee Appetite Regulation

Official URL: https://risc01.sabanciuniv.edu/record=b3205745

Honey bee (Apis mellifera) foragers rely heavily on rapidly changing energetic demands during their foraging activities, which is reflected by the amount of trehalose that is in the hemolymph, and are particularly susceptible to stressors while foraging away from the hive. To meet their dynamic needs of energy while foraging, we suspect that these bees have a direct link between hemolymph trehalose levels and appetite regulation, mediated by octopamine levels in the brain. In order to establish a cause-and-effect relationship between octopamine and appetite regulation, we used CRISPR-Cas9 vectored by baculovirus, to knockdown the octopamine beta subtype 2 receptor in the brain, and then measured appetite levels after starvation, using the Proboscis Extension Response (PER) assay. We found that at three days post injection, there were significantly lower appetite levels and octopamine beta subtype 2 receptor gene expression, while there was higher GFP (Green Fluorescence Protein) signal in the brain. Taken together, our findings suggest that we successfully delivered the CRISPR-Cas9 system and knockdowned the octopamine beta subtype 2 receptor in the brain that is likely to be found in the Kenyon cells. In addition, we established that the octopamine beta subtype 2 receptor is involved in appetite regulation of the honey bee and this is likely to be independent of the glucose-signaling pathway found in vertebrates. With this proof of concept established, other targets for gene editing in adult honey bees are now possible using the CRISPR-Cas9 system.