The Role Of RNA Binding Proteins And CRISPR/CAS9 As A Gene Editing Tool In Drosophila Nociception

Drosophila melanogaster is a powerful model organism to study nociception. The compact and easily manipulated genome provides an opportunity to determine the function of molecules involved in basal and sensitized nociception in both larval and adult animals. Using the GAL4/UAS system, genetic knockdown with RNAi and knockout with CRISPR/Cas9 are possible to pinpoint specific molecular mechanisms and cellular processes within nociceptors that are implicated in nociception. The three main objectives of this work were to: elucidate the impact that three RNA binding proteins have on basal nociception, establish a transgenic fly line capable of inducing Cas9-mediated knockout of specific genes, and to validate a protocol based on a previously published assay to measure thermal nociception in adults. The expression of SC35, an exon-inclusion splicing factor; LaRP4B, a translation stimulator; and eIF2a, a translation regulator, were each required for thermal nociception. Cas9 expression led to sgRNA-independent effects such as severe defects in dendrite morphology. The adult thermal nociception assay was validated and similar results to the original publication were reproduced. Importantly, the findings made with the Drosophila model can be directly applicable to chronic pain in humans due to DNA sequence homology and the conserved function of proteins across species.