An accelerated miRNA-based screen implicates Atf-3 in odorant receptor expression

Abstract

Large scale genetic screening is tedious and time-consuming. To address this problem, we propose a novel two-tiered screening system comprising an initial \"pooling\" screen that identifies miRNAs whose tissue-specific over-expression causes a phenotype of interest followed by a more focused secondary screen that uses gene-specific RNAi. As miRNAs inhibit translation or direct the destruction of their target mRNAs, any phenotype observed with miRNA over-expression can be attributed to the loss-of-function of one or more target mRNAs. Since miRNA-target pairing is sequence-specific, a list of predicted targets for miRNAs identified in the initial screen serves as a list of candidates for the secondary RNAi-based screen. These predicted miRNA targets can be prioritized by expression pattern, and if multiple miRNAs produce the same phenotype, overlapping target predictions can be given higher priority in the follow-up screen.\n\nSince miRNAs are short, miRNA misexpression will likely uncover artifactual miRNA-target relation-ships. Thus, we are using miRNAs as a tool to accelerate genetic screening rather than focus on the biology of miRNAs themselves. This two-tiered system allows us to rapidly identify individual target genes involved in a phenomenon of interest, often in less than 200 crosses. Here we demonstrate the effectiveness of this method by identifying miRNAs that alter Drosophila odorant receptor expression. With subsequent miRNA target prediction and follow-up RNAi screening we identify and validate a novel role for the transcription factor Atf3 in the expression of the socially relevant receptor Or47b.