RNA-seq Profiling of Small Numbers of Drosophila Neurons

Drosophila melanogaster has a robust circadian clock, which drives a rhythmic behavior pattern: locomotor activity increases in the morning shortly before lights on (M peak) and in the evening shortly before lights off (E peak). This pattern is controlled by similar to 75 pairs of circadian neurons in the Drosophila brain. One key group of neurons is the M-cells (PDF+ large and small LN(v)s), which control the M peak. A second key group is the E-cells, consisting of four LN(d)s and the fifth small LNv, which control the E peak. Recent studies show that the M-cells have a second role in addition to controlling the M peak; they communicate with the E-cells (as well as DN(1)s) to affect their timing, probably as a function of environmental conditions (Guo, Cerullo, Chen, & Rosbash, 2014). To learn about molecules within the M-cells important for their functional roles, we have adapted methods to manually sort fluorescent protein-expressing neurons of interest from dissociated Drosophila brains. We isolated mRNA and miRNA from sorted M-cells and amplified the resulting DNAs to create deep-sequencing libraries. Visual inspection of the libraries illustrates that they are specific to a particular neuronal subgroup; M-cell libraries contain timeless and dopaminergic cell libraries contain ple/TH. Using these data, it is possible to identify cycling transcripts as well as many mRNAs and miRNAs specific to or enriched in particular groups of neurons.