Characterization of MicroRNA's mediating olfactory neurogenesis

MicroRNAs (miRNAs) are a class of small RNAs expressed throughout metazoan species and have recently been recognized as a mechanism for post-transcriptional gene control. Although they are encoded by hundreds of genes and regulate thousands of targets in each species, the mechanisms by which these miRNAs shape developmental processes are not well characterized. In order to elucidate one such mechanism, we investigated whether miRNAs regulate an experimentally and genetically tractable model of neural development, olfactory neurogenesis in rodents. We characterized the repertoire of miRNAs expressed during olfactory epithelium development and identified several olfactory-enriched miRNAs. In order to establish that miRNAs are critical for olfactory neurogenesis, we developed a genetic strategy to conditionally eliminate the functions of all miRNAs in either olfactory progenitor cells or olfactory sensory neurons. These analyses revealed that miRNAs are essential for olfactory neurogenesis, but are not essential for the function of mature neurons. In order to probe the contributions of individual miRNAs to olfactory neurogenesis, we developed an antisense morpholino knock down strategy to specifically ablate the function of individual olfactory miRNAs in zebrafish, a species whose olfactory system is remarkably similar to that of rodents. Functional ablation of a specific family (miR-200) recapitulated the olfactory neurogenesis defects observed when all miRNA functions are eliminated in mouse or zebrafish olfactory progenitors, indicating that the miR-200 family is required for proper olfactory neurogenesis. In order to identify the molecular targets of miR-200 family members and other olfactory miRNAs, we systematically identified genes that accumulated in mouse olfactory progenitor cells lacking miRNA function. These analyses revealed a set of genes whose 3'UTRs are highly enriched for binding sites of olfactory miRNAs, including miR-200 family members. Moreover, the genes within this set are predominantly negative regulators of cell growth and of specific signaling pathways that have been previously described to be critical for olfactory neurogenesis. Thus, miR-200 and other olfactory expressed miRNAs appear to control the expression of a suite of negative regulators during olfactory neurogenesis that allows these cells to terminally differentiate into olfactory neurons.