An olfactory sensory neuron (OSN) expresses selectively one member from a repertoire of approximately 1000 odorant receptor (OR) genes and projects its axon to a specific glomerulus in the olfactory bulb. The axonal target depends on the OR gene that the neuron expresses. Eleven years after the discovery of OR genes, the mechanisms underlying OR gene choice and expression remain enigmatic. Transgenic model systems were developed for MOR23 and M71 OR genes. Minigenes of 9 kb recapitulate the features of OR gene expression, including punctate, zonally restricted expression and axonal projection to the cognate glomerulus. Data from comparative genomics and transgenesis suggested that 300 bp promoter regions displaying characteristic sequence motifs mediate choice and expression of these two OR genes. We propose to show conclusively that these small regions, now in the absence of the OR coding region, are sufficient to drive expression of the corresponding transgene, and to extend our analysis to other OR genes.
Our Specific Aim 1 is to define for five different OR genes expressed in three distinct zones the minimal cis-acting sequences that confer OR gene choice and expression, with the features of punctate and zonally restricted expression. These sequences are hypothesized to specify spatial (appropriate epithelial zone) and temporal parameters of OR expression, as well as of transcription competence. We next propose to test the impact these defined sequences have on control of axon guidance. Genetic manipulations of OR genes replacing the coding sequence of a recipient OR locus with a donor OR sequence resulted in a shift of the tagged glomerulus to a new position corresponding neither to the donor or recipient OR. This lead to the notion that the OR has an instructive role but is not the sole determinant in target determination. We hypothesize that conferring additionally to the donor OR a 'native' control of expression will redirect axonal projections to the donor glomeruli.
Our Specific Aim 2 is to test whether the simultaneous swaps at an OR recipient locus of both the coding sequence and the approximately300 bp control elements (as defined in Specific Aim 1) surrounding the TSS of a donor OR gene, redefine the glomerular address to that of the donor OR. In summary, we hypothesize that the OR, together with the parameters (spatial, temporal and of """"""""cell type"""""""") of its expression determine axonal targeting. Insight into the principles underlying OSN identity and connectivity may bear relevance to other important neuronal populations.
