Expression and physiological role of stomatin-domain proteins in the olfactory epithelium

The olfactory system is a chemosensory system that detects volatile molecules, called odorants, responsible for valuable information from the environment such as sources of food, predators or hazardous substances. The detection of odorants is carried out by the olfactory sensory neurons, the primary cellular sensors. The vast variety of molecules that the olfactory system can detect is due to the expression of a huge repertoire of molecular receptors that directly bind the odorants, called olfactory receptors. These are the primary molecular sensors which are expressed mainly in the ciliary membrane of the olfactory sensory neurons. Each olfactory sensory neuron expresses only one type of olfactory receptor, and all neurons transduce the odorant signals by a common signaling pathway. The core elements of this signaling pathway are well established: the binding of odorants by an olfactory receptor leads to the sequential activation of Gαolf and adenylyl cyclase III. The increase of ciliary cAMP opens CNG channels, and the entry of calcium through these channels leads to the opening of the calcium-activated chloride channel TMEM16B allowing an efflux of chloride ions from the cell. The opening of CNG and TMEM16B channels underlies the depolarizing receptor potential induced by the detection of odorants in olfactory sensory neurons. In addition to the core elements of the odorant transduction machinery, other proteins could play a role in the detection of odorants, especially as potential regulators of the transduction machinery. For example, some members of the stomatin family are highly expressed in the olfactory epithelium. Stomatin-domain proteins have emerged as general regulators of ion channels and transporters and have been shown to be involved in transduction mechanisms in other sensory systems. Stomatin-domain proteins are highly conserved and anciently evolved membrane proteins. In the mammalian genome, five members have been identified: Stomatin, stomatin-like protein-1 (STOML-1), stomatin-like protein-2 (STOML-2) stomatin-like protein-3 (STOML -3) and Podocin. Some members of this family have been shown to be expressed in the olfactory epithelium and Stomatin and STOML-3 mRNAs are present at very high levels. Stomatin and STOML-3 proteins have been isolated in calcium-dependent signaling complexes in cilia extracts from the olfactory epithelium and STOML-3 proteins have been shown to localize mainly in the cilia of the neurons, where transduction of odorants takes place, and to interact with ACIII. One of the main aims in this thesis was confirm and extend our understanding about the expression and localization of the stomatin family in the olfactory epithelium. we confirmed the localization of STOML-3 in the cilia of the olfactory sensory neurons. Moreover, we found that Stomatin is present also in these structures. In contrast, we revealed that STOML-1 and STOML-2 localizes in the central layer of the olfactory epithelium, in the soma of the neurons. To study the role of these proteins in the function of the olfactory epithelium we used a loss of expression approach. We used two KO mice models: a STOML-3 KO and a Stomatin/STOML-1/STOML-3 triple KO. First, we studied in detail the development in these KO animals. We did not find differences in the number of olfactory neurons, apoptotic, globose or horizontal basal cells and in the morphology of the cilia of the neurons. We did not observe changes in the expression and the localization of several members of the transduction machinery and potential targets of stomatin-domain proteins. Then, we measured the physiological response to odorants in KO animals by performing electro-olfactogram recordings. We found a reduction in the amplitude of the response in both KO animals, especially in older mice. The magnitude of the reduction was similar in both KO mice, suggesting that STOML-3 is the main responsible for this effect. The results in this thesis establish the first evidences that stomatin-domain proteins modulate the odorant response in mice and provide the foundation for future work aimed at clarifying the physiological role of the stomatin family in the olfactory system.