UC Berkeley

The gustatory system is primarily involved in feeding, allowing animals to distinguish nutrients from toxins. In the fruit fly Drosophila melanogaster, neurons of the gustatory system also function during egg-laying and courtship behavior, and are broadly involved in contact-mediated chemical recognition. Gustatory input in Drosophila occurs at structures called taste bristles, which are located on the proboscis, legs, wing margins, and ovipositor. Four chemosensory neurons are housed at the base of each bristle, three of which faithfully respond to specific modalities: sugars, bitters, or water. However, the function of the fourth chemosensory neuron, and the molecular mechanism responsible for its activation, remained elusive. Further, the physical properties of molecules responsible for sensory neuron detection is poorly understood.

Microarray analyses identified two genes encoding epithelial sodium channels (deg/ENaCs), PPK23 and PPK29, which show co-expression in the fourth taste neuron and are not expressed in any other tissues. These genes were exciting candidates for defining sensory neuron specificity, as a related channel, PPK28, confers water sensitivity to taste neurons. Initial expression studies revealed that these genes were co-expressed with the transcription factor fruitless, a critical molecule in the organization or sexual dimorphism in the fly. We hypothesized that PPK23 and/or PPK29 may function in pheromone detection and contribute to aspects of courtship behavior. The second chapter of this thesis demonstrates that these genes are imperative for mate discrimination during courtship, and the cells that express them serve as one of the earliest filters for the neural circuitry that leads to courtship enhancement or courtship inhibition. Finally, cells that express these genes respond directly to pheromone application in a male or female specific manner, suggesting the balance between attraction and repulsion at the sensory level may contribute to the overall decision of the organism.

How do pickpocket subunits contribute to mate discrimination behavior? The third chapter of this thesis is an exploration into subunit expression specificity. Calcium imaging studies suggest a heterogeneity in PPK23 cells. In order to separate attractive and repulsive circuits at the sensory level, we ascertained the behavioral genetic interactions between multiple subunits. We found that the population of cells expressing the subunit PPK25 responds exclusively to female pheromone molecules and is also critical for female receptivity. This provides direct genetic access to this component of the courtship circuit.

The function of pickpocket ion channels in signal transduction has yet to be demonstrated. PPK28 detects low osmolarity and has been shown to function when misexpressed in neurons as well as in heterologous systems. However whether it is a direct transducer of osmolarity or perhaps an amplifier of another protein is not known. In the fourth chapter, we delve into the mechanism of gating of pickpocket channels using PPK28 as a model. Our data suggest it is a direct transducer of mechanical force on the membrane and provides an exciting example to study other Deg/ENaC proteins in the role of mechanosensation.