Chemosensitive areas of the mammalian brainstem regulate breathing and are stimulated by small increases in carbon dioxide (CO2) levels in arterial blood. The genetic and molecular bases for this chemosensitivity are poorly understood. C. elegans is an excellent genetic model for probing this question because it possesses CO2-sensitive neurons - the BAG neurons - that mediate stereotyped behavior and whose cell physiology can be studied in vivo. BAG cell fate is determined by ETS-5, an ETS-domain transcription factor. Strikingly, the mammalian homolog of ETS-5, Pet1, is required for the development of CO2-chemosensitive brain regions, and Pet1 mutants have defects regulating breathing in response to CO2 challenges. To find molecules important in CO2 sensing in C. elegans and perhaps in vertebrates, I analyzed the direct transcriptional targets of ETS-5 using ChIP-seq. Then, to clarify which of these targets are being directly regulated by ETS-5 in BAG, I performed mRNA-seq on wild type versus ets-5 mutant BAG neurons. To determine which of these targets are functionally important for CO2 sensing, I performed a behavioral screen for CO2 avoidance defects. RGS-6 is a G-protein activating protein that is a direct ETS-5 transcriptional target that appears to be down-regulated in an ets-5 mutant background, and rgs-6 mutation also results in an avoidance defect. To better understand the role of RGS-6 in CO2 sensing, I will (1) determine the expression pattern, site, and time of action of rgs-6 in CO2 sensing, (2) place RGS-6 in a GPCR pathway, and (3) determine the physiological role of RGS-6 in CO2 sensing. The proposed studies will integrate molecular genetics, genomics, and in vivo functional imaging to elucidate the role of a novel RGS protein in CO2 sensing. Because Pet1-like factors are conserved between C. elegans and humans, our studies will likely elucidate mechanisms required for the function of chemosensitive neurons in the brain, which play critical roles in regulating breathing and whose dysfunction is linked to fatal apneas.
Neurons that sense the respiratory gas carbon dioxide (CO2) play critical roles in animal physiology, most notably in the control of breathing. We propose studies of an invertebrate model organism with powerful genetics - the nematode C. elegans - to determine molecular mechanisms required for the development and function of CO2-sensing neurons. Because a conserved factor is required for development of CO2-sensing neurons in both C. elegans and mammals, our studies will advance understanding of a critical chemosensory modality whose dysfunction in humans can cause fatal defects in the respiratory motor program.
