A novel calcium-regulated membrane guanylate cyclase transduction system in the olfactory neuroepithelium

This report defines the identity of a calcium-regulated membrane guanylate cyclase transduction system in the cilia of olfactory sensory neurons, whic h is the site of odorant transduction. The membrane fraction of the neuroep ithelial layer of the rat exhibited Ca2+-dependent guanylate cyclase activi ty, which was eliminated by the addition of EGTA. This indicated that the c yclase did not represent a rod outer segment guanylate cyclase (ROS-GC), wh ich is inhibited by free Ca2+. This interpretation was supported by studies with the Ca2+ binding proteins, GCAPs (guanylate cyclase activating protei ns), which stimulate photoreceptor ROS-GC in the absence of Ca2+. They did not stimulate the olfactory neuroepithelial membrane guanylate cyclase. The olfactory neuroepithelium contained a Ca2+ binding protein, neurocalcin, w hich stimulated the cyclase in a Ca2+-dependent fashion. The cyclase was cl oned from the neuroepithelium and was found to be identical in structure to that of the previously cloned cyclase termed GC-D. The cyclase was express ed in a heterologous cell system, and was reconstituted with its Ca2+-depen dent activity in the presence of recombinant neurocalcin. The reconstituted cyclase mimicked the native enzyme. Immunocytochemical studies showed that the guanylate cyclase coexists with neurocalcin in the apical region of th e cilia. Deletion analysis showed that the neurocalcin-regulated domain res ides at the C-terminal region of the cyclase. The findings establish the bi ochemical, molecular, and functional identity of a novel Ca2+-dependent mem brane guanylate cyclase transduction system in the cilia of the olfactory e pithelium, suggesting a mechanism of the olfactory neuroepithelial guanylat e cyclase regulation fundamentally distinct from the phototransduction-link ed ROS-GC.