I have been studying an apparently mutualistic endosymbiosis, between a widely distributed family of marine invertebrate animals (molgulid ascidian tunicates) and a funguslike protist, Nephromyces. Nephromyces inhabits the ductless "renal sac", surrounding the urate-containing concretions conspicuously accumulated in this molgulid organ. Nephromyces cells possess high levels of urate oxidase activity, and are thus metabolizing the renal sac urate. The intracellular site of this activity in Nephromyces is unknown; paradoxically, peroxisomes, the usual site of urate oxidase in eukaryotes, are not obvious in Nephromyces. I have unexpectedly discovered a third partner in this endosymbiosis: intracellular bacteria in the cytoplasm of Nephromyuces. I hypothesize that the activities of these bacteria are of profound consequence to the biology of the symbiosis as a whole. By adapting techniques developed to study compartmentation of ureide metabolism in legume-rhizobial symbioses, I plan to develop cytochemical and immunological techniques to study the activities of these bacteria in my system. I well test the hypothesis that the bacteria are the site of urate oxidase activity in Nephromyces, and, more generally, that the bacteria are serving as Nephromyces' peroxisomes. I will also test for the presence of allantoinase activity (the next step in the uricolytic pathway) among all three partners of the symbiosis. The general technical approaches developed in this work should be applicable to eventual elucidation of other steps in urate synthesis and degradation in the symbiosis, and of other possible metabolic activities of the bacteria; data gathered through these techniques will also be relevant to elucidation of the physiological dynamics of he several other, less studied, urate-associated endosymbioses found in both marine and terrestrial ecosystems.
