UC San Diego

Host vacuolar-type ATPase (VHA)-mediated carbon-concentrating mechanisms (CCMs)that promote photosynthesis have been observed in coral-, sea anemone-, and giant clam- Symbiodiniaceae symbiosis. Similarly, analogous CCMs operate in some free-living secondary endosymbiotic algae, where VHA is present in phagocytic-origin membranes surrounding the chloroplasts. This thesis explored the potential role of VHA in the CCM of cultured Symbiodiniaceae algae Breviolum psygmophilum and Miliolidium leei, which establish symbiosis with corals and foraminifera, respectively. VHA subunit B (VHAB) transcripts were detected in B. psygmophilum and symbiotic algae in the coral Stylophora pistillata. B. psygmophilum acclimated to 60 and 400 µmol photons m-2 s-1 contained similar amounts of VHAB proteins per cell; however, VHAB constituted a higher proportion of total protein under the high light condition. Respirometry experiments indicated that VHA inhibition decreased O2 production in B. psygmophilum cultures when illuminated by the light level they were acclimated to, suggesting a VHA-mediated CCM under these conditions. However, the effects of pharmacological inhibitors varied across replicates and times, suggesting that the CCM is highly responsive to light level, pH, inorganic carbon concentration, algal circadian rhythm and growth phases. Effects of drug concentration and drug incubation time should also be carefully considered. Unfortunately, B. psygmophilum ruptured in the chemical fixative, which prevented establishing the subcellular localization of VHAB. Successful immunolocalization of VHAB in M. leei that fixed properly revealed that VHAB surrounded intracellular spherical structures resembling vacuoles but not the chloroplast. These results highlight morphological and physiological differences between these two species within Symbiodiniaceae.