Symbiotic dinoflagellates of the giant clam, Tridacna squamosa, express an extracellular alpha carbonic anhydrase associated with the plasma membrane to promote HCO3− dehydration and CO2 uptake during illumination

Giant clams generally harbor phototrophic Symbiodiniaceae dinoflagellates of genera Symbiodinium, Cladocopium, and Durusdinium . The coccoid symbiotic dinoflagellates (zooxanthellae) reside extracellularly inside the lumen of zooxanthellal tubules in the colorful outer mantle. They obtain from the host inorganic carbon (C i ) for photosynthesis and supply photosynthate to the host. The outer mantle has a host-derived carbon concentration mechanism (CCM) to facilitate the transport of C i from the hemolymph into the luminal fluid. To regulate C i uptake, the symbionts probably possess their own CCMs that comprise an extracellular alpha carbonic anhydrase ( αCA ) and a proton transporter. Indeed, we obtained from the outer mantle of the giant clam, Tridacna squamosa , three complete cDNA coding sequences of a membrane-associated αCA derived from Symbiodinium ( Symb-αCA ) , Cladocopium ( Clad-αCA ), and Durusdinium ( Duru-αCA ), which consisted of 2808, 2847, and 2829 bp, respectively. The respective encoded proteins had 935 (104.7 kDa), 948 (106.1 kDa), and 942 (105 kDa) amino acids, each containing a transmembrane domain. The outer mantle had the highest level of Duru-αCA transcripts. Phenogramic analyses denoted Duru-αCA as an extracellular CA closely associated with human CA4 and had a dinoflagellate-origin. Using an antibody that could react comprehensively with zooxanthellae-αCAs (Zoox-αCA) derived from all three genera of dinoflagellate, immunofluorescence microscopy revealed immuno-labeling at the dinoflagellate’s plasma membrane. As Symb-αCA, Clad-αCA, and Duru-αCA possessed extracellular catalytic domains, they could catalyze the dehydration of HCO 3 − to CO 2 in the luminal fluid. Importantly, illumination led to significant increases in the gene and protein expression levels of Zoox-αCA /Zoox-αCA in the outer mantle of T. squamosa . Taken together, Zoox-αCA could be part of the symbiont’s light-enhanced external CCM to promote and regulate the acquisition of C i from the host for photosynthesis. Our results also suggested that the potentials of generating CO 2 adjacent to the symbionts’ plasma membrane could differ among different phylotypes of Symbiodinium and Cladocopium .