[B-CG05] Diversity and ecology of marine protists -from past to present

[BCG05-03] Carbonic anhydrase in Foraminifera: Perspective on one of major role of ‘vital effect’

*Yurika Ujiie¹, Yoshiyuki Ishitani², Yukiko Nagai^2,3, Yoshihiro Takaki², Takashi Toyofuku^2,4, Shun´ichi Ishii² (1.CMCR, Kochi University, 2.X-star, JAMSTEC, 3.National Museum of Nature and Science, 4.Tokyo University of Marine Science and Technology)

Keywords:carbonic anhydrase, calcification, stable oxygen and carbon isotopes

Foraminifera account for ~25% of calcium carbonate (CaCO₃) production in global carbon cycle, owing to their abundance in both seafloor and pelagic environments. Rotaliida, the major foraminiferal order in the class Globothalamea, has been widely used in geological studies as a proxy for modern and paleo-environments because the chemical and isotopic composition of its CaCO₃ tests (i.e., stable oxygen and carbon isotopes: d¹⁸O, and d¹³C) is influenced by temperature and salinity of ambient seawater. However, the non-equilibrium state of chemical compounds between foraminiferal tests and the external environment poses a problem for such measurements due to ‘vital effect’: physiology and diet of organisms dictat the uptake of inorganic carbon along with environments. To date, the mechanism of this vital effect has been masked because it is unclear how inorganic carbon may be rerouted from other metabolic functions toward calcification. Here we conducted comparative transcriptomics between calcification and non-calcifying stages of Rotaliida foraminifer to identify calcification-related genes and reconstructed their metabolic pathway concerning calcification.
Carbonic anhydrase (CA) and aquaporin 4 (AQP4) genes showed significant expression during calcification in the transcriptomics. The enzyme CA catalyzes the interconversion of H₂O + CO₂ to HCO₃^- + H⁺. The integral membrane protein AQP4 allows the passage of water molecules through the cell membrane and supplies H₂O for further HCO₃^-production around the cell membrane. CO₂ can be diffused from seawater as well as calcification sites and produced during metabolic processes. The CA and AQP4 expressions imply the intracellular generation of HCO₃^-. The phylogenetic analysis of CAs indicated that Rotaliida foraminifer has two out of five CA families: α-CA and b-CA members. α-CAs are involved in CO₂/HCO₃^- interconversion and calcification in mollusks, sea urchins, and corals. In mollusks, calcification-related α-CAs contain acidic low-complexity regions (LCRs), which could bind Ca²⁺ leading to CaCO₃ nucleation. While, Rotaliida’s α-CAs formed two clades, one of which (Rotaliida-αCA1) is a novel independent clade from metazoans and some has acidic LCRs. Rotaliida-αCA1 seems to catalyze CO₂/HCO₃^- interconversion which is related to foraminiferal calcification. Moreover, our in silico protein motif predictions indicated that Rotaliida-αCA1 comprised both secretory α-CAs and intracellular α-CAs. The former is likely involved in HCO₃^- generation and calcium-binding at the extracellular calcification site. The latter does not interact with Ca²⁺ as they lack acidic LCRs; hence, they likely participate in HCO₃^- production in the cytosol. These different distributions of α-CA enable the utilization of CO₂ derived from both extracellular and intracellular origins. The multiple sources, in particular those of CO₂, serve as a physiological control mechanism for d¹⁸O and d¹³C in foraminiferal tests via HCO₃^- production. Such physiology could be partly responsible for the non-equilibrium state of d¹⁸O and d¹³C between foraminiferal tests and the environment observed in geochemical studies.