Catalysis of carbonic anhydrase IX and pH control in MDA‐MB‐231 breast cancer cells

Abstract

Carbonic anhydrase IX (CAIX) is a membrane‐bound glycoprotein whose expression is considered a marker of tumor hypoxia and associated with tumor acidification. Our goal was to determine if the catalytic properties of native CAIX in MDA‐MB‐231 breast cancer cells are equivalent to those previously described in vitro and to evaluate its catalytic role in pH control. CAIX, like other CA family members, catalyzes the reversible hydration of carbon dioxide: CO2 + H2O ↔ H+ + HCO3−. We directly analyzed CAIX activity by using 18O exchange between CO2 and water as measured by membrane inlet mass spectrometry (MIMS). Data from membrane ghosts showed that the kinetic constants of CAIX catalysis in the membrane environment are very similar to those measured for purified, recombinant, truncated forms, suggesting that CAIX is not affected by the proteoglycan domain or the membrane environment. In intact cells, hypoxia increases CAIX activity compared to normoxia which is associated with increased expression. In the dehydration direction, the catalytic activity of CAIX increases as pH is decreased from pH 9 to pH 6.5. This suggests that at the pH of the tumor microenvironment (pH 6.8), CAIX uses protons to generate CO2. Thus, CAIX may contribute to both the development and maintenance of the acidic microenvironment of hypoxic tumors and thus to the survival fitness of tumor cells. BC073020 (SCF), DK45035 (SCF), and GM25254 (DNS)