Tolerance of Oocystis nephrocytioides to copper: intracellular distribution and extracellular complexation of copper

Several mechanisms have been proposed to explain how algae can tolerate heavy metals. In order to better understand the mechanisms determining metal tolerance, we examined the interaction of copper with two strains of the copper-tolerant green algae Oocystis nephrocytioides, isolated from algal communities differing only in copper exposure. The strains were cultured in chemically-defined media containing 0.04 μM Cutotal (pCu 12.4) or 2 μM Cutotal (pCu 10.6). Growth, photosynthesis rate, content of chlorophyll a and b, copper accumulation, its cellular distribution and ultrastructural localization, as well as the influence of algal growth on extracellular copper complexation were determined. Both strains had comparable growth and photosynthesis rates. The cellular content of both chlorophyll a and b was reduced, by roughly the same extent, at pCu 10.6 compared to pCu 12.4. Copper titration of the media indicated the production of copper-complexing ligands by O. nephrocytioides cultured at pCu 12.4 that increased with increased algal density during cell growth. No additional ligands were detected at pCu 10.6. Copper-complexing ligands had a conditional stability constant of K = 1013 at pH 7.3. The intracellular concentration of copper in O. nephrocytioides was 80 μM at pCu 12.4 and increased to 7.5 mM at pCu 10.6. The proportion of intracellular Cu accumulated increased from 8% of total Cu content at pCu 12.4 to 60% at pCu 10.6. By electron spectroscopic imaging, intracellular Cu was detected in the thylakoids and the pyrenoid of O. nephrocytioides cells. The results indicate that the tolerance of O. nephrocytioides to Cu is constitutive and does not need to be induced by previous exposure to Cu. We propose that accumulation and sequestration of Cu in thylakoids and, to a lesser extent, adsorption of copper to the algal cell surface represent the most important tolerance mechanism, for O. nephrocytioides.