Pools of cadmium in Chlamydomonas reinhardtii revealed by chemical imaging and XAS spectroscopy

The green micro-alga Chlamydomonas reinhardtii is commonly used as a model to investigate metallic stress in photosynthetic organisms. The aim of this study was to explore processes implemented by three C. reinhardtii strains to cope with cadmium (Cd), and particularly to evidence Cd sequestration in the cell. For that, we used a combination of subcellular fractionation and chemical imaging (micro X-ray fluorescence (μXRF) and transmission electron microscopy (TEM/X-EDS)) to identify subcellular compartments of Cd accumulation, and X-ray absorption spectroscopy (XAS) to determine chemical Cd speciation. C. reinhardtii wild type strain 11/32b (wt), a newly design strain (pcs1) expressing a modified phytochelatin synthase in the chloroplast and a cell wall less strain CC400 (cw15) were exposed to 70 μM Cd. At this Cd concentration, cell vitality was not affected, however, the strains showed various strategies to cope with Cd stress. In wt, most of Cd was diffused in the whole cell, and complexed by thiol ligands, while the other part was associated with phosphate in vacuolar Ca polyphosphate granules. Thiol ligands increased with exposure time, confirming their important role in Cd stress. In pcs1, Cd was also present as vacuolar Ca polyphosphate granules, and diffused in the cell as Cd-thiol complexes. In addition, while it should be regarded with caution, a minor proportion of Cd complexed by carboxyl groups, was potentially provided by starch produced around the pyrenoid and in the chloroplast. Results suggested that pcs1 uses thiol compounds such as PC to a lesser extent for Cd sequestration than wt. In cw15, an excretion of Cd, Ca polyphosphate granules has to be considered. Finally, Cd was detected in the pyrenoid of all strains