Carbon allocation and metal use in four Chlamydomonas mutants defective in CCM-related genes

Four mutants of Chlamydomonas reinhardtii with defects in different components of the CO2 concentrating mechanism (CCM) or in rubisco activase were grown autotrophically at high pCO2 and then transferred to low pCO2, in order to study the role of different components of the CCM on carbon allocation and elemental composition. To study carbon allocation, we measured the relative size of the main organic pools by Fourier Transform Infrared spectroscopy. Total reflection X-ray fluorescence was used to analyze the elemental composition of algal cells. Our data show that although the organic pools increased their size at high CO2 in all strains, their stoichiometry was highly homeostatic, i.e. the ratios of carbohydrates to proteins, lipid and proteins and carbohydrates and lipids, did not change significantly. The only exception was the wild type 137c, in which proteins decreased relative to carbohydrates and lipids, when the cells were transferred to low CO2. It is noticeable, therefore, that the two wild types responded differently to the transition from high to low CO2. Malfunctions of the CCM influenced the concentration of several elements, somewhat altering cell elemental stoichiometry: especially the C/P and N/P ratios changed appreciably in almost all strains as a function of growth CO2, except in 137c and the rubisco activase mutant rca1. In strain cia3, defective in the luminal carbonic anhydrase (CA), the cell quotas of P, S, Ca, Mn, Fe and Zn were about 5-fold higher at low CO2 than at high CO2. A Principle Components Analysis showed that, mostly because of its elemental composition, cia3 behaved in a substantially different way from all other strains, at low CO2. The luminal CA thus plays a crucial role, not only for the correct functioning of the CCM, but also for element utilization. Not surprisingly, growth at high CO2 attenuated differences among strains.