Background
Algae that are living in aquatic environments experience restraints in photosynthetic carbon assimilation because gases are diffusing much slower in water than in air. A slow diffusion of inorganic carbon may eventually lead to a deficiency of CO₂ inside the cells where the photosynthetic reduction of CO₂ occurs. Photosynthetic organisms like algae are fixing the CO₂ mainly by using the enzyme Ribulose-1-5-bisphosphate Carboxylase/Oxygenas (rubisco). This slow and inefficient enzyme is also catalyzing the fixation of oxygen (oxygenase reaction) in a reaction competing with CO₂ assimilation. The efficiency of photosynthesis is therefore ultimately linked to the ratio between CO₂ and O₂ at the active site of rubisco. It is therefore not surprising that organisms experiencing inorganic carbon limitation have evolved mechanisms to increase the concentration of CO₂ in the chloroplast. Most cyanobacteria and microalgae seem to have developed a unique mechanism to concentrate inorganic carbon inside the cells. This mechanism, generally called the Carbon Concentrating Mechanism (CCM) is operating via an energy dependent transport of inorganic carbon species (CO₂ and/or HCO_3-) and an efficient formation of CO₂ at the active site of rubisco from the HCO_3- pool created inside the chloroplasts. CCM is an inducible mechanism and cells transferred from high CO₂ (1-5% CO₂ in air) to ambient air concentrations of CO₂ induce the mechanism within a few hours. Cells with a fully induced CCM have a much higher affinity for inorganic carbon than non-induced cells and hence their photosynthesis is very efficient even under very low inorganic carbon conditions. When C. reinhardtii is transferred to air from high CO₂-conditions, a set of genes are immediately activated and at least 8-10 new gene products can be identified using SDS PAGE. It is believed that these gene products are essential for the cell's ability to concentrate carbon.

Objectives
In my group we are mainly working with the green alga Chlamydomonas reinhardtii as a molecular model system to study CCM. During many years we have been focusing on the role of Carbonic Anhydrases (CA) in CCM. We have been able to identify two novel CAs in C. reinhardtii, one in the thylakoid lumen and one in the mitochondia. They belong to two different gene families, alpha-CA (lumen) ß-CA (mitochondria). The mitochondrial CA is strongly induced by a lowering of the CO₂ concentration in the medium while the lumen CA is expressed in a constitutive manner. One of the key issues of my research now is to resolve the signal transduction pathway between the lowering of CO₂ in the medium to the gene expression of CCM related genes. The ultimate goal is to identify genes and gene products involved in sensing and transducing the information about the concentration of inorganic carbon in the medium to the nucleus. We have taken the advantage of the fact that the mitochondrial CA gene is inducible and so we have fused a reporter gene behind the mitochondrial CA promoter and transferred it back into the cells of C. reinhardtii. The mutant formed, CAP1, is further the target for random mutagenesis and transformed cells are screened for defects in CO₂ sensing and transduction. This is an efficient system and we can with relative ease screen millions of cells. So far one mutant has been obtained which a decreased and delayed induction. Preliminary results indicate that the mutated gene is coding for an enhancer protein. In another project the higher plant Arabidopsis thaliana is used to identify novel CAs. Recently we have been able to show that a novel alpha-CA is expressed in the chloroplast stroma. The long-term goal is to resolve its role for carbon flow in the cells of this higher plant model system.

Selected publications:

Eriksson, M., Ramazanov, Z., Gardeström, P. and Samuelsson, G. () Discovery of an algal mitochondrial carbonic anhydrase: Molecular cloning and characterization of a low-CO₂- induced polypeptide in Chlamydomonas reinhardtii. PNAS 93:

Villand, P., Eriksson, M. and Samuelsson, G. () Carbon dioxide and light regulation of promoters controlling expression of mitochondrial carbonic anhydrase in Chlamydomonas reinhardtii are regulated by carbon dioxide and light but not by oxygen. Biochemical Journal 327:51-57

Hiltonen, T., Björkbacka, H., Forsman, C., Clarke, A. and Samuelsson, G. () Intracellular beta-carbonic anhydrase in the unicellularbgreen alga Coccomyxa. Plant Physiology 117:

Karlsson, J., Clarke, A., Chen, Z., Hugghins, S., Park, Y.-I., Husic, D., Moroney, J.V. and Samuelsson, G. () A noveltype carbonic anhydrase associated with the thylakoid membrane in Chlamydomonas reinhardtii is required for growth at ambient CO₂. EMBO Journal 17 (5)

Park, Y.-I., Karlsson, J., Rojdestvenski, I., Pronina, N., Klimov, V., Öquist, G. and Samuelsson, G. () The role of the novel photosystem II-associated carbonic anhydrase in carbon concentrating mechanism (CCM) in Chlamydomonas reinhardtii. FEBS letters 444:102-105