Genetic and physiological analysis of the CO2-concentrating system of Chlamydomonas reinhardii.

When grown photoautotrophically at air levels of CO2, Chlamydomonas reinhardii possesses a system involving active transport of inorganic carbon which increases the intracellular CO2 concentration considerably above ambient, thereby stimulating photosynthetic CO2 assimilation. In previous investigations, two mutant strains of this unicellular green alga deficient in some component of this CO2-concentrating system were recovered as strains requiring high levels of CO2 to support photoautotrophic growth. One of the mutants, ca-1-12-1C, is a leaky (nonstringent) CO2-requiring strain deficient in carbonic anhydrase (EC 4.2.1.1) activity, while the other, pmp-1-16-5K, is a stringent CO2-requiring strain deficient in inorganic carbon transport. In the present study a double mutant (ca pmp) was constructed to investigate the physiological and biochemical interaction of the two mutations. The two mutations are unlinked and inherited in a Mendelian fashion. The double mutant was found to have a leaky CO2-requiring phenotype, indicating that the mutation ca-1 overcomes the stringent CO2-requirement conferred by the mutation pmp-1. Several physiological characteristics of the double mutant were very similar to the carbonic-anhydrase-deficient mutant, including high CO2 compensation concentration, photosynthetic CO2 response curve, and deficiency of carbonic-anhydrase activity. However, the labeling pattern of metabolites during photosynthesis in (14)CO2 was more like that of the bicarbonatetransport-deficient mutant, and accumulation of internal inorganic carbon was intermediate between that of the two original mutants. These data indicate a previously unsuspected complexity in the Chlamydomonas CO2-concentrating system.