Identification and functional role of the carbonic anhydrase Cah3 in thylakoid membranes of pyrenoid of Chlamydomonas reinhardtii

Abstract

The distribution of the luminal carbonic anhydrase Cah3 associated with thylakoid membranes in the chloroplast and pyrenoid was studied in wild-type cells of Chlamydomonas reinhardtii and in its cia3 mutant deficient in the activity of the Cah3 protein. In addition, the effect of CO(2) concentration on fatty acid composition of photosynthetic membranes was examined in wild-type cells and in the cia3 mutant. In the cia3 mutant, the rate of growth was lower as compared to wild-type, especially in the cells grown at 0.03% CO(2). This might indicate a participation of thylakoid Cah3 in the CO(2)-concentrating mechanism (CCM) of chloroplast and reflect the dysfunction of the CCM in the cia3 mutant. In both strains, a decrease in the CO(2) concentration from 2% to 0.03% caused an increase in the content of polyunsaturated fatty acids in membrane lipids. At the same time, in the cia3 mutant, the increase in the majority of polyunsaturated fatty acids was less pronounced as compared to wild-type cells, whereas the amount of 16:4ω3 did not increase at all. Immunoelectron microscopy demonstrated that luminal Cah3 is mostly located in the thylakoid membranes that pass through the pyrenoid. In the cells of CCM-mutant, cia3, the Cah3 protein was much less abundant, and it was evenly distributed throughout the pyrenoid matrix. The results support our hypothesis that CO(2) might be generated from HCO(3)(-) by Cah3 in the thylakoid lumen with the following CO(2) diffusion into the pyrenoid, where the CO(2) fixing Rubisco is located. This ensures the maintenance of active photosynthesis under CO(2)-limiting conditions, and, as a result, the active growth of cells. The relationships between the induction of CCM and restructuring of the photosynthetic membranes, as well as the involvement of the Cah3 of the pyrenoid in these events, are discussed. This article is part of a Special Issue entitled: Photosynthesis Research for Sustainability: from Natural to Artificial.

Zinc Deficiency Impacts CO2 Assimilation and Disrupts Copper Homeostasis in Chlamydomonas reinhardtii

“…It is possible that the proteins are indeed synthesized, but they might be degraded in the absence of the essential zinc cofactor, or they might be actively degraded as part of a zinc-sparing program (24,48). Carbonic anhydrases are important for photosynthesis under low CO 2 conditions based on the effect of enzyme inhibitors (acetazolamide and DBS) and based on the phenotype of a mutant lacking lumenal carbonic anhydrase Cah3 (66), and the expression of several CAH genes is increased in cells grown at air levels of CO 2 as part of the CCM (59). The conditional CO 2 -repressed phenotype of zinc-deficient Chlamydomonas cells establishes the carbonic anhydrases as key targets of zinc deficiency and therefore, in an environmental context, emphasizes the importance of the ability of diatoms to substitute cobalt or cadmium in the active site of CAs (22,67) to maintain photosynthetic growth in face of zinc deficiency.…”

Functions, Compositions, and Evolution of the Two Types of Carboxysomes: Polyhedral Microcompartments That Facilitate CO 2 Fixation in Cyanobacteria and Some Proteobacteria

“…It is known that some pyrenoids contain pyrenoid-localized CA enzymes (210), whereas others contain thylakoid invaginations/ strands which may contain thylakoid-localized CA enzymes (211). Indeed, Markelova et al (212) showed that the CAH3 CA enzyme was localized to both the thylakoid lumen and the pyrenoid body in Chlamydomonas reinhardtii, with this CA enzyme supporting CO 2 fixation only when correctly localized to the pyrenoid-traversing thylakoids (213).…”

Carbon Supply and Photoacclimation Cross Talk in the Green Alga Chlamydomonas reinhardtii

“…The essence of these processes lies in the active pumping of inorganic carbon in the cell via a number of transporters that concentrate it in the pyrenoid, a ball-like structure containing Rubisco, Rubisco activase, and intrapyrenoid thylakoids and surrounded by a starch sheath. In the pyrenoid, HCO 3 2 is converted to CO 2 by CARBONIC ANHYDRASE3 (CAH3; Blanco-Rivero et al, 2012;Sinetova et al, 2012) and then fixed by Rubisco in the Calvin-Benson-Bassham cycle. CAH3 also is suggested to provide HCO 3 2 in the proximity of the oxygen-evolving complex, where it may function as a proton carrier, removing H + from water splitting to avoid photoinhibition (Villarejo et al, 2002;Shutova et al, 2008).…”