Abstract
AbstractCarbonic anhydrase (CA), an enzyme conserved across species, is pivotal in the interconversion of inorganic carbon (Ci; CO2and HCO3−). Compared to the well-studied intracellular CA, the specific role of extracellular CA in photosynthetic organisms is still not well understood. In the green algaChlamydomonas reinhardtii, CAH1, located at the periplasmic space, is strongly induced under CO2-limiting conditions by the Myb transcription factor LCR1. While it has been observed that thelcr1mutant shows decreased Ci-affinity, the detailed mechanisms behind this phenomenon are yet to be elucidated. In this study, we aimed to unravel the LCR1-dependent genes essential for maintaining high Ci-affinity. To achieve this, we identified a total of 12 LCR1-dependent inducible genes under CO2-limiting conditions, focusing specifically on the most prominent ones -CAH1,LCI1,LCI6, andCre10.g426800. We then created mutants of these genes using the CRISPR-Cas9 system, all from the same parental strain, and compared their Ci-affinity. Contrary to earlier findings (Van and Spalding, 1999) that reported no reduction in Ci-affinity in thecah1mutant, our newly createdcah1-1 mutant exhibited a significant decrease in Ci-affinity under high HCO3−/CO2-ratio conditions. Additionally, when we treated wild-type cells with a CA inhibitor with low membrane permeability, a similar reduction in Ci-affinity was observed. Moreover, the addition of exogenous CA to thecah1mutant restored the decreased Ci-affinity. These results, highlighting the crucial function of the periplasmic CAH1 in maintaining high Ci-affinity inChlamydomonascells, provide new insights into the functions of periplasmic CA in algal carbon assimilation.One-sentence summaryCAH1, a periplasmic carbonic anhydrase inChlamydomonas reinhardtii, plays a crucial role in maintaining a high affinity for inorganic carbon, particularly under CO2-limiting conditions.
Publisher
Cold Spring Harbor Laboratory