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While early hypotheses proposed that CAs enhance CO2 assimilation by rapidly accessing the HCO3\u2212 pool, direct evidence has been lacking. In this study, we examined Arabidopsis (Arabidopsis thaliana) mutants lacking both chloroplast-localized \u03b2CA1 (AT3G01500) and \u03b2CA5 (AT4G33580) to assess their impact on plant growth and photosynthetic performance. Our results show that plants deficient in chloroplast CA activity are unable to grow under ambient CO2 conditions (400\u2005\u03bcL\u2005L\u207b1) but can complete their life cycle under elevated CO2 levels (\u226512,000\u2005\u03bcL\u2005L\u207b1). However, CO2 assimilation rates and \u03a6II measurements in CA-deficient plants were comparable to those in wild-type plants under 0.04% (400\u2005\u03bcL\u2005L\u207b1), 0.4% (4,000\u2005\u03bcL\u2005L\u207b1), and 4% CO2 (40,000\u2005\u03bcL\u2005L\u207b1) concentrations, indicating that chloroplast CAs are not essential for photosynthetic CO2 fixation. Instead, our findings suggest that chloroplast CA activity is critical for supporting other metabolic pathways, namely amino acid, nucleic acid, and fatty acid biosynthesis. Expression of the Chlamydomonas (Chlamydomonas reinhardtii) bicarbonate transporter LCIA in chloroplast CA mutants partially rescued the growth phenotype under near-ambient CO2 conditions. These LCIA-complemented lines showed no difference in photosynthesis, further supporting the role of CAs in non-photosynthetic reactions. This work provides direct evidence that while chloroplast CAs are dispensable for photosynthesis, they are essential for plant growth and development under ambient CO2 due to their role in increasing the bicarbonate concentration for specific anaplerotic pathways.<\/jats:p>","DOI":"10.1093\/plphys\/kiag048","type":"journal-article","created":{"date-parts":[[2026,1,30]],"date-time":"2026-01-30T12:47:30Z","timestamp":1769777250000},"source":"Crossref","is-referenced-by-count":4,"title":["Plastid carbonic anhydrases are essential for growth of Arabidopsis on ambient air but not for photosynthesis"],"prefix":"10.1093","volume":"200","author":[{"ORCID":"https:\/\/orcid.org\/0009-0000-6129-0177","authenticated-orcid":false,"given":"Hiruni N","family":"Weerasooriya","sequence":"first","affiliation":[{"name":"Department of Biological Sciences, Louisiana State University , Baton Rouge, LA 70803 ,","place":["United States"]},{"name":"MSU-DOE Plant Research Laboratory , 612 Wilson Rd., East 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