Expression activation and functional analysis of HLA 3, a putative inorganic carbon transporter in C hlamydomonas reinhardtii
Summary The CO 2 concentrating mechanism (CCM) is a key component of the carbon assimilation strategy of aquatic microalgae. Induced by limiting CO 2 and tightly regulated, the CCM enables these microalgae to respond rapidly to varying environmental CO 2 supplies and to perform photosynthetic CO 2 assimilation in a cost‐effective way. A functional CCM in eukaryotic algae requires Rubisco sequestration, rapid interconversion between CO 2 and catalyzed by carbonic anhydrases (CAs), and active inorganic carbon (Ci) uptake. In the model microalga Chlamydomonas reinhardtii , a membrane protein HLA3 is proposed to be involved in active Ci uptake across the plasma membrane. In this study, we use an artificially designed transcription activator‐like effector ( dTALE ) to activate the expression of HLA3. The successful activation of HLA3 expression demonstrates dTALE as a promising tool for gene‐specific activation and investigation of gene function in Chlamydomonas . Activation of HLA3 expression in high CO 2 acclimated cells, where HLA3 is not expressed, resulted in increased Ci accumulation and Ci‐dependent photosynthetic O 2 evolution specifically in very low CO 2 concentrations, which confirms that HLA3 is indeed involved in Ci uptake, and suggests it is mainly associated with transport in very low CO 2 concentrations, conditions in which active CO 2 uptake is highly limited.