Ccm1 , a regulatory gene controlling the induction of a carbon-concentrating mechanism in Chlamydomonas reinhardtii by sensing CO ₂ availability

Abstract

Aquatic photosynthetic organisms, including the green algaChlamydomonas reinhardtii, induce a set of genes for a carbonconcentrating mechanism (CCM) to acclimate to CO 2-limiting conditions. This acclimation is modulated by some mechanisms in the cell to sense CO2 availability. Previously, a high-CO2-requiring mutant C16 defective in an induction of the CCM was isolated from C. reinhardtii by gene tagging. By using this pleiotropic mutant, we isolated a nuclear regulatory gene, Ccm1, encoding a 699-aa hydrophilic protein with a putative zinc-finger motif in its Nterminal region and a Gln repeat characteristic of transcriptional activators. Introduction of Ccm1 into this mutant restored an active carbon transport through the CCM, development of a pyrenoid structure in the chloroplast, and induction of a set of CCM-related genes. That a 5,128-base Ccm1 transcript and also the translation product of 76 kDa were detected in both high-and low-CO2 conditions suggests that CCM1 might be modified posttranslationally. These data indicate that Ccm1 is essential to control the induction of CCM by sensing CO2 availability in Chlamydomonas cells. In addition, complementation assay and identification of the mutation site of another pleiotropic mutant, cia5, revealed that His-54 within the putative zinc-finger motif of the CCM1 is crucial to its regulatory function.zinc-finger motif ͉ carbon transport ͉ signal transduction ͉ photosynthesis ͉ acclimation P hotosynthetic organisms sense environmental changes, e.g., light, temperature, and various nutrient availabilities, to modulate and optimize photosynthetic activities. A number of aquatic photosynthetic organisms are able to concentrate dissolved inorganic carbon (DIC) intracellularly, allowing rapid growth despite low-CO 2 availability externally (1). This carbonconcentrating mechanism (CCM) shows acclimation to external DIC to optimize CO 2 fixation efficiency (2). During acclimation these organisms induce the expression of a set of genes required for various aspects of the CCM. In Chlamydomonas reinhardtii, several genes have been shown to be regulated in response to changes in external CO 2 concentration, including periplasmic carbonic anhydrase (CA; ref.3), mitochondrial CA (4), and a chloroplast envelope protein, LIP-36 (5). A development of pyrenoid structure in the chloroplasts also is modulated by the supply of CO 2 (6). This acclimation to CO 2 -limiting conditions suggests the existence of sensory mechanisms by which cells perceive the shortage of CO 2 and pathways by which the signal is transduced into specific gene regulation. However, the regulation of gene expression during this acclimation in eukaryotic organisms is still poorly understood. One reason is the paucity of mutants impaired in induction processes. We have isolated previously one C. reinhardtii mutant (C16) by gene-tagging mutagenesis, which was high-CO 2 requiring (7), and we showed that C16 exhibited a defect in CCM induction, with a low affinity for DIC and a low level of DIC accumulation under C...

Concerted Up-regulation of Aldehyde/Alcohol Dehydrogenase (ADHE) and Starch in Chlamydomonas reinhardtii Increases Survival under Dark Anoxia

“…In these conditions, the cells rely on CCM to concentrate CO 2 at the active site of the Rubisco (44). The ADHE abundance was first assessed in two CCM mutant strains: strain CIA5, which lacks Ccm1, a master gene regulator that controls the induction of CCM by sensing CO 2 availability in C. reinhardtii (45,46), and strain CIA3, which lacks CAH3, the thylakoid lumen carbonic anhydrase, which provides CO 2 to the Rubisco (47). The ADHE abundance in these two strains grown under atmospheric conditions was similar, being 1.5-1.8-fold higher than that in wild-type strain CC-124 (Table 4).…”

Lack of the Rhesus protein Rh1 impairs growth of the green alga Chlamydomonas reinhardtii at high CO 2

“…The growth defect of the RNAi lines provides further evidence that Rh1 is a gas channel for CO 2 . At high CO 2 , the CCM is not induced (36,(39)(40)(41)(42)(43)(44) and there is no need to prevent CO 2 regenerated near Rubisco from leaking away. (It is CO 2 , rather than HCO 3 Ϫ , which is the substrate for this enzyme, but probably HCO 3 Ϫ that is accumulated and transferred to the chloroplast by cells grown in air).…”

“…Chlamydomonas genes are highly expressed during growth in air, a circumstance under which the so-called CCM is activated (36,(39)(40)(41)(42)(43)(44). These include genes coding for a periplasmic carbonic anhydrase Cah1 (36,38), the mitochondrial carbonic anhydrase mtCA (32,45), and the chloroplast envelope protein LIP-36 (33,46).…”

Light and CO2/cAMP Signal Cross Talk on the Promoter Elements of Chloroplastic β-Carbonic Anhydrase Genes in the Marine Diatom Phaeodactylum tricornutum

“…The enhancer region is controlled by two critical elements denoted enhancer-element consensus (Kucho et al, 2003), to which a class of basic-ZIP transcription factors, enhancer-element-consensus binding proteins, interact (Yoshioka et al, 2004). A zinc finger protein, CCM1/Cia5, has been found to be involved in the up-regulation of most low-CO 2 -inducible genes, strongly suggesting that CCM1/Cia5 is a master regulator for the transcriptional response of the Chlamydomonas CCM, but the mechanism of the initial CO 2 signal perception is still unknown (Fukuzawa et al, 2001;Xiang et al, 2001). …”