Abstract
The chloroplast protein CP12 is involved in the dark/light regulation of the Calvin–Benson–Bassham cycle, in particular, in the dark inhibition of two enzymes: glyceraldehyde−3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK), but other functions related to stress have been proposed. We knocked out the unique CP12 gene to prevent its expression in Chlamydomonas reinhardtii (ΔCP12). The growth rates of both wild-type and ΔCP12 cells were nearly identical, as was the GAPDH protein abundance and activity in both cell lines. On the contrary, the abundance of PRK and its specific activity were significantly reduced in ΔCP12, as revealed by relative quantitative proteomics. Isolated PRK lost irreversibly its activity over-time in vitro, which was prevented in the presence of recombinant CP12 in a redox-independent manner. We have identified amino acid residues in the CP12 protein that are required for this new function preserving PRK activity. Numerous proteins involved in redox homeostasis and stress responses were more abundant and the expressions of various metabolic pathways were also increased or decreased in the absence of CP12. These results highlight CP12 as a moonlighting protein with additional functions beyond its well-known regulatory role in carbon metabolism.
Highlights
In all photosynthetic eukaryotes, the Calvin–Benson–Bassham (CBB) cycle involved in CO2 assimilation is dependent upon reducing power and ATP production from the photochemical phase of photosynthesis
The Absence of CP12 Protein Has No Effect on C. reinhardtii Growth
Once the targeted mutation of the CP12 gene was confirmed, we tested the growth of wild-type (WT) CC4349 C. reinhardtii and ∆CP12-Cr strains
Summary
The Calvin–Benson–Bassham (CBB) cycle involved in CO2 assimilation is dependent upon reducing power and ATP production from the photochemical phase of photosynthesis. CP12 is reduced, and its disulfide bridges are disrupted The reduction in these disulfide bridges triggers conformational changes and leads to a fully disordered CP12, while, under its oxidized state, CP12 oscillates between a disordered and a partly ordered state [10,11]. This redox-dependent conditional disorder, described for other proteins such as Cox and Hsp33 [12], allows the regulation of the CBB cycle and, in particular, of PRK and GAPDH activities. Upon CP12 reduction, the two enzymes are released from the supramolecular complex and become fully active, allowing the CBB cycle to function
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