Pyruvate orthophosphate dikinase is required for the acclimation to high bicarbonate concentrations in Phaeodactylum tricornutum

Abstract

The model marine diatom P. tricornutum possesses efficient CO2-concentrating mechanisms (CCMs) involving the bicarbonate transport and the rapid inter-conversion of HCO3− and CO2, and it has a complete set of enzymes essential for biochemical CCM C4 pathway. However, the role of potential C4 pathway in the diatom is still controversial. Based on the analysis of previously published proteome and transcriptome data in P. tricornutum, the expression of pyruvate orthophosphate dikinase (PPDK), a key enzyme generating the primary acceptor for bicarbonate fixation in C4 pathway, was upregulated in long-term high CO2-selected populations. In our study, P. tricornutum PPDK (PtPPDK) transcript abundance was higher with short-term treatment of high NaHCO3 concentrations and overexpression of PtPPDK resulted in a cell density increase of 5–16 % from day 2 to 8. Knockdown (KD) or knockout of PtPPDK inhibited the growth and the maximal photosystem II electron transport rate, and the inhibited growth became more and more evident with the increasing NaHCO3 concentration from 1 to 8 mM in PtPPDK KD lines. Furthermore, elevating NaHCO3 from 1 to 16 mM dramatically decreased the cell density both in wild-type and PtPPDK mutants, suggesting that 1 mM NaHCO3 is enough for the photosynthesis carbon assimilation. PtPPDK was localized in the plastid stroma by eGFP fusion protein analysis at 2 mM NaHCO3, while PtPPDK-eGFP fluorescence was observed both in the stroma and periplasmic space at higher NaHCO3. It is suggested that PPDK is necessary for consuming excessive inorganic carbon (Ci) both from the plastid and extracellular environment to maintain pH homeostasis, indicating a potential function of adapting to high Ci concentrations.