Change in expression levels of NAD kinase-encoding genes in Flaveria species

Nicotinamide adenine dinucleotides (NAD(H)) and NAD phosphates (NADP(H)) are electron carriers involved in redox reactions and metabolic processes in all organisms. NAD kinase (NADK) is the only enzyme that phosphorylates NAD⁺ into NADP⁺, using ATP as a phosphate donor. In NADP-dependent malic enzyme (NADP-ME)-type C₄ photosynthesis, NADP(H) are required for dehydrogenation by NADP-dependent malate dehydrogenase (NADP-MDH) in mesophyll cells, and decarboxylation by NADP-ME in bundle sheath cells. In this study, we identified five NADK genes (FbNADK1a, 1b, 2a, 2b, and 3) from the C₄ model species Flaveria bidentis. RNA-Seq database analysis revealed higher transcript abundance in one of the chloroplast-type NADK2 genes of C₄F. bidentis (FbNADK2a). Comparative analysis of NADK activity in leaves of C₃, C₃-C₄, and C₄Flaveria showed that C₄Flaveria (F. bidentis and F. trinervia) had higher NADK activity than the other photosynthetic-types of Flaveria. Taken together, our results suggest that chloroplastic NAD kinase appeared to increase in importance as C₃ plants evolved into C₄ plants in the genus Flaveria.