Dynamic changes of genome sizes and gradual gain of cell-specific distribution of C4 enzymes during C4 evolution in genus Flaveria

Abstract

C₄ plants are believed to have evolved from C₃ plants through various C₃ -C₄ intermediate stages in which a photorespiration-dependent CO₂ concentration system known as C₂ photosynthesis operates. Genes involved in the C₄ cycle were thought to be recruited from orthologs present in C₃ species and developed cell-specific expression during C₄ evolution. To understand the process of establishing C₄ photosynthesis, we performed whole-genome sequencing and investigated expression and mesophyll- or bundle-sheath-cell-specific localization of phosphoenolpyruvate carboxylase (PEPC), NADP-malic enzyme (NADP-ME), pyruvate, orthophosphate dikinase (PPDK) in C₃ , C₃ -C₄ intermediate, C₄ -like, and C₄ Flaveria species. While genome sizes vary greatly, the number of predicted protein-coding genes was similar among C₃ , C₃ -C₄ intermediate, C₄ -like, and C₄ Flaveria species. Cell-specific localization of the PEPC, NADP-ME, and PPDK transcripts was insignificant or weak in C₃ -C₄ intermediate species, whereas these transcripts were expressed cell-type specific in C₄ -like species. These results showed that elevation of gene expression and cell-specific control of pre-existing C₄ cycle genes in C₃ species was involved in C₄ evolution. Gene expression was gradually enhanced during C₄ evolution, whereas cell-specific control was gained independently of quantitative transcriptional activation during evolution from C₃ -C₄ intermediate to C₄ photosynthesis in genus Flaveria.