Identification and characterization of a frequent genetic alteration toward the evolution of C2-photosynthesis in the genus Moricandia

During the process of photosynthesis plants fix atmospheric CO2 into organic carbohydrates. The photosynthetic assimilation rates resulting from this reaction differ among different photosynthetic variants, in which C3 photosynthesis represents the most simple and ancestral photosynthetic variant, whereas C4 is the more advanced variant and more correlated with increased biomass. C4 photosynthesis evolved more than 60 times independently via intermediate C3-C4 variants that are referred to as C2 photosynthesis and are considered as a bridge between C3 and C4 photosynthesis. Understanding of the molecular basis of the evolution of C4 holds the potential of improving crop productivity. This thesis is mainly concerned with one of the earliest steps of development of the C4-trait, the restriction of glycine decarboxylase (GDC) complex to the bundle sheath (BS) of C2 plants. By using promoter deletion experiments this work shows that restriction of the GDC to the BS of C3 plants is possible. The cis-element responsible for this spatial expression was isolated from the promoter of AtGldp1, one of the two genes encoding the GDC-P subunit in Arabidopsis thaliana. The identified element is referred to as the M-box. Using bioinformatic analysis this work generalizes the presence of the M-box in the promoters of most C3 Gldp1 genes from Brassicaceae and suggests its loss from promoters of most C2 Gldp genes. Subsequently, this offers a possible molecular mechanism for restriction of GDC to BS in at least in genus Moricandia. The bioinformatic analysis was further validated experimentally by isolating additional C3 and C2 promoters from other species of the Moricandia that were not characterized or isolated before by restriction genome walking. We show that the M-box is found in the promoter of a C3 Moricandia species and absent in three Gldp promoters for three different Moricandia C2 species. The isolated promoters were further studied in Arabidopsis. Furthermore the M-box binding factor was defined to be a member of GATA transcription factor family via a yeast-one-hybrid screening coupled with several coexpression analyses and the processing of previous genome-wide footprint sequencing data. Collective results from this study identify and characterize a simple mechanism for establishing C2-specific gene expression in a C3 plant like Arabidopsis