cis-Regulatory elements for mesophyll-specific gene expression in the C4 plant Flaveria trinervia, the promoter of the C4 phosphoenolpyruvate carboxylase gene

C-4 photosynthesis depends on the strict compartmentalization of CO2 assimilatory, enzymes. cis-regulatory mechanisms are described that ensure mesophyll-specific expression of the gene encoding the C-4 isoform of phosphoenolpyruvate carboxylase (ppcA1) of the C-4 dicot Flaveria trinervia. To elucidate and understand the anatomy of the C-4 ppcA1 promoter, detailed promoter/reporter gene studies were performed in the closely related C-4 species F. bidentis, revealing that the C-4 promoter contains two regions, a proximal segment up to -570 and a distal part from -1566 to -2141, which are necessary but also sufficient for high mesophyll-specific expression of the beta-glucuronidase reporter gene. The distal region behaves as an enhancer-like expression module that can direct mesophyll-specific expression when inserted into the ppcA1 promoter of the C-3 plant F. pringlei. Mesophyll expression determinants were restricted to a 41-by segment, referred to as mesophyll expression module 1 (Mem1). Evolutionary and functional studies identified the tetranucleotide sequence CACT as a key component of Mem1.