Timing of gene expression and recruitment in independent origins of CAM in the Agavoideae (Asparagaceae).

CAM photosynthesis has evolved repeatedly across the plant tree of life, yet our understanding of the genetic convergence across independent origins remains hampered by the lack of comparative studies. CAM is furthermore thought to be closely linked to the circadian clock in order to achieve temporal separation of carboxylation and sugar production. Here, we explore gene expression profiles in eight species from the Agavoideae (Asparagaceae) encompassing three independent origins of CAM. Using comparative physiology and transcriptomics, we examined the variable modes of CAM in this subfamily and the changes in gene expression across time of day and between well-watered and drought-stressed treatments. We further assessed gene expression and molecular evolution of genes encoding phosphoenolpyruvate carboxylase (PPC), an enzyme required for primary carbon fixation in CAM. Most time-of-day expression profiles are largely conserved across all eight species and suggest that large perturbations to the central clock are not required for CAM evolution. In contrast, transcriptional response to drought is highly lineage specific. Yucca and Beschorneria have CAM-like expression of PPC2, a copy of PPC that has never been shown to be recruited for CAM in angiosperms, and evidence of positive selection in PPC genes implicates mutations that may have facilitated the recruitment for CAM function early in the evolutionary history of the Agavoideae. Together the physiological and transcriptomic comparison of closely related C3 and CAM species reveals similar gene expression profiles, with the notable exception of differential recruitment of carboxylase enzymes for CAM function.