Endoreduplication is not involved in bundle-sheath formation in the C4 species Cleome gynandra.

There is currently significant interest in engineering the two-celled C-4 photosynthesis pathway into crops such as rice in order to increase yield. This will require alterations to the biochemistry of photosynthesis in both mesophyll (M) and bundle-sheath (BS) cells, but also alterations to leaf anatomy. For example, the BS of C-4 species is enlarged compared with that in C-3 species. Because cell and nucleus size are often correlated, this study investigated whether nuclear endoreduplication is associated with increased differentiation and expansion of BS cells. Nuclei in the BS of C-4 Cleome gynandra were tagged with green fluorescent protein. Confocal laser-scanning microscopy and flow cytometry of isolated nuclei were used to quantify size and DNA content in BS cells. The results showed a significant endoreduplication in BS cells of C. gynandra but not in additional C-4 lineages from both the monocotyledonous and dicotyledenous plants. Furthermore, in the C-3 species Arabidopsis thaliana, BS cells undergo endoreduplication. Due to this significant endoreduplication in the small BS cells of C-3 A. thaliana, it was concluded that endoreduplication of BS nuclei in C-4 plants is not linked to expansion and differentiation of BS cells, and therefore that alternative strategies to increase this compartment need to be sought in order to engineer C-4 traits into C-3 crops such as rice.