Endoreduplication is not involved in bundle-sheath formation in the C(4) 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.