Increased chloroplast area in the rice bundle sheath through cell-specific perturbation of brassinosteroid signaling

In the leaves of C3 species such as rice (Oryza sativa), mesophyll cells contain the largest compartment of photosynthetically active chloroplasts. In contrast, plants that use the derived and more efficient C4 photosynthetic pathway have a considerable chloroplast compartment in both bundle sheath and mesophyll cells. Accordingly, the evolution of C4 photosynthesis from the ancestral C3 state required an increased chloroplast compartment in the bundle sheath. Here, we investigated the potential to increase chloroplast compartment size in rice bundle sheath cells by manipulating brassinosteroid signaling. Treatment with brassinazole, a brassinosteroid biosynthesis inhibitor, raised leaf chlorophyll content and increased the number but decreased the area of chloroplasts in bundle sheath cells. Ubiquitous overexpression of the transcription factor-encoding BRASSINAZOLE RESISTANT 1 (OsBZR1) increased bundle sheath chloroplast area by up to 45%, but these plants became chlorotic. However, when OsBZR1 expression was driven by a bundle sheath-specific promoter, the negative effects on growth and viability were alleviated while chloroplast area still increased. In summary, we report a role for brassinosteroids in controlling chloroplast area and number in rice and conclude that cell-specific manipulation of brassinosteroid signaling can be used to manipulate the chloroplast compartment in rice bundle sheath cells.