Early Initiation of Bundle Sheath Cells During Leaf Development as Visualised by SCARECROW Expression in Dicotyledonous C Plants.

The C type of dicotyledonous plants exhibit a higher density of reticulate veins than the C type, with a nearly 1:1 ratio of mesophyll cells (MCs) to bundle sheath cells (BSCs). To understand how this C-type cell pattern is formed, we identified two SCARECROW (SCR) genes in C Flaveria bidentis, FbSCR1 and FbSCR2, that fully or partially complement the endodermal cell layer-defective phenotype of Arabidopsis scr mutant. We then created FbSCRs promoter β-glucuronidase reporter (GUS) lines of F. bidentis, which showed GUS expression in BSCs and their progenitor cells. The GUS expression pattern in F. bidentis transformants and comparison with the closely related C-type Flaveria pringlei revealed that higher-order veins were initiated in the early leaf developmental stage. Treatment with an auxin polarity transport inhibitor decreased the MC area and led to vein formation without free ends, resulting in the formation of BSCs in positions adjacent to other BSCs. However, BSC differentiation was not affected, as evidenced by BSC specific FbSCR1 expression and RuBisCO accumulation. These results indicate that polar auxin transport is important for MC proliferation and/or differentiation, which leads to the formation of a C-type cell pattern in which MCs and BSCs are equally adjacent.