Overexpression of miR165 affects apical meristem formation, organ polarity establishment and vascular development in Arabidopsis.

The class III homeodomain leucine-zipper (HD-ZIP III) genes are thought to be targets of microRNAs (miRNAs) 165 and 166, but it is not known whether all the developmental processes affected by mutations of the HD-ZIP III genes could be recapitulated by an alteration in the expression of miR165 and miR166. Previous work showed that overexpression of miR166 by activation tagging results in down-regulation of the ATHB-9/PHV, ATHB-14/PHB and ATHB-15 genes, and concomitantly causes an enlargement of shoot apical meristems (SAMs) and an enhancement in vascular development. Here we demonstrated that overexpression of miR165 causes a drastic reduction in the transcript levels of all five HD-ZIP III genes in Arabidopsis. The miR165 overexpressors display prominent phenotypes reminiscent of loss-of-function mutants of rev phb phv and rev/ifl1, including loss of SAM, alteration of organ polarity, abnormal formation of carpels, inhibition of vascular development and aberrant differentiation of interfascicular fibers. Global gene expression analysis revealed a link between miR165 overexpression and altered expression of genes involved in auxin signaling and vascular development. Our results demonstrate that overexpression of miR165 recapitulates the phenotypes caused by loss-of-function mutations of HD-ZIP III genes, such as loss of SAM, altered organ polarity and defects in development of vascular tissues and interfascicular fibers.