Dose-dependent long-distance movement of microRNA399 duplex regulates phosphate homeostasis in Arabidopsis

Abstract

MicroRNA399 (miR399), a phosphate (Pi) starvation-induced long-distance signal, is first produced in shoots and moves to roots to suppress PHO2 encoding a ubiquitin conjugase, leading to enhanced Pi uptake and root-to-shoot translocation. However, the molecular mechanism underlying miR399 long-distance movement remains elusive. Hypocotyl grafting with various Arabidopsis mutants or transgenic lines expressing artificial miR399f was employed. The movement of miR399 across graft junction and the rootstock PHO2 transcript and scion Pi levels were analyzed to elucidate the potential factors involved. Our results showed that miR399f precursors are cell-autonomous and mature miR399f movement is independent of its biogenesis, sequence context, and length (21 or 22 nucleotides). Expressing viral silencing suppressor P19 in the root stele or blocking unloading in the root phloem pore pericycle (PPP) antagonized its silencing effect, suggesting that the miR399f/miR399f* duplex is a mobile entity unloaded through PPP. Notably, the scion miR399f level positively correlates with its amount translocated to rootstocks, implying dose-dependent movement. This study uncovers the molecular basis underlying the miR399-mediated long-distance silencing in coordinating shoot Pi demand with Pi acquisition and translocation activities in the roots.

Keywords: PHO2; long-distance movement; microRNA399; phosphate transport; small RNA duplex.