Ectopic assembly of an auxin efflux control machinery shifts developmental trajectories

Polar auxin transport in the Arabidopsis root tip maintains high auxin levels around the stem cell niche that gradually decrease in dividing cells but increase again once they transition towards differentiation. Protophloem differentiates earlier than other proximal tissues and employs a unique auxin ‘canalization’ machinery that is thought to balance auxin efflux with retention. It consists of a proposed activator of PIN auxin efflux carriers, the AGC kinase PAX; its inhibitor, BRX; and PIP5K enzymes, which promote polar PAX and BRX localization. Because of dynamic PAX-BRX-PIP5K interplay, the net cellular output of this machinery remains unclear. Here we deciphered the dosage-sensitive regulatory interactions between PAX, BRX and PIP5K by their ectopic expression in developing xylem vessels. The data suggest that the dominant collective output of the PAX-BRX-PIP5K module is a localized reduction in PIN abundance. This requires PAX-stimulated clathrin-mediated PIN endocytosis by site-specific phosphorylation, which distinguishes PAX from other AGC kinases. Importantly, ectopic assembly of the PAX-BRX-PIP5K module is sufficient to cause cellular auxin retention and affects root growth vigor by accelerating the trajectory of xylem vessel development. Our data thus provide direct evidence that local manipulation of auxin efflux alters the timing of cellular differentiation in the root.