Recent evidence indicates a correlation between orientation of the plant cortical microtubule cytoskeleton and localization of polar cargoes1. However, the molecules and mechanisms that create this correlation have remained unknown. Here we show that, inArabidopsis thaliana, the microtubule orientation regulators CLASP2,3and MAP65 (refs3,4) control the abundance of polarity regulator PINOID kinase5,6at the plasma membrane. By localized upregulation of clathrin-dependent endocytosis7at cortical microtubule- and clathrin-rich domains orthogonal to the axis of polarity, PINOID accelerates the removal of auxin transporter PIN proteins8,9,10from those sites. This mechanism links directional microtubule organization to the polar localization of auxin transporter PIN proteins, and clarifies how microtubule-enriched cell sides are kept distinct from polar delivery domains. Our results identify the molecular machinery that connects microtubule organization to the regulation of the axis of PIN polarization.