The cortical array of microtubules inside the cell and arabinogalactan proteins on the external surface of the cell are each implicated in plant morphogenesis. To determine whether the cortical array is influenced by arabinogalactan proteins, we first treated Arabidopsis roots with a Yariv reagent that binds arabinogalactan proteins. Cortical microtubules were markedly disorganized by 1 microM beta-D-glucosyl (active) Yariv but not by up to 10 microM beta-D-mannosyl (inactive) Yariv. This was observed for 24-h treatments in wild-type roots, fixed and stained with anti-tubulin antibodies, as well as in living roots expressing a green fluorescent protein (GFP) reporter for microtubules. Using the reporter line, microtubule disorganization was evident within 10 min of treatment with 5 microM active Yariv and extensive by 30 min. Active Yariv (5 microM) disorganized cortical microtubules after gadolinium pre-treatment, suggesting that this effect is independent of calcium influx across the plasma membrane. Similar effects on cortical microtubules, over a similar time scale, were induced by two anti-arabinogalactan-protein antibodies (JIM13 and JIM14) but not by antibodies recognizing pectin or xyloglucan epitopes. Active Yariv, JIM13, and JIM14 caused arabinogalactan proteins to aggregate rapidly, as assessed either in fixed wild-type roots or in the living cells of a line expressing a plasma membrane-anchored arabinogalactan protein from tomato fused to GFP. Finally, electron microscopy of roots prepared by high-pressure freezing showed that treatment with 5 microM active Yariv for 2 h significantly increased the distance between cortical microtubules and the plasma membrane. These findings demonstrate that cell surface arabinogalactan proteins influence the organization of cortical microtubules.