Auxin transport in membrane vesicles from Cucurbita pepo L

Abstract

Association of (14)C-labelled indole-3-acetic acid (IAA) with membrane particles from zucchini (Cucurbita pepo L.) hypocotyls - previously described as "site III binding" (M. Jacobs and R. Hertel, 1978, Planta 142, 1-10) - is reinterpreted as a carrier-mediated uptake into closed and sealed vesicles driven by a pH gradient. Accumulation of the radioactive auxin is saturable, sensitive to the protonophore, carbonyl cyanide p-trifluoromethoxyphenyl hydrazone (FCCP), and to nigericin, and requires a pH gradient across the membranes with proton concentration greater outside than inside. The pH gradient decays within 1-2 h at 4°C and can be restored by re-equilibration of the particle preparation at more alkaline pH followed by return to more acidic medium. Osmotic shock and sonication release the IAA from the vesicles. 1-N-naphthylphthalamic acid (NPA) and 2,3,5-triiodobenzoic acid (TIBA), both inhibitors of auxin transport in vivo, increase the amount of net IAA accumulation in the vesicles, presumably by blocking efflux. Analogs of NPA less active or inactive in vivo are respectively less active or inactive in vitro. It is proposed that these membrane particles are outside-out plasma membrane vesicles, and that they perform the essential functions of auxin transport according to the chemiosmotic theory, with a specific, saturable proton symport uptake and an export anion carrier which is inhibited by NPA and TIBA.