Polar auxin transport in the intact pea (Pisum sativum L.)

The `chemiosmotic polar diffusion hypothesis' of the mechanism of polar auxin transport in plants satisfactorily accounts for the observed properties of transmembrane auxin transport in isolated tissues, cell suspensions and membrane vesicle preparations, but the extent to which it can adequately describe the characteristics of the long-distance, root-directed transport of auxin from the apex of the intact plant remains unclear. The principal objective of the work described in this thesis was to investigate the degree to which the properties of transmembrane auxin transport predicted by the hypothesis are consistent with the known features of the long distance transport of indol-3yl-acetic acid (IAA) applied to the apex of in the intact pea plant (Pisum sativum L.). The transport properties of apically-applied [1-14C]IAA in the stem of intact pea plants were identical to those associated with its polar transport in isolated shoot segments. [1-14C]IAA moved through the stem at velocities of 8-15 mm h-1 and this movement was strongly inhibited by local applications to the stem of non-competitive inhibitors of IAA transmembrane efflux carriers (N-1-naphthylphthalamic acid, NPA; and 2,3,5-triiodobenzoic acid, TIBA) and by high concentrations of competing unlabelled auxins (IAA; 1-naphthaleneacetic acid, 1-NAA; and 2,4-dichlorophenoxyacetic acid, 2,4-D). The net uptake of IAA by small segments of the same tissue was shown to involve all the transmembrane auxin transport components predicted by the chemiosmotic polar diffusion hypothesis. Segments excised from the stem of intact plants transporting apically-applied [1-14C]IAA effluxed 14C from their basal ends but not from their apical ends; basal efflux was significantly inhibited by inclusion of 3 μM NPA in the efflux medium. It is concluded that the rootward migration of IAA from the shoot tip of the intact plant is a true polar transport and occurs by the same mechanism as that postulated for transport in isolated stem segments. Tissue from correlatively-inhibited shoots and decapitated mainstems of pea lost the ability to support polar transport of [1-^14C]IAA, but transport was restored in both cases when an apical supply of auxin was reestablished. The loss of polar transport in these systems was shown not to result from a reduced rate of IAA uptake, loss or inactivation of specific IAA carrier systems, inability of the cells to maintain transmembrane pH gradients, or changed patterns of IAA metabolism. This reversible loss of polar transport may reflect a gradual randomization of auxin efflux carrier distribution in the plasma membrane following withdrawal of an auxin supply. The transport of phenylacetic acid (PAA) in intact plants and stem segments was also investigated. Long known to possess auxin-like growth regulatory activity, PAA has recently been shown to be a natural constituent of higher plants. Unlike IAA, apically applied [1-^14C]PAA did not undergo significant long-distance transport in the stem, despite ready accumulation by apical tissues and a similar pattern of metabolism in the shoot tip to that exhibited by IAA. Although readily taken up by leaves and root system of intact pea plants, PAA was not transported in the xylem or phloem. Local applications of PAA to the stem of intact plants inhibited the basipetal movement of apically-applied [1-^14C]IAA. Pea stem segments readily accumulated [1-^14C]PAA from buffered external solutions by diffusion of the undissociated acid, but PAA was found not to be a transported substrate for carriers mediating IAA transmembrane transport. Nevertheless, PAA interacted with these carriers to inhibit both the uptake and efflux of [1-^14C]IAA. It is proposed that endogenous PAA may function as a regulator of the polar transport and/or accumulation of IAA.