Use of mutants and transformed plants to study the action of auxins

We describe here the results of a comparison of the properties of several plant genotypes differing in their reaction to auxins. The hormonal response used to compare the genotypes is the auxin-induced variation of the transplasmalemma electrical potential difference (delta Em) exhibited by protoplasts isolated from leaves or root tips. Using this membrane response, we have shown that large variations in the sensitivity to auxins can be induced either by mutagenesis in tobacco or by transformation of various materials by Agrobacterium rhizogenes. Tobacco protoplasts isolated from an auxin resistant mutant selected by M. Caboche (INRA, Versailles), when compared to protoplasts from the control genotype display an auxin-induced hyperpolarization with a 10 fold decrease in their sensitivity to the hormone. Conversely, protoplasts isolated from root tips of various plants transformed by A. rhizogenes or from the leaves of tobacco plants regenerated from transformed roots, exhibit a 100 to 1000 fold increase in their sensitivity to auxins. The single gene, rol B, from the Ri plasmid harboured by A. rhizogenes is able to induce the dramatic modification in the sensitivity to auxins of tobacco protoplasts. The auxin-induced hyperpolarization of tobacco protoplasts is inhibited by antibodies directed against an auxin-binding protein (ABP) purified from corn coleoptile membranes. Immunotitrations of ABP antigens at the external surface of tobacco protoplasts show that transformed protoplasts could have more surface receptors than protoplasts from the normal genotype, whereas protoplasts isolated from the auxin-tolerant mutant could have less receptors than their wild-type counterpart. These results suggest that the sensitivity of tobacco protoplasts to auxin could be dependent on the number of functional receptors exposed at the outer surface of the plasmalemma.