GIBBERELLIC ACID-INDUCED MICROTUBULE REORIENTATION IN DWARF PEAS IS ACCOMPANIED BY RAPID MODIFICATION OF AN ALPHA-TUBULIN ISOTYPE

To test whether shifts induced in microtubule orientation by gibberell ic acid (GA(3)) involved changes in tubulin isotypes, pea stem cells w ere examined when elongation had been enhanced by GA(3). The behaviour of a dwarf recessive mutation (le), with very low endogenous levels o f gibberellin, was compared with the tall (Le) plant. Two hours after adding GA(3), cells were measurably longer than controls and this coin cided with a net shift of microtubule orientation from longitudinal an d oblique to transverse - an effect that was more pronounced in the dw arf. There were always more cells with net-transverse microtubules in GA(3)-treated tissue than in controls, but as growth ceased, the major orientation of the microtubule arrays became oblique in both samples. Microtubule reorientation was rapid and was closely correlated with t he growth of the cells. Although changes in orientation and isotype we re monitored over a 40 h period, immunoblotting 2D gels with the well characterized antibodies YL1/2 and YOL1/34 confirmed that alterations to the alpha-tubulin constellation could be detected as early as the 2 h time point. Again the effect was especially pronounced in dwarf pla nts. In the presence of added GA(3), one alpha-tubulin isotype (design ated alpha 1) retained its position in the alpha-tubulin constellation (as determined by total protein staining and with YOL1/34 that recogn izes detyrosinated as well as tyrosinated tubulin). It was no longer r ecognized, however, by the anti-tyrosinated alpha-tubulin antibody YL1 /2. This indicates that as GA(3) begins to cause a reorientation of th e cortical microtubules (and to enhance the rate of cell elongation) t he alpha 1 isotype is rapidly changed, probably by post-translational modification.