EFFECTS OF TAXOL ON THE POLYMERIZATION AND POSTTRANSLATIONAL MODIFICATION OF CLASS-III BETA-TUBULIN IN P19 EMBRYONAL CARCINOMA-CELLS

Undifferentiated P19 embryonal carcinoma cells and P19 cells induced t o differentiate along a neuronal pathway by 10(-6) M retinoic acid wer e treated with taxol to examine the effects of this microtubule-stabil izing drug on the subcellular sorting of class III beta-tubulin and on neurite outgrowth. P19 cells were grown on cover slips and then treat ed with taxol at concentrations of 10(-6) to 10(-9) M for 24 h. The mi crotubule cytoskeleton was examined after double-immunofluorescence la belling with a monoclonal antibody to alpha-tubulin (YOL 1/34) and a m onoclonal neuron-specific class III beta-tubulin antibody (TuJ1). Trea tment of undifferentiated P19 cells with concentrations of taxol great er than 4 x 10(-8) M caused microtubule bundling and multiple aster fo rmation and promoted polymerization of the low levels of class III bet a-tubulin found in these cells. In neurons, at 2 x 10(-8) M taxol, bun dling of microtubules at the base of the neurite was apparent. At taxo l concentrations greater than 1 x 10(-7) M, enhanced assembly of class III beta-tubulin was apparent, although long neurites were not observ ed. Using isoelectric focusing followed by western blotting, we detect ed an additional isoform of class III beta-tubulin after treatment wit h 10(-6) M taxol. These results indicate taxol treatment alters the no rmal subcellular sorting of tubulin isotypes, promotes the polymerizat ion and posttranslational modification of class III beta-tubulin, and interferes with neurite outgrowth.