Ralstonia Solanacearum Species Complex and Bacterial Wilt Disease

Abstract

In many cell types, a small subset of mlcrotubules (MfTs) are unusually long-lived compared with the major- ity of the MTs. These "stable" MTs may be Important mediators of differentative events since they are usually found aligned with developing asymmetries of cels undergoing morphogenesis. In addition to their longevity, the stable MTs are more resitant to drug depolymerization and are enriched in post- transationally detyrosinated tubulin (Glu-tubulin). To determine the role of protein phosphorylatlon in the regulation of these stable MTs, we treated NIH 3T3 fibroblasts and TC-7 moukey kidney epitheal cells with okadaic acid (OA) and calyculnA, potent inhibitors of protein phosphatases 1 and 2A (PP1 and PP2A), and then localized dynamic MTs and stable MTs with antibodies specfflc for tyrosinated tubuln (Tyr- tubulin) and Glu-tubulHn, respectively. OA at 0.1-10 pM caused a rapid and complete breakdown of Glu-MTs (MTs enriched in Glu-tubulin) in both cell types without substantially affecting the number of Tyr-MTs. While all concentrations of OA over this range resulted in a complete loss of Glu-MTs, the onset of Glu-MT breakdown was proportional to the logarithm of the OA concentration. The inactive analog of OA, 1-norokadaone, had no effect at any concentration. Calyculin A also caused a selective loss of Glu-MTs but was effective at 10nM, consistent with Its more potent inhibition of PP1. That the loss of Glu-MTs reflected the loss of stable MTs from the cells was shown by the absence ofnocodazole-resistantMTs in OA-treated cells. OA did not appear to actvate a MT-severing activity, since no MT fragments were observed after OA treatment of cells pretreated with taxol.These results suggest that PP1 and perhaps PP2A are involved in the regulation of MT stability in cells and show that the dynamic and stable subsets of MTs are regulated differentially by protein phosphorylation.