The Effect of Paclitaxel on Protein Phosphorylation and Reorganization of Intermediate Filaments in Rat Brain Tumor Cells and Human Non-small Cell Lung Cancer Cells

• Main Authors: Jao-Jia Chu, 朱兆嘉
• Other Authors: Yiu-Kai Lai
• Format: Others
• Language: en_US
• Published: 2005
• Online Access: http://ndltd.ncl.edu.tw/handle/46991103396193447955

博士 === 國立清華大學 === 生命科學系 === 93 === we report that taxol induces hyperphosphorylation and reorganization of the vimentin intermediate filament in 9L rat brain tumor cells, in concentration- and time-dependent manner. Phosphorylation of vimentin was maximum at10-6 M of taxol treatment for 8 h and diminished at higher (10-5 M) concentration. Enhanced phosphorylation of vimentin was detectable at 2 h treatment with 10-6 M taxol and was maximum after 12 h of treatment. Taxol-induced phosphorylation of vimentin was largely abolished in cells pretreated with staurosporine and bisindolymaleimide but was unaffected by H-89, KT-5926, SB203580, genistein, and olomoucine. Thus, protein kinase C may be involved in this process. Hyperphosphorylation of vimentin was accompanied by rounding up of cells as revealed by scanning electron microscopy. Moreover, there was a concomitant reorganization of the vimentin intermediate filament in the taxoltreated cells, whereas the microtubules and the actin microfilaments were less affected. Based on these finding, we further study the mechanism of paclitaxel resistance in paclitaxel resistance. We have established a paclitaxel-resistant mutant cell line called H460/TAX, which was derived from human non-small cell lung cancers (NSCLC) H460. A 64-fold more resistant was shown in our assay compared with the parental cells. High specificity of drug resistance was also observed in that this mutant was not cross-resistant to several other anticancer drugs. Drug acccumulation in H460/TAX was significantly less than that in H460. Many endogenous protein profiles were intact, including the expression level of P-glycoprotein, multidrug resistance-associated protein, the 70 kDa heat shock proteins as well as the phosphorylation of Bcl-2 in H460/TAX cells except that total amount of a- and b-tubulins was higher in H460/TAX than in H460 cells. Higher drug concentration and longer treatment for paclitaxel were required in H460/TAX to exert the phosphorylation of keratin 19 which was then accompanied by reorganization of the intermediate filament and the microtubule networks. The fact that H460/TAX defects only in paclitaxel-targeted pathway and leaving intact with those previous proposed factors suggesting a yet identified factor might be independently involved in the mechanism of paclitaxel resistance in human lung cancers.