Synthesis and Accumulation of αB Crystallin in C6 Glioma Cells Is Induced by Agents That Promote the Disassembly of Microtubules

Abstract

When C6 cells in culture were exposed at 37 degrees C to 1 microM colchicine or to 1 microM colcemid, a tubulin-binding antimitotic alkaloid, levels of alphaB crystallin in cells began to increase after about 10 h, reaching a maximum of more than 1 microg/mg protein after 24 h. The level of alphaB crystallin returned to near the control level within two subsequent days of culture in the normal medium. Northern blot analysis showed that the accumulation of alphaB crystallin was preceded by an increase in the level of the mRNA for alphaB crystallin. Nuclear run-off transcription assays showed that colchicine induced new synthesis of mRNA for alphaB crystallin. Immunofluorescence staining revealed that alphaB crystallin accumulated in the peripheral areas of cells, as did the depolymerized tubulin, after several hours of treatment with colcemid, and then it gradually became more conspicuous in the cytoplasm. Vinblastine and nocodazole, which also promote the disassembly of microtubules by binding to tubulins, also induced the synthesis of alphaB crystallin. Furthermore, induction of alphaB crystallin by these drugs was observed in quiescent cells that had been cultured in serum-free medium. However, taxol, a microtubule-stabilizing antimitotic agent, did not stimulate the synthesis of alphaB crystallin, but rather, it suppressed the induction of synthesis of alphaB crystallin by the microtubule-disrupting drugs. Induction of alphaB crystallin by colchicine or by other drugs that promote the disassembly of microtubules was sensitive to staurosporine, an inhibitor of protein kinases, and the induction was completely suppressed in the presence of 10 nM staurosporine. These results suggest that the expression of alphaB crystallin is stimulated, via phosphorylation reactions that are sensitive to staurosporine, when the depolymerization of microtubules is enhanced.