MODULATION OF THE STRESS-INDUCED SYNTHESIS OF HSP27 AND ALPHA-B-CRYSTALLIN BY CYCLIC-AMP IN C6 RAT GLIOMA-CELLS

The possible participation of cyclic AMP in the stress-induced synthes is of two small stress proteins, hsp27 and alpha B-crystallin, in C6 r at glioma cells was examined by specific immunoassays, western blot an alysis, and northern blot analysis. When C6 cells were exposed to arse nite (50-100 mu(M) for 1 h) or heat (42 degrees C for 30 min), express ion of hsp27 and alpha B-crystallin was stimulated, with levels of the two proteins reaching a maximum after 10-16 h of culture. Induction o f hsp27 was markedly enhanced when cells were exposed to arsenite in t he presence of isoproterenol (20 mu M) or epinephrine (20 mu M) but no t in the presence of phenylephrine. The stimulatory effects of isoprot erenol and epinephrine were blocked completely by propranolol, an anta gonist of beta-adrenergic receptors. Cholera toxin (2 mu g/ml), forsko lin (20 mu M), and dibutyryl cyclic AMP (2.5 mM), all of which are kno wn to increase intracellular levels of cyclic AMP, also stimulated the arsenite- or heat-induced accumulation of hsp27. Treatment of cells w ith each of these modulators alone did not result in the induction of hsp27. The level of hsp70 in C6 cells, as estimated by western blot an alysis, was also enhanced by arsenite or heat stress. However, inducti on of hsp70 by stress was barely stimulated by isoproterenol. By contr ast, induction of alpha B-crystallin by heat or arsenite stress was su ppressed when isoproterenol, cholera toxin, forskolin, or dibutyryl cy clic AMP was present during the stress period. Northern blot analysis of the expression of mRNAs for hsp70, hsp27, and alpha B-crystallin sh owed that the modulation of the stress-induced accumulation of the thr ee hsps by the various agents was regulated at the level of the corres ponding mRNA. These results indicate that stress responses of hsp70, h sp27, and alpha B-crystallin in C6 rat glioma cells are regulated diff erently and, moreover, that when the level of cyclic AM P increases in cells, the response to stress of hsp27 is stimulated but that of alph a B-crystallin is suppressed.