PROTEIN GLYCOSYLATION IN A HEAT-RESISTANT RAT FIBROBLAST CELL MODEL EXPRESSING HUMAN HSP70

Thermotolerance and heat resistance are frequently associated with ele vated levels of heat shock proteins (HSPs). Elevated heat resistance i s also found to be associated with the overexpression of high levels o f HSP70, as seen in M21 cells, derived from the Rat-1 line. In the pre sent study, we report that M21 cells also feature an increase in gener al protein glycosylation and specific expression of the stress glycopr otein, GP62, both of which correlate with cellular heat resistance. Th e expression of GP50, a major stress glycoprotein in cell lines such a s CHO, however, did not correlate with cellular heat resistance in M21 cells. Protein glycosylation that occurs during acute heat stress ('' prompt'' glycosylation) was associated with the glycosylation of a maj or ''prompt'' stress glycoprotein, P-SG64 (M(r) of 64,000), that was i dentified by immunoblotting as a glycosylated form of calreticulin. Th e higher level of protein glycosylation in M21 cells correlated well w ith increased D-[2-H-3]mannose incorporation into precursor pools of d olichyl phosphomannose and dolichyl pyrophosphoryl oligosaccharides an d into glycoproteins. Thus, heat resistance in M21 cells is associated not only with expression of high levels of HSP70, but also with a con comitant increase in protein glycosylation. These data support the hyp othesis that stress-induced protein glycosylation is a component of ce llular stress response, either in association with HSPs or as an indep endent mechanism. (C) 1997 Academic Press.