RESPONSES TO HEAT-SHOCK OF ALPHA-B-CRYSTALLIN AND HSP28 IN U373-MG HUMAN GLIOMA-CELLS

Responses to heat shock of alphaB crystallin and small heat-shock protein HSP28, which are expressed at considerable levels in human astroglioma U373 MG cells (2-4 mug of each per mg soluble protein in confluent cultures), were analysed quantitatively by specific immunoassays. Concentrations of alphaB crystallin and HSP28 in soluble extracts of U373 MG cells decreased to about 50% of original values, with an increase in the insoluble fraction during heat treatment for 15 min at 45-degrees-C. The concentrations of alphaB crystallin and HSP28 increased gradually upon return to 37-degrees-C, reaching and then exceeding the control levels within 5 h and 10 h, respectively, after heat shock. During centrifugation on sucrose density gradients both alphaB crystallin and HSP28 in extracts from untreated and heat-treated cells sedimented at the same position, which corresponded to a molecular mass of > 540 kDa. This result suggests that the sizes of aggregates of the two proteins in the cytoplasm are not affected by heat shock. Both alphaB crystallin and HSP28 in an extract of U373 MG cells were trapped on and coeluted from an affinity column prepared with antibodies against alphaB crystallin. These results suggest that the two proteins are also associated in U373 MG cells.