HEAT-SHOCK-INDUCED ASSEMBLY OF HSP30 FAMILY MEMBERS INTO HIGH-MOLECULAR-WEIGHT AGGREGATES IN XENOPUS-LAEVIS CULTURED-CELLS

In this study, we have examined whether members of the small heat shoc k protein family, hsp30, were capable of forming heat-induced aggregat es in Xenopus laevis A6 kidney epithelial cells. Race-zonal centrifuga tion coupled with immunoblot analysis demonstrated the presence of hsp 30 aggregates with an estimated sedimentation coefficient of 10-16S. A lso, pore exclusion limit electrophoretic analysis of labeled protein from heat-shocked A6 cells revealed four heat-induced aggregates, incl uding a prominent 510 kDa aggregate, as well as weaker 350, 290, and 2 40 kDa aggregates. Immunoblot analysis of the aggregates employing an hsp30C antibody suggested that the 510 and 350 kDa aggregates were com prised of hsp30 protein. One- and two-dimensional SDS-PAGE analysis of the proteins isolated from the 510 kDa region of the pore exclusion l imit electrophoretic gel confirmed the presence of 30 kDa heat-induced protein. A total of eight small hsps were present in this aggregate, suggesting that virtually all of the major small hsps in Xenopus A6 ce lls were involved in aggregate formation. This study also detected the presence of heat-inducible hsp70 in the 510 kDa gel fraction containi ng the small hsps, but it could not be determined whether it was part of the multimer complex. (C) 1998 Elsevier Science Inc.