High-molecular-mass complexes formed in vivo contain smHSPs and HSP70 and display chaperone-like activity

Stress can have profound effects on the cell. The elicitation of the stress response in the cell is often accompanied by the synthesis of high-molecul ar-mass complexes, sometimes termed heat shock granules (HSGs). The presenc e of the complexes has been shown to be important for the survival of cells subjected to stress. We purified these complexes from heat-stressed BY-2 t obacco cells. HSG complexes formed in vivo contain predominantly smHSPs, HS P40 and HSP70 and display chaperone-like activity. Tubulins as well as othe r proteins may be part of the complex or its substrate. The proteins, excep t smHSPs and to some extent HSP70, were hypersensitive to proteolysis, sugg esting that they were partially denatured and not an integral part of the H SG complexes. When citrate synthase was used as the substrate, in vivo gene rated HSG complexes exhibited strong nucleotide-dependent in vitro chaperon e activity. Measurable ATP-mediated hydrolytic activity was detected. Isola ted HSG complexes are stable until ATP is added, which leads to rapid disso ciation of the complex into subunits. It is proposed that smHSPs form the c ore of the complex in association with ATP-dependent HSP70 and HSP40 cochap erones. Implications of these findings are discussed.