Stressful environments often lead to protein unfolding and the formation of cytotoxic aggregates that can compromise cell survival. The molecular chaperone heat shock protein (HSP) 101 is a protein disaggregase that co-operates with the small HSP (sHSP) and HSP70 chaperones to facilitate removal of such aggregates and is essential for surviving severe heat stress. To better define how HSP101 protects plants， we investigated the localization and targets of this chaperone in Arabidopsis (). By following HSP101 tagged with GFP， we discovered that its intracellular distribution is highly dynamic and includes a robust， reversible sequestration into cytoplasmic foci that vary in number and size among cell types and are potentially enriched in aggregated proteins. Affinity isolation of HSP101 recovered multiple proteasome subunits， suggesting a functional interaction. Consistent with this， the GFP-tagged 26S proteasome regulatory particle non-ATPase (RPN) 1a transiently colocalized with HSP101 in cytoplasmic foci during recovery. In addition， analysis of aggregated (insoluble) proteins showed they are extensively ubiquitylated during heat stress， especially in plants deficient in HSP101 or class I sHSPs， implying that protein disaggregation is important for optimal proteasomal degradation. Many potential HSP101 clients， identified by mass spectrometry of insoluble proteins， overlapped with known stress granule constituents and sHSP-interacting proteins， confirming a role for HSP101 in stress granule function. Connections between HSP101， stress granules， proteasomes， and ubiquitylation imply that dynamic coordination between protein disaggregation and proteolysis is required to survive proteotoxic stress caused by protein aggregation at high temperatures.