A pathway of multi-chaperone interactions common to diverse regulatory proteins: estrogen receptor, Fes tyrosine kinase, heat shock transcription factor Hsf1, and the aryl hydrocarbon receptor

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Hop is a 60-kDa protein characterized by its ability to bind the two chaperones, hsp70 and hsp90. We have tested the function of Hop using an assay for the refolding of denatured firefly luciferase. We show that Hop is involved in the process of refolding thermally denatured firefly luciferase in rabbit reticulocyte lysate. Hop also stimulates refolding by hsp70 and Ydj-1 in a purified refolding system. Hsp90 can also stimulate refolding, and optimal refolding is observed in the presence of both Hop and hsp90. Similar stimulation was observed when Hop was replaced by its yeast homolog Sti1. In assays of the binding of Hop to hsp70 and hsp90, Hop preferentially forms a complex with ADP-bound hsp70, and this process is unaffected by the presence of hsp90. Hop does not alter the ATPase activity or the rate of ADP dissociation of hsp70. Hop also appears to bind to the ADP-bound form of hsp90, blocking the ATP-dependent conversion of hsp90 to a form capable of interacting with p23. Conversely, once p23 is bound to hsp90, Hop binding is diminished. These results confirm that Hop provides a physical link between hsp70 and hsp90 and also indicate that Hop modulates the activities of both of these chaperone proteins.

mentioning

confidence: 99%

Hop Modulates hsp70/hsp90 Interactions in Protein Folding

Johnson

Schumacher

Ross

et al. 1998

338

297

“…For instance, the dioxin receptor is a ligand-dependent transcription factor which, in the absence of ligand, is found in the cytoplasmic compartment of the cell or, depending on cell type, evenly distributed between the cytoplasm and the nucleus (1,2). This non-activated form of the receptor interacts with the hsp90 1 -dependent molecular chaperone complex (3) and its associated proteins, such as the hsp90-associated factor p23 (4,5) and the dioxin receptor specific immunophilin XAP-2 (6 -8).…”

mentioning

confidence: 99%

Two Parallel Pathways Mediate Cytoplasmic Localization of the Dioxin (Aryl Hydrocarbon) Receptor

Berg

Pongratz

2002

The dioxin receptor is a ligand-dependent transcription factor that mediates the biological effects of dioxin and related environmental pollutants. In the absence of ligand the receptor is present in the cytoplasmic compartment of the cell associated with the hsp90-dependent chaperone complex. This complex regulates several functions of the receptor such as ligand binding and nuclear import. Furthermore, intracellular localization of the receptor is modulated by multiple factors such as the export protein CRM-1 and the hsp90-associated immunophilin XAP-2. We have identified the mechanism of XAP-2-induced cytoplasmic localization of the receptor and studied the potential cross-talk between CRM-1 and XAP-2. We show that XAP-2 anchors the ligand-free receptor to cytoskeletal structures. This effect is blocked upon treatment with the actin inhibitor cytochalasin B, whereas the tubulin inhibitor colchicine had no effect on receptor localization. In addition, we show that the receptor interacts with CRM-1 both in the presence and absence of ligand. CRM-1-mediated nuclear export occurs independently of XAP-2. Our data provide evidence that CRM-1 and XAP-2 act in parallel through different mechanisms and target different interfaces of the receptor. These results suggest that two pathways cooperate to localize the non-activated receptor in the cytoplasmic compartment of the cell.In mammalian cells, multiple systems such as ligand binding, dimerization, and post-translational modifications have evolved to regulate the function of transcription factors. Recently, intracellular compartmentalization has emerged as a major pathway to modulate transcription factor activity. For instance, the dioxin receptor is a ligand-dependent transcription factor which, in the absence of ligand, is found in the cytoplasmic compartment of the cell or, depending on cell type, evenly distributed between the cytoplasm and the nucleus (1, 2). This non-activated form of the receptor interacts with the hsp90 1 -dependent molecular chaperone complex (3) and its associated proteins, such as the hsp90-associated factor p23 (4, 5) and the dioxin receptor specific immunophilin XAP-2 (6 -8).The interaction between the dioxin receptor and hsp90 is mediated by two distinct structural determinants, the DNA binding bHLH domain (9), and the ligand binding region within the PAS-B subdomain of the receptor (10). In the presence of ligand the receptor accumulates in the nucleus (1, 11) where it interacts with the general dimerization partner factor ARNT (12). This event induces release of the hsp90 complex (4, 13). Nuclear accumulation of the receptor is regulated by a nuclear localization signal present in the bHLH domain (14) and by the hsp90 complex which regulates the interaction between the import machinery and the activated form of the receptor (1). In addition, we have recently shown that the non-activated form of the receptor is shuttling between the nucleus and the cytoplasm (2). In this respect, nuclear export of the dioxin receptor is likely to be me...

“…Another example of a differential chaperone requirement for steroid receptor activation involves Cdc37. Cdc37 is required for the folding of several protein kinases and was proposed to be kinase specific based on its failure to interact with aryl hydrocarbon receptor, estrogen receptor, GR, or PR in vitro (15)(16)(17). On the other hand, genetic studies using the yeast Saccharomyces cerevisiae as a model system showed that Cdc37 was important for hormone-dependent activity by AR but not by GR (18).…”

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confidence: 99%

Functional Interaction of Human Cdc37 with the Androgen Receptor but Not with the Glucocorticoid Receptor

Rao

Lee

Benzeno

et al. 2001

Cdc37 is a molecular chaperone closely associated with the folding of protein kinases. Results from studies using a yeast model system showed that it was also important for activation of the human androgen receptor (AR). Based on results from the yeast model system (Fliss, A. E., Fang, Y., Boschelli, F., and Caplan, A. J. (1997) Mol. Biol. Cell 8, 2501-2509), we initiated studies to address whether AR and Cdc37 interact with each other in animal cell systems. Our results show that Cdc37 binds to AR but not to glucocorticoid receptors (GR) synthesized in rabbit reticulocyte lysates. This binding occurs via the ligand-binding domain of the AR in a manner that is partially dependent on Hsp90 and the presence of hormone. Further studies using the yeast system showed that Cdc37 is not interchangeable with Hsp90, suggesting that it functions at a distinct step in the activation pathway. Expression of a dominant negative form of Cdc37 in animal cells down-regulates full-length AR but has very little effect on an AR truncation lacking the ligand-binding domain or fulllength GR. These results reveal differences in the mechanisms by which AR and GR become active transcription factors and strengthen the notion that Cdc37 has a wider range of polypeptide clients than was realized previously.