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J. Dinny Graham,2,,
M. Greg Abel,2
Twila A. Jackson,2
David F. Gordon,2
William M. Wood,2
Sara J. Felts,1
Kathryn B. Horwitz,2 and
David Toft1*
Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, Minnesota,1 Division of Endocrinology, Department of Medicine, University of Colorado Health Science Center, Denver, Colorado2
Received 13 September 2005/ Returned for modification 9 October 2005/ Accepted 19 December 2005
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ABSTRACT |
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INTRODUCTION |
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Cell-free systems using the purified proteins have been of crucial importance in dissecting the ordered pathway that leads to the hormone-responsive state of the receptor (36, 39, 52). This hsp90-dependent chaperoning pathway is initiated by hsp40 and hsp70 binding to PR, followed by the binding of Hop-hsp90 to hsp70 (16). This intermediate complex is then modified by loss of hsp70 and Hop and recruitment of p23, resulting in a receptor able to bind hormone. In the cell, this last step also incorporates the cochaperone FKBP51, FKBP52, or Cyp40, but this step has eluded dissection in vitro.
Although the overall mechanism might be as described above, details and dynamics of this process are still unclear. Furthermore, several additional proteins have been discovered recently that interact with hsp90 or hsp70, such as Chip, Bag1, TPR2, and Aha1 (23, 31, 36, 39, 52). These are all likely to have important roles in some aspect of hsp70/hsp90 chaperoning to provide a more intricate and versatile process than that described in the current model. There are also a number of proteins that interact with the PR as transcriptional coregulators or to relate PR functions with other cell signaling pathways (25).
In a search for additional PR binding factors, we used yeast two-hybrid analysis and the PR hormone-binding domain as a target. This revealed the protein GCUNC-45 that is thought to act in the folding and functioning of myosins (37). Here we show that GCUNC-45 can bind directly and simultaneously to either PR isoform and to hsp90. The latter involves a tetratricopeptide repeat (TPR) domain of GCUNC-45 and has revealed a new site for TPR protein regulation in the N-terminal domain of hsp90. GCUNC-45 can inhibit the activation of hsp90's ATPase activity by Aha1 and progression of PR chaperoning in vitro. However, this effect is relieved by FKBP52, and GCUNC-45 clearly acts as a positive effector of PR activity in the cell.
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MATERIALS AND METHODS |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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gt-11 cDNA library. GST pull-down. The GCUNC-45 cDNA was subcloned into pGEX-4T-1 (Promega) and expressed in BL21-CodonPlus Escherichia coli (Stratagene, LaJolla, CA) as a C-terminal fusion to glutathione S-transferase (GST). The expressed GCUNC-45 GST fusion or GST alone bound to glutathione-Sepharose 4B (Amersham-Pharmacia Biotech) was incubated with HeLa whole-cell extracts containing transfected PR-A, PR-B, or glucocorticoid receptor (GR) for 2 h at 4°C. Where indicated, samples were incubated in the presence of 10 nM R5020, 100 nM RU486, 10 nM dexamethasone, or ethanol vehicle. In this instance, the PR- or GR-transfected HeLa cells had also been exposed to the same treatment for 1 h prior to harvesting and preparation of whole-cell protein extracts. Samples were washed twice with binding buffer (20 mM HEPES, pH 7.9, 60 mM NaCl, 0.1 mM EDTA, 6 mM MgCl2, 1 mM dithiothreitol [DTT], 0.5 mM phenylmethylsulfonyl fluoride, 10% glycerol, complete protease inhibitor) containing 0.05% NP-40 and twice with binding buffer alone. Bound proteins were separated on denaturing 7.5% polyacrylamide gels and visualized by immunoblotting.
Cell culture and transcription assays. The T47Dco and T47D-Y breast cancer cell lines have been described previously (18, 41). HeLa cells (American Type Culture Collection, Manassas, VA) were maintained as continuous monolayer cultures in modified Eagle's medium (MEM) supplemented with 6 ng/ml insulin (Invitrogen-Gibco BRL) and 5% fetal bovine serum (FBS; HyClone Laboratories, Logan, UT) at 37°C and 5% CO2. Transient transfections were carried out in phenol red-free MEM containing 5% charcoal-stripped FBS, using a standard calcium phosphate precipitate-mediated method followed by 24 h of hormone treatment. The cells, 2 x 105/10-cm dish, were harvested and lysed, and ß-galactosidase and luciferase activities were measured. The pA3-PRE2LUC construct was used to measure PR activation. This contained two copies of the PRE from the tyrosine aminotransferase gene promoter (22) cloned upstream of a firefly luciferase gene (51). The pCH110 (Amersham-Pharmacia) construct, constitutively expressing ß-galactosidase, was used to normalize for transfection efficiency.
siRNA knockdown techniques. HeLa cells were used containing a chloramphenicol acetyltransferase (CAT) reporter gene composed of two copies of the PRE (22) upstream of the CAT gene (42). Cells at 30 to 40% confluence in 6-well plates were transfected with 100 nM small interfering RNA (siRNA) duplexes to GCUNC-45, SRC-1, SRC-2, FKBP51, or FKBP52 using DharmaFect 1 reagent according to the manufacturer's suggestions (Dharmacon). The siRNA sequences for FKBP51 and FKBP52 were generously supplied by Michael Chinkers. The nonspecific control VIII was used to assess the non-sequence-specific effects. This control was chosen because it has the same GC content (52%) as that of the functional GCUNC-45 siRNA used and it lacks homology with known gene targets. Cells were harvested at 90 h after transfection and were treated with R5020 (0 to 250 nM; Perkin Elmer) for the last 18 h. Protein expression in the cell lysates was assessed by Western blotting from 25 µg protein, and the level of CAT protein was measured by enzyme-linked immunosorbent assay (Roche) using 50 µg protein, as instructed by the supplier (Roche).
GCUNC-45 expression and purification. GCUNC-45 with a thrombin cleavage site followed by a 6x His tag at its C terminus was cloned in pET-23a vector using NdeI and NotI restriction sites. The cDNA was transformed into BL21-CodonPlus (DE3)-RIL cells that were induced with 0.5 mM isopropyl-ß-D-thiogalactopyranoside for 3 h at 30°C. Cells were suspended in 20 ml Buffer A (10 mM Tris, 50 mM KCl, 2.5% glycerol) supplemented with protease inhibitors [0.1 mM leupeptin, 0.1 mg/ml bacitracin, 77 µg/ml aprotinin, 1.5 µM of pepstatin, 1 mM 4-(2-aminoethyl) benzensulfonyl fluoride]. After sonication, the cell lysate was supplemented with 5 mM imidazole, 400 mM KCl and clarified by ultracentrifugation (100,000 x g) for 45 min. The supernatant was loaded onto a TALON metal affinity column (BD Bioscience) equilibrated with Buffer A supplemented with 5 mM imidazole and 400 mM KCl. Unbound protein was removed, and GCUNC-45 was eluted with Buffer A containing 100 mM imidazole. GCUNC-45 was further purified using a UnoQ column (Bio-Rad), a heparin-Sepharose column where GCUNC-45 was collected in the unbound protein fraction, and finally a Superdex-200 column (Pharmacia).
Protein purification. Human hsp90 beta was overexpressed in SF9 cells and purified as described previously (48). Hsp70, Ydj1, Hop, and p23 were expressed and purified as described previously (24).
Monoclonal antibody production. Purified GCUNC-45 was used to produce a mouse monoclonal immunoglobulin G antibody named AbS1, using conventional methods (47) with the assistance of Thomas Beito in the Mayo Antibody Core Facility.
Progesterone receptor complex assembly with purified proteins.
PR from chick oviduct was adsorbed onto PR22 antibody-protein A-Sepharose and was assembled into complexes as described by Kosano et al. (24). The incubation contained 
0.05 µM PR plus 1.4 µM hsp70, 0.8 µM hsp90 dimer, 0.2 µM Ydj-1, and 0.08 µM Hop plus 2.6 µM p23. The samples also contained 20 mM Tris, pH 7.5, 5 mM MgCl2, 2 mM dithiothreitol, 0.01% NP-40, 50 mM KCl, and 5 mM ATP. After incubation for 7 min at 37°C, 0.1 µM [3H]progesterone (American Radiolabeled Chemicals, Inc., St. Louis, MO) was added for incubation on ice for 3 h. The complexes were then isolated on antibody resin and assessed for bound progesterone and for protein composition.
Site-directed mutagenesis. The pET23a-hGCUNC-45 plasmid was used as a template for the generation of double-amino-acid substitution mutants in the TPR and NR regions using the QuikChange site-directed mutagenesis kit (Stratagene, La Jolla, CA). In the TPR1 motif, lysine 13 was changed to glutamic acid and alanine 20 was changed to aspartic acid. In TPR2, both equivalent residues were changed to glutamic acid. In the four NR motifs, residues 1 and 4 were changed from leucines to serines. All mutant clones were sequenced to confirm the mutations and the authenticity of the remaining GCUNC-45 sequence.
Cytosol preparation and immunoprecipitation. Cells were lysed in Buffer A with protease inhibitors. After centrifugation (100,000 x g, 45 min), lysates were used for immunoprecipitations of PR complexes, hsp90 complexes, or GCUNC-45 using antibodies AB52 for PR, H9010 for human hsp90, and AbS1 for GCUNC-45. Immunoprecipitation reactions were carried out by incubation of immunoresins with the cytosol for 90 min in ice. After five 1-ml washes with binding buffer, bound proteins were eluted with sodium dodecyl sulfate (SDS) sample buffer, resolved by SDS-10% polyacrylamide gel electrophoresis (PAGE), and transferred to polyvinylidene difluoride membranes. Proteins were then detected by Western blotting with the appropriate antibody.
GCUNC-45 binding assay. GCUNC-45 was covalently linked to Sepharose 4B as described by Cornillot et al. (8). Bovine serum albumin (BSA) was also immobilized and used as a control. To test the binding of hsp90 and its fragments to GCUNC-45 resin, we used 25 µl resin and 20 µg protein in a 200-µl final reaction volume of incubation buffer. Samples were incubated for 30 min at 37°C and washed four times with 1 ml buffer. Bound proteins were extracted with sample buffer and resolved by SDS-PAGE.
ATPase assay.
The ATPase activity of hsp90 was measured using [
-32P]ATP as described by Owen et al. (32).
Quantification of protein levels. Arbitrary densitometric values (AU) for the indicated electrophoretic bands of protein were obtained by using IP Lab Gel 1.5e software (Signal Analytics). For evaluation of the binding of GCUNC-45 to hsp90, the mean AU values were converted to micromolar equivalents by normalizing them against 1-µg samples of purified GCUNC-45 run on the same gel. To obtain the values of free GCUNC-45, the amount of bound GCUNC-45 was subtracted from the initial amount added to the sample. The Scatchard plot was linear, with an equation of Y = –22.9 x 0.5951 corresponding to a Kd of 43 nm with the scatter illustrated by R2 = 0.9914.
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RESULTS |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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GCUNC-45 binds directly to PR. Immunoprecipitation was used to further study the binding of GCUNC-45 to both receptor forms A and B. Two variants of the human breast cancer cell line T47D were analyzed; one, YA, expressed exclusively the PR-A isoform, and the other, YB, expressed exclusively PR-B. Antibodies were used to immunoprecipitate PR-A, PR-B, or GCUNC-45 from cell lysates, and the proteins were resolved by SDS-PAGE and analyzed by Western blotting. As shown in Fig. 1C, GCUNC-45 was detected in complexes with each PR isoform. When an antibody to GCUNC-45 was used for immunoprecipitation, coprecipitation of PR-A was clearly observed, but that for PR-B was barely detectable with this technique. Thus, both receptor isoforms can be observed in complexes with GCUNC-45, although their abundance in these complexes may differ.
The results in Fig. 1C could not distinguish GCUNC-45-PR binding from indirect interactions of GCUNC-45 with other PR-associated proteins. To test for direct interaction, PR-A and PR-B were immunoprecipitated from chicken oviduct cytosol, stripped of associated proteins, and incubated with purified GCUNC-45. Chicken PR was used because preliminary studies showed that human and chicken PR interact in a similar manner with GCUNC-45 (data not shown), and methods for chicken PR isolation and analysis in vitro are much more clearly defined than those for human PR. Binding of GCUNC-45 to PR was observed by antibody pull-down and was very temperature dependent, with optimal binding at 37°C (Fig. 1D). The interaction was relatively inefficient at temperatures below 30°C, indicating that the binding may be accompanied by significant conformational changes in either GCUNC-45 or PR. In the experiment of Fig. 1B, GCUNC-45 binding was observed at 4°C, and higher temperatures were not feasible because of PR instability in the lysate. It is possible that the binding is facilitated by other associated factors in cell lysates. Note that the binding of GCUNC-45 is seen as an increased intensity of the bands in the PR-B region, since the two proteins have similar mobilities.
GCUNC-45 binds directly to hsp90 through its TPR domain. Previous studies have shown that the UNC-45 from Caenorhabditis elegans, which has 50% sequence identity with human GCUNC-45, binds to hsp90 through its TPR domain (1). To test the direct binding of GCUNC-45 to hsp90 in vitro, we used antibody pull-down experiments with purified hsp90 and GCUNC-45. As shown in Fig. 2A and B, GCUNC-45 binds to hsp90, and this is saturable and of high affinity (Kd = 43 nM). We next tested N-terminal and C-terminal fragments of GCUNC-45 for their binding to hsp90 (Fig. 2C). The binding of residues 1 to 284 (designated fragment 1-284), which contains the TPR domain, (lane 4) was similar to that of the full-length protein (lane 2), but no binding of fragment 616-944 to hsp90 was observed (lane 6). Attempts to test fragments containing only the TPR motifs failed because these were poorly expressed and unstable.
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GCUNC-45 is a positive modulator of PR activity in vivo.
Because NR box-containing proteins have been shown to act as coactivators, we tested GCUNC-45 for its ability to regulate transcriptional activation by PR under conditions of ligand excess using a HeLa cell model. HeLa cells were used because they provided a model where the PR requirement for effects could be readily tested, and, in comparison to T47D cells, they were found to be more sensitive to PR activation and allowed more effective transfection experiments. When transfected into PR-negative HeLa cells with PR-B and a progestin-responsive luciferase reporter, reporter activity was weakly, but consistently, enhanced (2-fold) with increasing amounts of GCUNC-45 (Fig. 3A). This suggested that GCUNC-45 slightly increased the availability of PR to activate transcription on its reporter.
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GCUNC-45 is a predominantly cytoplasmic protein. To measure the cellular localization of GCUNC-45, a fusion protein construct with GFP (green fluorescent protein) linked to the N terminus of GCUNC-45 was expressed in T47D-YB cells expressing a high level of PR-B. The cells were treated with either R5020 or RU486 to see if it would alter localization of GCUNC-45. Figure 4 shows that GCUNC-45 is distributed throughout the cytoplasm and is predominant in the perinuclear region. Little protein is evident in the nucleus, and this distribution is unaffected by R5020 or RU486 treatment. PR was also visualized immunohistochemically and was localized mainly in the nuclei with some weak cytoplasmic staining in the untreated cells (results not shown). Thus, GCUNC-45 does not appear to colocalize with ligand-activated nuclear PR, which is consistent with a role in the chaperoning of PR.
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GCUNC-45 regulates the ATPase activity of hsp90. The properties of hsp90 are strongly influenced by factors that regulate its conformation. For example, binding of the cochaperone p23 to hsp90 requires bound ATP, and this is inhibited by ADP or the hsp90 inhibitor geldanamycin (48). However, Hop favors hsp90 that lacks ATP, and its binding blocks the binding and hydrolysis of ATP (17, 38). The weak ATPase activity of hsp90 is important for its biological functions (14, 28, 29). It is stimulated by the hsp90 cochaperone Aha1 (31) and inhibited by Hop (38). Using highly purified proteins, the possible influence of GCUNC-45 on the ATPase activity of hsp90 was tested and compared to that of Aha1 and to that of Hop. GCUNC-45 did not itself have any ATPase activity and had no effect on the ATPase activity of hsp90 alone (data not shown). However, GCUNC-45 blocked the stimulation of ATPase activity by Aha1 (Fig. 6A). This inhibition was dose dependent and nearly as effective as that of Hop (Fig. 6B). However, the immunophilin FKBP52, which binds to the C terminus of hsp90 through its TPR domain (33), showed no inhibitory activity.
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DISCUSSION |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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GCUNC-45 has only recently been identified as a protein closely related to SMUNC-45, and it appears to be unique to vertebrates (37). Because all other members of the UCS family have been implicated in the processing and functions of myosins, GCUNC-45 was proposed to be a myosin chaperone for non-muscle cells that would be involved in cytoskeletal functions (37). It has been shown to differ from SMUNC-45 in its tissue distribution and pattern of expression during embryogenesis. Antisense experiments indicated that a loss of GCUNC-45 suppressed myotube formation in muscle cells and reduced the rate of proliferation in non-muscle cells (37). Our results show that GCUNC-45 activities are not confined to myosin interaction. One can even consider the possibility that the myosin-binding activity might be important to the PR chaperone complex, perhaps as a means for localization or trafficking of the complex.
The presence of potential NR box motifs initially suggested that GCUNC-45 might be a transcriptional coregulator of PR. We have not excluded this possibility, but its primary localization in the cytoplasm and its association with hsp90 indicates a role in PR chaperoning. We have found that the LXXLL sequences are not essential for binding of GCUNC-45 to PR in vitro, although their mutation reduces binding somewhat (results not shown).
UNC-45 from Caenorhabditis elegans has been reported to interact with hsp90 through its TPR motifs (1). This interaction appeared to involve the C terminus of hsp90, based on competition by a peptide containing the C-terminal sequence MEEVD. Because of these results and the proposed mechanism for the interaction of hsp90 with UNC-45 (1) and other cochaperones (6, 43, 53), it seemed likely that the TPR recognition motif in the C terminus of hsp90 would be involved in the interaction with GCUNC-45. However, instead, a new site within the N-terminal ATP-binding domain of hsp90 was revealed. The existence of a second TPR-binding site on hsp90 opens new possibilities for identifying factors that might bind to this site to regulate hsp90 functions and may stimulate a reassessment of known chaperone-cochaperone interactions. Perhaps some cochaperones can recognize either TPR-binding site depending on the conformational state of hsp90. An intriguing parallel may exist with the complex of Hip-hsp70-Hop. Like hsp90, hsp70 has an EEVD sequence at its C terminus which appears to be part of the Hop-binding domain near the C terminus of hsp70 (5, 43). However, while the cochaperone Hip binds hsp70 through its TPR domain, it binds to a site, yet to be identified, in the N-terminal ATPase domain of hsp70 (10, 34).
GCUNC-45 has a positive role in the hsp90 chaperoning pathway for PR. In the cell, its overexpression caused only a small enhancement of PR transcriptional activity to induce a reporter gene, but the loss of endogenous GCUNC-45 resulted in a marked reduction of PR activity. This reduction of PR activity appears linked to the loss of GCUNC-45, since the protein levels of PR, hsp90, hsp70, Hop, and FKBP52 were not altered by the treatment. However, in simplified cell-free systems, GCUNC-45 inhibits the ATPase activity of hsp90 and blocks progression of PR chaperoning to its hormone-binding state. We have only observed the inhibition of ATPase activity when this activity was enhanced by the presence of Aha1. Aha1 has been shown to bind to the middle domain of hsp90 and is thought to facilitate repositioning of a catalytic loop of this domain that interacts with the 
phosphate of ATP bound in the N-terminal domain (28). Thus, the binding of GCUNC-45 to hsp90 may interfere with the conformational changes needed to bring the catalytic loop of the middle domain into contact with bound ATP, or it may block the binding of Aha1 to hsp90 even though the two proteins do not seem to share a common binding site.
Since ATP binding and hydrolysis are essential to the activity of hsp90 (14, 29, 30), their inhibition explains the loss of proper PR chaperoning in the presence of GCUNC-45 as illustrated by a loss of hormone-binding activity. However, this appears not to be the case in the complete chaperoning system of the cell. The cochaperone Hop is also a potent inhibitor of the hsp90 ATPase activity, but this inhibition is confined to one step in the chaperoning process and is reversed by the association of Hop-hsp90 with hsp70 (17). As a result, Hop has a very positive effect on the chaperoning activity of hsp90. The cochaperone Cdc37 is essential for the chaperoning of many protein kinases, yet it inhibits the ATPase activity of hsp90 in vitro (45). It has been suggested that this inhibited state is more optimal for initial interaction of hsp90 with client proteins. In like manner, we propose that GCUNC-45 enters the chaperone complex at an intermediate stage where its inhibitory action serves a purpose, but that this inhibition is relieved by subsequent events that are missing from the in vitro system (Fig. 8). One such event is the binding of the immunophilin FKBP52 in the PR complex. This protein is observed in PR complexes from cell lysates and appears to interact with both PR and hsp90. Its function is still unclear, but it may be involved in modulating PR conformation or trafficking (35, 46). Future studies may reveal additional interactions between GCUNC-45 and other hsp90 cochaperones and clients.
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ACKNOWLEDGMENTS |
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This work was supported by NIH grants RO1 DK 59284 and RO1 CA26869 and by the National Foundation for Cancer Research.
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FOOTNOTES |
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A.C. and J.D.G. contributed equally to this study.
Present address: Westmead Institute for Cancer Research, University of Sydney Westmead Hospital, Westmead, New South Wales, Australia.
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REFERENCES |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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2. Blatch, G. L., and M. Lassle. 1999. The tetratricopeptide repeat: a structural motif mediating protein-protein interactions. BioEssays 21:932-939.[CrossRef][Medline]
3. Brinker, A., C. Scheufler, F. von der Mülbe, B. Fleckenstein, C. Herrmann, G. Jung, I. Moarefi, and F. U. Hartl. 2002. Ligand discrimination by TPR domains. J. Biol. Chem. 277:19265-19275.
4. Carrello, A., E. Ingley, R. F. Minchin, S. Tsai, and T. Ratajczak. 1999. The common tetratricopeptide repeat acceptor site for steroid receptor-associated immunophilins and Hop is located in the dimerization domain of hsp90. J. Biol. Chem. 274:2682-2689.
5. Carrigan, P. E., G. M. Nelson, P. J. Roberts, J. N. Stoffer, D. L. Riggs, and D. F. Smith. 2004. Multiple domains of the co-chaperone Hop are important for Hsp70 binding. J. Biol. Chem. 279:16185-16193.
6. Chen, S., W. P. Sullivan, D. O. Toft, and D. F. Smith. 1998. Differential interactions of p23 and the TPR-containing proteins Hop, Cyp40, FKBP52 and FKBP51 with Hsp90 mutants. Cell Stress Chaperones 3:118-129.[CrossRef][Medline]
7. Collingwood, T. N., F. D. Urnov, and A. P. Wolffe. 1999. Nuclear receptors: coactivators, corepressors and chromatin remodeling in the control of transcription. J. Mol. Endocrinol. 23:255-275.[Abstract]
8. Cornillot, J. D., M. Caron, R. Joubert-Caron, and D. Bladier. 1992. Use of an immobilized human endogenous lectin for the purification of complementary ligands. Int. J. Biochem. 24:1585-1589.[CrossRef][Medline]
9. Das, A. K., P. T. W. Cohen, and D. Barford. 1998. The structure of the tetratricopeptide repeats of protein phosphatase 5: implications for TPR-mediated protein-protein interactions. EMBO J. 17:1192-1199.[CrossRef][Medline]
10. Desmond, J., J. Lüders, and J. Höhfeld. 1998. The carboxy-terminal domain of Hsc70 provides binding sites for a distinct set of chaperone cofactors. Mol. Cell. Biol. 18:2023-2028.
11. Elbi, C., D. A. Walker, G. Romero, W. P. Sullivan, D. O. Toft, H. L. Hager, and D. B. DeFranco. 2004. Molecular chaperones function as steroid receptor nuclear mobility factors. Proc. Natl. Acad. Sci. USA 101:2876-2881.
12. Freeman, B. C., and K. R. Yamamoto. 2002. Disassembly of transcriptional regulatory complexes by molecular chaperones. Science 296:2232-2235.
13. Glass, C. K., D. W. Rose, and M. G. Rosenfeld. 1997. Nuclear receptor coactivators. Curr. Opin. Cell Biol. 9:222-232.[CrossRef][Medline]
14. Grenert, J. P., B. D. Johnson, and D. O. Toft. 1999. The importance of ATP binding and hydrolysis by Hsp90 in formation and function of protein heterocomplexes. J. Biol. Chem. 274:17525-17533.
15. Heery, D. M., E. Kalkhoven, S. Hoare, and M. G. Parker. 1997. A signature motif in transcriptional co-activators mediates binding to nuclear receptors. Nature 387:733-736.[CrossRef][Medline]
16. Hernández, M. P., A. Chadli, and D. O. Toft. 2002. HSP40 binding is the first step in the HSP90 chaperoning pathway for the progesterone receptor. J. Biol. Chem. 277:11873-11881.
17. Hernández, M. P., W. P. Sullivan, and D. O. Toft. 2002. The assembly and intermolecular properties of the hsp70-Hop-hsp90 molecular chaperone complex. J. Biol. Chem. 277:38294-38304.
18. Horwitz, K. B., and G. R. Freidenberg. 1985. Growth inhibition and increase of insulin receptors in antiestrogen-resistant T47DCO human breast cancer cells by progestins: implications for endocrine therapies. Cancer Res. 45:167-173.
19. Horwitz, K. B., T. A. Jackson, D. L. Bain, J. K. Richer, G. C. Takimoto, and L. Tung. 1996. Nuclear receptor coactivators and corepressors. Mol. Endocrinol. 10:1167-1177.[Abstract]
20. Hutagalung, A. H., M. L. Landsverk, M. G. Price, and H. F. Epstein. 2002. The UCS family of myosin chaperones. J. Cell Sci. 115:3983-3990.
21. Jackson, T. A., J. K. Richer, D. L. Bain, G. S. Takimoto, L. Tung, and K. B. Horwitz. 1997. The partial agonist activity of antagonist-occupied steroid receptors is controlled by a novel hinge domain-binding coactivator L7/SPA and the corepressors N-CoR or SMRT. Mol. Endocrinol. 11:693-705.
22. Jantzen, H.-M., U. Strahle, B. Gloss, F. Stewart, W. Schmid, M. Boshart, R. Miksicek, and G. Schutz. 1987. Cooperaativity of glucocorticoid response elements located far upstream of the tyrosine aminotransferase gene. Cell 49:29-38.[CrossRef][Medline]
23. King, F. W., A. Wasrzynow, J. Höhfeld, and M. Zylicz. 2001. Co-chaperones Bag-1, Hop and Hsp40 regulate Hsc70 and Hsp90 interactions with wild-type or mutant p53. EMBO J. 20:6297-6305.[CrossRef][Medline]
24. Kosano, H., B. Stensgard, M. C. Charlesworth, N. McMahon, and D. O. Toft. 1998. The assembly of progesterone receptor-hsp90 complexes using purified proteins. J. Biol. Chem. 273:32973-32979.
25. Lange, C. A., J. K. Richer, and K. B. Horwitz. 1999. Hypothesis: progesterone primes breast cancer cells for cross-talk with proliferative or antiproliferative signals. Mol. Endocrinol. 13:829-836.
26. Lapouge, K., S. J. M. Smith, P. A. Walker, S. J. Gamblin, S. J. Smerdon, and K. Rittinger. 2000. Structure of the TPR domain of p67phox in complex with Rac-GTP. Mol. Cell 6:899-907.[Medline]
27. McKenna, N. J., and B. W. O'Malley. 2002. Combinatorial control of gene expression by nuclear receptors and coregulators. Cell 108:465-474.[CrossRef][Medline]
28. Meyer, P., C. Prodromou, C. Liao, B. Hu, S. M. Roe, C. K. Vaughan, I. Vlasic, B. Panaretou, P. W. Piper, and L. H. Pearl. 2004. Structural basis for recruitment of the ATPase activator Aha1 to the hsp90 chaperone machinery. EMBO J. 23:1402-1410.[CrossRef][Medline]
29. Obermann, W. M. J., H. Sondermann, A. A. Russo, N. P. Pavletich, and F. U. Hartl. 1998. In vivo function of Hsp90 is dependent on ATP binding and ATP hydrolysis. J. Cell Biol. 143:901-910.
30. Panaretou, B., C. Prodromou, S. M. Roe, R. O'Brien, J. E. Ladbury, P. W. Piper, and L. H. Pearl. 1998. ATP binding and hydrolysis are essential to the function of the Hsp90 molecular chaperone in vivo. EMBO J. 17:4829-4836.[CrossRef][Medline]
31. Panaretou, B., G. Siligardi, P. Meyer, A. Maloney, J. K. Sullivan, S. Singh, S. H. Millson, P. A. Clarke, S. Naaby-Hansen, R. Stein, R., Cramer, M. Mollapour, P. Workman, P. W. Piper, L. H. Pearl, and C. Prodromou. 2002. Activation of the ATPase activity of Hsp90 by the stress-regulated cochaperone Aha1. Mol. Cell 10:307-1318.[CrossRef][Medline]
32. Owen, B. A. L., W. P. Sullivan, S. J. Felts, and D. O. Toft. 2002. Regulation of heat shock protein 90 ATPase activity by sequences in the carboxyl terminus. J. Biol. Chem. 277:7086-7091.
33. Owens-Grillo, J. K., M. J. Czar, K. A. Huchison, K. Hoffmann, G. H. Perdew, and W. B. Pratt. 1996. A model of protein targeting mediated by immunophilins and other proteins that bind to hsp90 via tetratricopeptide repeat domains. J. Biol. Chem. 271:13468-13475.
34. Prapapanich, V., S. Chen, E. J. Toran, R. A. Rimerman, and D. F. Smith. 1996. Mutational analysis of the hsp70-interacting protein Hip. Mol. Cell. Biol. 16:6200-6207.[Abstract]
35. Pratt, W. P., M. D. Galigniana, J. M. Harrell, and D. B. DeFranco. 2004. Role of hsp90 and the hsp90-binding immunophilins in signalling protein movements. Cell. Signal. 16:857-872.[CrossRef][Medline]
36. Pratt, W. P., and D. O. Toft. 2003. Regulation of signaling protein function and trafficking by the hsp90/hsp70-based chaperone machinery. Exp. Biol. Med. 228:111-133.
37. Price, M. G., M. L. Landsverk, J. M. Barral, and H. F. Epstein. 2002. Two mammalian UNC-45 isoforms are related to distinct cytoskeletal and muscle-specific functions. J. Cell Sci. 115:4013-4023.
38. Prodromou, C., G. Siligardi, R. O'Brien, D. N. Woolfson, L. Regan, B. Panaretou, J. E. Ladbury, P. W. Piper, and L. H. Pearl. 1999. Regulation of Hsp90 ATPase activity by tetratricopeptide repeat (TPR)-domain co-chaperones. EMBO J. 18:754-762.[CrossRef][Medline]
39. Richter, K., and J. Buchner. 2001. Hsp90: chaperoning signal transduction. J. Cell Physiol. 188:281-290.[CrossRef][Medline]
40. Russell, L. C., S. R. Whitt, M.-S. Chen, and M. Chinkers. 1999. Identification of conserved residues required for the binding of a tetracopeptide repeat domain to heat shock protein 90. J. Biol. Chem. 174:20060-20063.
41. Sartorius, C. A., S. D. Groshong, L. A. Miller, R. L. Powell, L. Tung, G. S. Takimoto, and K. B. Horwitz. 1994. New T47D breast cancer cell lines for the independent study of progesterone B- and A-receptors: only antiprogestin-occupied B-receptors are switched to transcriptional agonists by cAMP. Cancer Res. 54:868-3877.
42. Sartorius, C. A., M. Y. Melville, A. R. Hovland, L. Tung, G. S. Takimoto, and K. B. Horwitz. 1994. A third transactivation function (AF3) of human progesterone receptors located in the unique N-terminal segment of the B-isoform. Mol. Endocrinol. 8:1347-1360.[Abstract]
43. Scheufler, C., A. Brinker, G. Bourenkov, S. Pegoraro, L. Moroder, H. Bartunik, F. U. Hartl, and I. Moarefi. 2000. Structure of TPR domain-peptide complexes: critical elements in the assembly of the Hsp70-Hsp90 multichaperone machine. Cell 101:99-210.
44. Shibata, H., T. E. Spencer, S. A. Onate, G. Jenster, S. Y. Tsai, M. J. Tsai, and B. W. O'Malley. 1997. Role of co-activators and co-repressors in the mechanism of steroid/thyroid receptor action. Recent Prog. Horm. Res. 52:141-164.[Medline]
45. Siligardi, G., B. Panaretou, P. Meyer, S. Singh, D. N. Woolfson, P. W. Piper, L. H. Pearl, and C. Prodromou. 2002. Regulation of Hsp90 ATPase activity by the co-chaperone Cdc37p/p50cdc37. J. Biol. Chem. 277:20151-20159.
46. Smith, D. F. 2004. Tetratricopeptide repeat cochaperones in steroid receptor complexes. Cell Stress Chaperones 9:109-121.[CrossRef][Medline]
47. Sullivan, W. P., T. G. Beito, J. Proper, C. J. Krco, and D. O. Toft. 1986. Preparation of monoclonal antibodies to the avian progesterone receptor. Endocrinology 119:1549-15557.[Abstract]
48. Sullivan, W. P., B. Stensgard, G. Caucutt, B. Bartha, N. McMahon, E. S. Alnemri, G. Litwack, and D. O. Toft. 1997. Nucleotides and two functional states of hsp90. J. Biol. Chem. 272:8007-8012.
49. Tranguch, S., J. Cheung-Flynn, T. Daikoku, V. Prapapanich, M. B. Cox, H. Xie, H. Wang, S. K. Das, D. F. Smith, and S. K. Dey. 2005. Cochaperone immunophilin FKBP52 is critical to uterine receptivity for embryo implantation. Proc. Natl. Acad. Sci. USA 102:14326-14331.
50. Tsai, M.-J., and B. W. O'Malley. 1994. Molecular mechanisms of action of steroid/thyroid receptor superfamily members. Annu. Rev. Biochem. 63:451-486.[CrossRef][Medline]
51. Wood, W. M., M. Y. Kao, D. F. Gordon, and E. C. Ridgway. 1989. Thyroid hormone regulates the mouse thyrotropin beta subunit gene promoter in transfected primary thyrotropes. J. Biol. Chem. 264:14840-14847.
52. Young, J. C., I. Moarefi, and F. U. Hartl. 2001. Hsp90: a specialized but essential protein-folding tool. J. Cell Biol. 154:267-273.
53. Young, J. C., M. J. Obermann, and F. U. Hartl. 1998. Specific binding of tetratricopeptide repeat proteins to the C-terminal 12-kDa domain of hsp90. J. Biol. Chem. 273:18007-18010.
54. Yu, Q., and S. I. Bernstein. 2003. UCS proteins: managing the myosin motor. Curr. Biol. 13:R525-R527.[CrossRef][Medline]
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