Functional Comparison of Human and Drosophila Hop Reveals Novel Role in Steroid Receptor Maturation

Hsp70/Hsp90 organizing protein (Hop) coordinates Hsp70 and Hsp90 interactions during assembly of steroid receptor complexes. Hop is composed of three tetratricopeptide repeat (TPR) domains (TPR1, TPR2a, and TPR2b) and two DP repeat domains (DP1 and DP2); Hsp70 interacts directly with TPR1 and Hsp90 with TPR2a, but the function of other domains is less clear. Human Hop and the Saccharomyces cerevisiae ortholog Sti1p, which share a common domain arrangement, are functionally interchangeable in a yeast growth assay and in supporting the efficient maturation of glucocorticoid receptor (GR) function. To gain a better understanding of Hop structure/function relationships, we have extended comparisons to the Hop ortholog from Drosophila melanogaster (dHop), which lacks DP1. Although dHop binds Hsp70 and Hsp90 and can rescue the growth defect in yeast lacking Sti1p, dHop failed to support GR function in yeast, which suggests a novel role for Hop in GR maturation that goes beyond Hsp binding. Chimeric Hop constructs combining human and Drosophila domains demonstrate that the C-terminal domain DP2 is critical for this previously unrecognized role in steroid receptor function. Hsp70/Hsp90 organizing protein (Hop) coordinates Hsp70 and Hsp90 interactions during assembly of steroid receptor complexes. Hop is composed of three tetratricopeptide repeat (TPR) domains (TPR1, TPR2a, and TPR2b) and two DP repeat domains (DP1 and DP2); Hsp70 interacts directly with TPR1 and Hsp90 with TPR2a, but the function of other domains is less clear. Human Hop and the Saccharomyces cerevisiae ortholog Sti1p, which share a common domain arrangement, are functionally interchangeable in a yeast growth assay and in supporting the efficient maturation of glucocorticoid receptor (GR) function. To gain a better understanding of Hop structure/function relationships, we have extended comparisons to the Hop ortholog from Drosophila melanogaster (dHop), which lacks DP1. Although dHop binds Hsp70 and Hsp90 and can rescue the growth defect in yeast lacking Sti1p, dHop failed to support GR function in yeast, which suggests a novel role for Hop in GR maturation that goes beyond Hsp binding. Chimeric Hop constructs combining human and Drosophila domains demonstrate that the C-terminal domain DP2 is critical for this previously unrecognized role in steroid receptor function. Functional maturation of steroid receptors requires multistep assembly with molecular chaperones (1Pratt W.B. Toft D.O. Endocr. Rev. 1997; 18: 306-360Crossref PubMed Scopus (1539) Google Scholar), and Hop, which binds both Hsp70 and Hsp90, can facilitate the progression through the intermediate stages of assembly (2Odunuga O.O. Longshaw V.M. Blatch G.L. BioEssays. 2004; 26: 1058-1068Crossref PubMed Scopus (178) Google Scholar). The early steps in assembling receptor/chaperone complexes are the binding of Hsp40 to a receptor monomer (3Hernandez M.P. Chadli A. Toft D.O. J. Biol. Chem. 2002; 277: 11873-11881Abstract Full Text Full Text PDF PubMed Scopus (120) Google Scholar) followed by recruitment of Hsp70. Hop-Hsp90 complexes are then recruited to bind the receptor-associated Hsp70. In a transition that remains poorly understood, Hsp90 becomes directly associated with the receptor, and Hsp70 and Hop leave the complex. The Hsp90 co-chaperone p23 binds to Hsp90, stabilizing its association with the receptor, and immunophilin-related co-chaperones bind Hsp90 at a site vacated by Hop. Only when the receptor has achieved assembly with Hsp90 and p23 does it attain full hormone binding ability (4Scherrer L.C. Dalman F.C. Massa E. Meshinchi S. Pratt W.B. J. Biol. Chem. 1990; 265: 21397-21400Abstract Full Text PDF PubMed Google Scholar, 5Smith D.F. Mol. Endocrinol. 1993; 7: 1418-1429Crossref PubMed Scopus (251) Google Scholar). Through its ability to simultaneously bind both Hsp70 and Hsp90, Hop serves as an adaptor to coordinate the recruitment of Hsp90 to intermediate receptor complexes containing Hsp70 (6Chen S. Smith D.F. J. Biol. Chem. 1998; 273: 35194-35200Abstract Full Text Full Text PDF PubMed Scopus (281) Google Scholar). Hop can also inhibit Hsp90 ATPase, but it acts somewhat differently toward yeast or vertebrate Hsp90. Yeast Hsp90 has a much higher basal ATPase activity, and Sti1p, the Hop ortholog in Saccharomyces cerevisiae (7Nicolet C.M. Craig E.A. Mol. Cell. Biol. 1989; 9: 3638-3646Crossref PubMed Scopus (206) Google Scholar), inhibits this activity (8Prodromou C. Siligardi G. O'Brien R. Woolfson D.N. Regan L. Panaretou B. Ladbury J.E. Piper P.W. Pearl L.H. EMBO J. 1999; 18: 754-762Crossref PubMed Scopus (355) Google Scholar, 9Richter K. Muschler P. Hainzl O. Reinstein J. Buchner J. J. Biol. Chem. 2003; 278: 10328-10333Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar). In contrast, human Hsp90 has very low basal ATPase activity that can be stimulated by binding to substrate, and Hop is able to inhibit the client-stimulated ATPase (10McLaughlin S.H. Smith H.W. Jackson S.E. J. Mol. Biol. 2002; 315: 787-798Crossref PubMed Scopus (214) Google Scholar). In some settings Hop can also affect Hsp70 ATPase. For example, Sti1p dramatically stimulates activity of the yeast Ssa family of Hsp70 proteins (11Wegele H. Haslbeck M. Reinstein J. Buchner J. J. Biol. Chem. 2003; 278: 25970-25976Abstract Full Text Full Text PDF PubMed Scopus (104) Google Scholar), although Hop was not observed to stimulate the ATPase activity of vertebrate Hsp70 (12Gebauer M. Zeiner M. Gehring U. Mol. Cell. Biol. 1998; 18: 6238-6244Crossref PubMed Scopus (33) Google Scholar). Unlike most other receptor-associated chaperone components, Hop does not possess independent chaperone activity by as determined by in vitro refolding assays (13Freeman B.C. Toft D.O. Morimoto R.I. Science. 1996; 274: 1718-1720Crossref PubMed Scopus (289) Google Scholar, 14Gebauer M. Zeiner M. Gehring U. FEBS Lett. 1997; 417: 109-113Crossref PubMed Scopus (87) Google Scholar), so it is unlikely to act directly on a receptor. In addition to cell-free assembly studies, yeast models have been used to show that deletion of Sti1p inhibits steroid receptor function (15Chang H.C. Nathan D.F. Lindquist S. Mol. Cell. Biol. 1997; 17: 318-325Crossref PubMed Scopus (193) Google Scholar). Yeast lacking functional Sti1p are viable but have retarded growth at elevated temperature or in the presence of minimal media (7Nicolet C.M. Craig E.A. Mol. Cell. Biol. 1989; 9: 3638-3646Crossref PubMed Scopus (206) Google Scholar). We have shown previously (6Chen S. Smith D.F. J. Biol. Chem. 1998; 273: 35194-35200Abstract Full Text Full Text PDF PubMed Scopus (281) Google Scholar) that Sti1p can functionally replace human Hop (hHop) 1The abbreviations used are: hHop, human Hop; dHop, Drosophila Hop; TPR, tetratricopeptide repeat; RL, reticulocyte lysate; PR, progesterone receptor; GR, glucocorticoid receptor; IP, immunoprecipitation; WT, wild type. using an in vitro receptor assembly system. We have also shown (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar) that hHop can fully rescue the growth phenotype in a yeast strain (sti1Δ0) lacking the gene for Sti1p and that hHop can fully restore steroid receptor function in the sti1Δ0 background. Thus, it appears that Sti1p and hHop are functionally interchangeable. Sti1p and hHop share over 50% amino acid sequence similarity and have a common domain structure consisting of three tetratricopeptide repeat domains (TPR1, TPR2a, and TPR2b) and two small domains (DP1 and DP2) containing a characteristic DP repeat motif (17Prapapanich V. Chen S. Smith D.F. Mol. Cell. Biol. 1998; 18: 944-952Crossref PubMed Scopus (50) Google Scholar). The domain arrangement is TPR1-DP1-TPR2a-TPR2b-DP2. Two of the Sti1p TPR domains have been individually crystallized (18Scheufler C. Brinker A. Bourenkov G. Pegoraro S. Moroder L. Bartunik H. Hartl F.U. Moarefi I. Cell. 2000; 101: 199-210Abstract Full Text Full Text PDF PubMed Scopus (1010) Google Scholar) and have overall folds similar to each other and to the TPR domains from several other Hsp90-binding co-chaperones (reviewed in Ref. 19Dornan J. Taylor P. Walkinshaw M.D. Curr. Top. Med. Chem. 2003; 3: 1392-1409Crossref PubMed Scopus (101) Google Scholar). Several lines of evidence suggest that the highly conserved EEVD sequence that terminates many eukaryotic Hsp90 and Hsp70 proteins directly interacts with a TPR binding pocket in co-chaperones. Hop TPR2a is required for Hsp90 binding (20Chen S. Prapapanich V. Rimerman R.A. Honore B. Smith D.F. Mol. Endocrinol. 1996; 10: 682-693Crossref PubMed Google Scholar, 21Lassle M. Blatch G.L. Kundra V. Takatori T. Zetter B.R. J. Biol. Chem. 1997; 272: 1876-1884Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar); the Hsp90-specific peptide MEEVD co-crystallizes with TPR2a (18Scheufler C. Brinker A. Bourenkov G. Pegoraro S. Moroder L. Bartunik H. Hartl F.U. Moarefi I. Cell. 2000; 101: 199-210Abstract Full Text Full Text PDF PubMed Scopus (1010) Google Scholar), and site-directed mutation of the EEVD terminus of Hsp90 is sufficient to disrupt binding with Hop (22Chen S. Sullivan W.P. Toft D.O. Smith D.F. Cell Stress Chaperones. 1998; 3: 118-129Crossref PubMed Scopus (169) Google Scholar, 23Carrello A. Ingley E. Minchin R.F. Tsai S. Ratajczak T. J. Biol. Chem. 1999; 274: 2682-2689Abstract Full Text Full Text PDF PubMed Scopus (102) Google Scholar, 24Ramsey A.J. Russell L.C. Whitt S.R. Chinkers M. J. Biol. Chem. 2000; 275: 17857-17862Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). Therefore, EEVD-TPR interaction is critical for Hsp90 binding to Hop. Hop TPR1 is required for Hsp70 binding (20Chen S. Prapapanich V. Rimerman R.A. Honore B. Smith D.F. Mol. Endocrinol. 1996; 10: 682-693Crossref PubMed Google Scholar, 21Lassle M. Blatch G.L. Kundra V. Takatori T. Zetter B.R. J. Biol. Chem. 1997; 272: 1876-1884Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar), and the Hsp70-specific peptide PTIEEVD co-crystallizes with TPR1 (18Scheufler C. Brinker A. Bourenkov G. Pegoraro S. Moroder L. Bartunik H. Hartl F.U. Moarefi I. Cell. 2000; 101: 199-210Abstract Full Text Full Text PDF PubMed Scopus (1010) Google Scholar). On the other hand, deletion of PTIEEVD and up to 40 additional amino acids from the C terminus of Hsp70 do not prevent binding of truncated Hsp70 to Hop (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). In contrast to Hsp90, therefore, there must be one or more additional interaction modes between Hop and Hsp70, although the EEVD-TPR1 interaction is probably functionally important. Based on analysis of Hop co-crystal structures (18Scheufler C. Brinker A. Bourenkov G. Pegoraro S. Moroder L. Bartunik H. Hartl F.U. Moarefi I. Cell. 2000; 101: 199-210Abstract Full Text Full Text PDF PubMed Scopus (1010) Google Scholar), positively charged amino acids in the TPR binding pocket form a carboxylate clamp that interacts with the negatively charged EEVD. Point mutations of carboxylate clamp residues have been shown to disrupt binding of TPR-containing proteins to Hsp90 (25Russell L.C. Whitt S.R. Chen M.S. Chinkers M. J. Biol. Chem. 1999; 274: 20060-20063Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 26Ward B.K. Allan R.K. Mok D. Temple S.E. Taylor P. Dornan J. Mark P.J. Shaw D.J. Kumar P. Walkinshaw M.D. Ratajczak T. J. Biol. Chem. 2002; 277: 40799-40809Abstract Full Text Full Text PDF PubMed Scopus (58) Google Scholar) or Hsp70 (27Odunuga O.O. Hornby J.A. Bies C. Zimmermann R. Pugh D.J. Blatch G.L. J. Biol. Chem. 2003; 278: 6896-6904Abstract Full Text Full Text PDF PubMed Scopus (79) Google Scholar). However, we also observed that carboxylate clamp mutations in Hop TPR domains had complex, unexpected effects on Hsp70 binding (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). We have further observed that mutation of the C-terminal DP2 domain of Hop disrupts Hsp70 binding (6Chen S. Smith D.F. J. Biol. Chem. 1998; 273: 35194-35200Abstract Full Text Full Text PDF PubMed Scopus (281) Google Scholar). Our working model is that multiple Hop domains contribute to Hsp binding and that domain-domain interactions within Hop are likely important. To further characterize the role of individual Hop domains in Hsp binding and in functional maturation of steroid receptors, we have undertaken a functional comparison between hHop and a Hop ortholog from Drosophila melanogaster (dHop), which shares ∼65% sequence similarity. One unique feature of dHop is the absence of DP1, a domain of poorly understood function found in hHop and Sti1p (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar), but there are sequence dissimilarities in other domains as well. In a variety of functional assays presented here, dHop is found to be similar to hHop and Sti1p, yet dHop fails to support steroid receptor maturation in vivo, and the deficiency maps to DP2. Antibodies—The following mouse monoclonal antibodies were used in these studies: F5 (28Smith D.F. Sullivan W.P. Marion T.N. Zaitsu K. Madden B. McCormick D.J. Toft D.O. Mol. Cell. Biol. 1993; 13: 869-876Crossref PubMed Scopus (247) Google Scholar), which recognizes vertebrate Hop; St2 (15Chang H.C. Nathan D.F. Lindquist S. Mol. Cell. Biol. 1997; 17: 318-325Crossref PubMed Scopus (193) Google Scholar), which is specific for yeast Sti1p; PR22 (29Sullivan W.P. Vroman B.T. Bauer V.J. Puri R.K. Riehl R.M. Pearson G.R. Toft D.O. Biochemistry. 1985; 24: 4214-4222Crossref PubMed Scopus (62) Google Scholar), which recognizes the progesterone receptor (PR); BuGR2 (Affinity Bioreagents, Golden, CO), which recognizes the glucocorticoid receptor (GR); and anti-L3, which reacts with a yeast ribosomal protein (30Vilardell J. Warner J.R. Mol. Cell. Biol. 1997; 17: 1959-1965Crossref PubMed Scopus (126) Google Scholar). Novel mouse monoclonal antibodies specific for dHop were prepared by standard procedures in the Mayo Clinic Scottsdale Immunology Core Facility. Mice were immunized with purified recombinant dHop, and hybridoma supernatants were initially screened by Western immunostaining for antibodies cross-reacting with dHop. Five monoclonal hybridoma cell lines producing IgG specific for dHop were established. The antibody selected for this study was termed DH1. Yeast Strains and Plasmids—All S. cerevisiae strains were generated in the W303a background (MATaleu2–112 ura3–1 trp1–1 his3–11, 15 ade2–1 can1–100 GAL SUC2). Plasmids were introduced into wild type or the previously described sti1Δ0 yeast strain (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar) using a lithium acetate/polyethylene glycol protocol. For each new strain construction, 2–4 independent transformants were compared for consistent Strains were in media consisting of yeast amino the and for the described previously D.L. Roberts P.J. J. Prapapanich V. Ratajczak T. R. D. Smith D.F. EMBO J. 2003; PubMed Scopus Google Scholar), the GR strain the which GR, and a in which is by Hop was into the which a a of and a D. R. M. PubMed Scopus Google Scholar). Yeast for assays were as described previously D.L. Roberts P.J. J. Prapapanich V. Ratajczak T. R. D. Smith D.F. EMBO J. 2003; PubMed Scopus Google Scholar). yeast strains were in media at to an at of was by for hormone addition to that the was in was to the at To assay for activity of was and to of the in at Five were at hormone addition and in a To the of were generated by the of the analysis of the of each a the of which is the of of Yeast strains were in of minimal at an of were and the cell was in of the addition of were by of between in a or of the were as determined by contrast The cell was in an during the at for the protein was as determined by protein assay and of Yeast monoclonal antibodies specific for hHop dHop and Sti1p were to protein of of For each yeast containing of protein was with of for at complexes were three in of in of and by For of yeast of of were by to a and for Sti1p, hHop, dHop, GR, or of Hop combining human and Drosophila Hop were by for the two were to the that the The were and used as in a with the to the The was into for in vitro or into a yeast In Hop were generated by in vitro in the presence of of each was by following protein were by For each of each were to of reticulocyte lysate; with an D.F. Toft D.O. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The was to the with of and for at complexes were by and complexes were in vitro as described previously and Smith (6Chen S. Smith D.F. J. Biol. Chem. 1998; 273: 35194-35200Abstract Full Text Full Text PDF PubMed Scopus (281) Google recombinant was from cell using the monoclonal antibody PR22 to protein assembly of of an of Hop or Hop in of containing an system. The assembly which was with the Hsp90 to of intermediate complexes containing Hop D.F. L. Chen S. Prapapanich V. Rimerman R.A. Mol. Cell. Biol. PubMed Scopus Google Scholar), was for at and the complexes were and then into were by and Yeast that Sti1p at but more at (7Nicolet C.M. Craig E.A. Mol. Cell. Biol. 1989; 9: 3638-3646Crossref PubMed Scopus (206) Google Scholar). We determined previously that hHop functionally for Sti1p as it growth at elevated temperature (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). To an of dHop we dHop can rescue the growth phenotype of the sti1Δ0 yeast shown in dHop and hHop both restore growth at in a sti1Δ0 it appears that dHop is fully functional in this growth functional analysis of dHop was using the yeast model for steroid receptor function. The M. Science. PubMed Scopus Google Scholar) that yeast a vertebrate steroid receptor the minimal to support of an The Lindquist (15Chang H.C. Nathan D.F. Lindquist S. Mol. Cell. Biol. 1997; 17: 318-325Crossref PubMed Scopus (193) Google Scholar) that Sti1p is for full receptor activity in the yeast and we (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar) that hHop fully rescue GR function in we by the ability of dHop to restore GR function In sti1Δ0 with activity remains the but activity is by In contrast to the human dHop was to restore GR function dHop can rescue the growth phenotype in Western that Sti1p is in sti1Δ0 that hHop and dHop are in yeast and that the of GR protein is similar in strains To gain into the of the defect that dHop support of GR we protein interactions of dHop. critical feature of assembly of receptor complexes is the binding of Hop to both Hsp70 and Hsp90. To dHop and hHop differently with Hsp70, Hsp90, or steroid receptor assays were in RL, a used for cell-free assembly of receptor complexes. in hHop and dHop with Hsp70 or Hsp90 hHop and dHop to in complexes The analysis was extended to yeast as shown in Sti1p, hHop, and dHop complexes were individually using specific monoclonal To for binding of chaperones to was at low which and at elevated which Hsp binding to Hop. using from yeast lacking the Hop Yeast Hsp90 by Western and Hsp70 and as determined by were with each of the three Hop hHop, and dHop complexes in were prepared from or sti1Δ0 with the and with antibodies specific for Sti1p, hHop, or dHop One of each was to to the complexes were and to The of was by a Western The Ssa was by to and but not other yeast Hsp70 To better dHop fails to support GR the amino acid for hHop, Sti1p, and dHop were to of the The most is the absence of in dHop. domains are although amino acid within these domains be an to the in hHop and dHop, a of constructs was generated in which domains were between The are by in and the are in of of a of hHop and dHop domains was as and proteins were generated in vitro and for with Hsp70 Hsp90 and In and Hop were functionally compared the sti1Δ0 GR strain with an for each Hop for the that generated background are in The from assays are in One of the is that of the dHop C-terminal DP2 domain to hHop is sufficient to GR Hsp binding and receptor assembly in vitro are is that the human domain into dHop disrupts Hsp binding. The hHop in which has been Hsp binding and GR of Hop to stimulate GR The sti1Δ0 GR strain was with an or the Hop form or of each were with and for of interactions and GR of from and GR activity from were as shown for hHop, dHop, and each of the constructs has unique of Hop function that to its role in GR maturation and function. The that dHop, similar to human and yeast can support yeast growth at elevated and yet dHop fails to support GR activity, a unique that Hop differently in these The molecular of Hop that are to growth have not been but multiple of Hsp70 and Hsp90 be by the of Hop and contribute to growth GR, must be by Hop in a dHop fails to support GR For Hop to support steroid receptor assembly and functional maturation it must bind both Hsp70 and Hsp90 (20Chen S. Prapapanich V. Rimerman R.A. Honore B. Smith D.F. Mol. Endocrinol. 1996; 10: 682-693Crossref PubMed Google Scholar). Hop does not directly with a receptor, it has been an adaptor protein that Hsp90 to complexes (6Chen S. Smith D.F. J. Biol. Chem. 1998; 273: 35194-35200Abstract Full Text Full Text PDF PubMed Scopus (281) Google Scholar). However, in of with in which binding to Hsp70 and Hsp90 remains of GR function is there must be an additional Hop activity other binding to Hsp70 and Hsp90 that is critical for GR function in additional activity maps most with DP2. We have shown that DP2 an independent domain that is by mutations in the conserved DP repeat motif G.M. H. Smith D.F. Cell Stress Chaperones. 2003; PubMed Scopus Google Scholar). of DP2 is of Hsp70 which evidence that DP2 and TPR1 in some to support Hsp70 binding The DP2 domain of dHop DP repeat and shares sequence similarity to hHop does the DP2 domain of Sti1p in this study is that DP2 between hHop and dHop does not affect Hsp70 with Drosophila DP2) fails to support GR activity, and the with human DP2) the ability to GR Therefore, DP2 other Hsp70 binding to GR function. is that Hop has a DP1, between TPR1 and TPR2a, and that is from dHop. does not to be critical for hHop function mutations in the DP repeat motif had on Hsp binding or GR activity (16Carrigan P.E. Nelson G.M. Roberts P.J. Stoffer J. Riggs D.L. Smith D.F. J. Biol. Chem. 2004; 279: 16185-16193Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar), and deletion of from hHop Hsp binding and GR On the other hand, of human into dHop binding to that the Drosophila protein has to with Hsp70 or Hsp90 when is understanding of Hop functional maturation of steroid receptors likely a better of domain-domain interactions within Hop and these interactions are by Hsp by the presence of a receptor to by additional Hsp co-chaperones that in the full assembly and by of Hop V.M. H.W. Blatch G.L. M. Biol. Chem. 2000; PubMed Scopus Google Scholar, V.M. M.S. Blatch G.L. J. Cell 2004; PubMed Scopus Google Scholar). in Hop structure contribute to the poorly transition of the receptor from intermediate complexes most associated with Hsp70 to functionally complexes most associated with Hsp90. We Roberts for D. O. Toft for antibody and J. R. Warner for the