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Chrisostomos Prodromou - One of the best experts on this subject based on the ideXlab platform.
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Chaperone ligand-discrimination by the TPR-Domain Protein Tah1
Biochemical Journal, 2008Co-Authors: Stefan H Millson, Cara K. Vaughan, Chao Zhai, Maruf M. U. Ali, Barry Panaretou, Peter W. Piper, Laurence H. Pearl, Chrisostomos ProdromouAbstract:Tah1 has been identified as a tetratricopeptide (TPR)-domain protein. TPR-domain proteins are involved in protein-protein interactions and a number have been characterized that interact either with Hsp70 or Hsp90, but a few can bind both chaperones. Independent studies suggest that Tah1 interacts with Hsp90, but whether it can also interact with Hsp70/Ssa1 has not been investigated. Amino acid sequence alignments suggest that Tah1 is most similar to the TPR2b-domain of Hop which when mutated reduces binding to both Hsp90 and Hsp70. Our alignments suggest that there are three TPR-domain motifs in Tah1 which is consistent with the architecture of the TPR2b-domain. We find that Tah1 is specific for Hsp90, able to bind tightly the yeast Hsp90, and the human Hsp90α and Hsp90β proteins, but not the yeast Hsp70, Ssa1 isoform. Tah1 acheives ligand discrimination by favourably binding the methionine residue in the conserved MEEVD-motif (Hsp90) and positively discriminating against the first valine residue in the VEEVD-motif (Ssa1). We also show that Tah1 can affect the ATPase activity of Hsp90, in common with some other TPR-domain proteins.
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Chaperone ligand-discrimination by the TPR-domain protein Tah1
The Biochemical journal, 2008Co-Authors: Stefan H Millson, Cara K. Vaughan, Chao Zhai, Maruf M. U. Ali, Barry Panaretou, Peter W. Piper, Laurence H. Pearl, Chrisostomos ProdromouAbstract:Tah1 [TPR (tetratricopeptide repeat)-containing protein associated with Hsp (heat-shock protein) 90] has been identified as a TPR-domain protein. TPR-domain proteins are involved in protein–protein interactions and a number have been characterized that interact either with Hsp70 or Hsp90, but a few can bind both chaperones. Independent studies suggest that Tah1 interacts with Hsp90, but whether it can also interact with Hsp70/Ssa1 has not been investigated. Amino-acid-sequence alignments suggest that Tah1 is most similar to the TPR2b domain of Hop (Hsp-organizing protein) which when mutated reduces binding to both Hsp90 and Hsp70. Our alignments suggest that there are three TPR-domain motifs in Tah1, which is consistent with the architecture of the TPR2b domain. In the present study we find that Tah1 is specific for Hsp90, and is able to bind tightly the yeast Hsp90, and the human Hsp90α and Hsp90β proteins, but not the yeast Hsp70 Ssa1 isoform. Tah1 acheives ligand discrimination by favourably binding the methionine residue in the conserved MEEVD motif (Hsp90) and positively discriminating against the first valine residue in the VEEVD motif (Ssa1). In the present study we also show that Tah1 can affect the ATPase activity of Hsp90, in common with some other TPR-domain proteins.
William B Pratt - One of the best experts on this subject based on the ideXlab platform.
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visualization and mechanism of assembly of a glucocorticoid receptor hsp70 complex that is primed for subsequent Hsp90 dependent opening of the steroid binding cleft
Journal of Biological Chemistry, 2003Co-Authors: Patrick J. Murphy, Yoshihiro Morishima, Mario D. Galigniana, Haifeng Chen, John F Mansfield, Stoney S Simons, William B PrattAbstract:A minimal system of five proteins, Hsp90, hsp70, Hop, hsp40, and p23, assembles glucocorticoid receptor (GR).Hsp90 heterocomplexes and causes the simultaneous opening of the steroid binding cleft to access by steroid. The first step in assembly is the ATP-dependent and hsp40 (YDJ-1)-dependent formation of a GR.hsp70 complex that primes the receptor for subsequent ATP-dependent activation by Hsp90, Hop, and p23. This study focuses on three aspects of the GR priming reaction with hsp70. First, we have visualized the primed GR.hsp70 complexes by atomic force microscopy, and we find the most common stoichiometry to be 1:1, with some complexes of a size approximately 1:2 and a few complexes of larger size. Second, in a recent study of progesterone receptor priming, it was shown that hsp40 binds first, leading to the notion that it targets hsp70 to the receptor. We show here that hsp40 does not perform such a targeting function in priming the GR. Third, we focus on a short amino-terminal segment of the ligand binding domain that is required for GR.Hsp90 heterocomplex assembly. By using two glutathione S-transferase (GST)/ligand binding domain fusions with (GST/520C) and without (GST/554C) Hsp90 binding and steroid binding activity, we show that the priming step with hsp70 occurs with GST/554C, and it is the subsequent assembly step with Hsp90 that is defective.
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regulation of signaling protein function and trafficking by the Hsp90 hsp70 based chaperone machinery
Experimental Biology and Medicine, 2003Co-Authors: William B Pratt, David O ToftAbstract:Nearly 100 proteins are known to be regulated by Hsp90. Most of these substrates or "client proteins" are involved in signal transduction, and they are brought into complex with Hsp90 by a multiprotein Hsp90/hsp70-based chaperone machinery. In addition to binding substrate proteins at the chaperone site(s), Hsp90 binds cofactors at other sites that are part of the heterocomplex assembly machinery as well as immunophilins that connect assembled substrate*Hsp90 complexes to protein-trafficking systems. In the 5 years since we last reviewed this subject, much has been learned about Hsp90 structure, nucleotide-binding, and cochaperone interactions; the most important concept is that ATP hydrolysis by an intrinsic ATPase activity results in a conformational change in Hsp90 that is required to induce conformational change in a substrate protein. The conformational change induced in steroid receptors is an opening of the steroid-binding cleft so that it can be accessed by steroid. We have now developed a minimal system of five purified proteins-Hsp90, hsp70, Hop, hsp40, and p23- that assembles stable receptor*Hsp90 heterocomplexes. An Hsp90*Hop*hsp70*hsp40 complex opens the cleft in an ATP-dependent process to produce a receptor*Hsp90 heterocomplex with Hsp90 in its ATP-bound conformation, and p23 then interacts with the Hsp90 to stabilize the complex. Stepwise assembly experiments have shown that hsp70 and hsp40 first interact with the receptor in an ATP-dependent reaction to produce a receptor*hsp70*hsp40 complex that is "primed" to be activated to the steroid-binding state in a second ATP-dependent step with Hsp90, Hop, and p23. Successful use of the five-protein system with other substrates indicates that it can assemble signal protein*Hsp90 heterocomplexes whether the substrate is a receptor, a protein kinase, or a transcription factor. This purified system should facilitate understanding of how eukaryotic hsp70 and Hsp90 work together as essential components of a process that alters the conformations of substrate proteins to states that respond in signal transduction.
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stoichiometry abundance and functional significance of the Hsp90 hsp70 based multiprotein chaperone machinery in reticulocyte lysate
Journal of Biological Chemistry, 2001Co-Authors: Patrick J. Murphy, Yoshihiro Morishima, Kimon C. Kanelakis, Mario D. Galigniana, William B PrattAbstract:Rabbit reticulocyte lysate contains a multiprotein chaperone system that assembles the glucocorticoid receptor (GR) into a complex with Hsp90 and converts the hormone binding domain of the receptor to its high affinity steroid binding state. This system has been resolved into five proteins, with Hsp90 and hsp70 being essential and Hop, hsp40, and p23 acting as co-chaperones that optimize assembly. Hop binds independently to hsp70 and Hsp90 to form an Hsp90.Hop.hsp70 complex that acts as a machinery to open up the GR steroid binding site. Because purified Hsp90 and hsp70 are sufficient for some activation of GR steroid binding activity, some investigators have rejected any role for Hop in GR.Hsp90 heterocomplex assembly. Here, we counter that impression by showing that all of the Hop in reticulocyte lysate is present in an Hsp90.Hop.hsp70 complex with a stoichiometry of 2:1:1. The complex accounts for approximately 30% of the Hsp90 and approximately 9% of the hsp70 in lysate, and upon Sephacryl S-300 chromatography the GR.Hsp90 assembly activity resides in the peak containing Hop-bound Hsp90. Consistent with the notion that the two essential chaperones cooperate with each other to open up the steroid binding site, we also show that purified Hsp90 and hsp70 interact directly with each other to form weak Hsp90.hsp70 complexes with a stoichiometry of 2:1.
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Stoichiometry, abundance, and functional significance of the Hsp90/hsp70-based multiprotein chaperone machinery in reticulocyte lysate.
The Journal of biological chemistry, 2001Co-Authors: Patrick J. Murphy, Yoshihiro Morishima, Kimon C. Kanelakis, Mario D. Galigniana, William B PrattAbstract:Rabbit reticulocyte lysate contains a multiprotein chaperone system that assembles the glucocorticoid receptor (GR) into a complex with Hsp90 and converts the hormone binding domain of the receptor to its high affinity steroid binding state. This system has been resolved into five proteins, with Hsp90 and hsp70 being essential and Hop, hsp40, and p23 acting as co-chaperones that optimize assembly. Hop binds independently to hsp70 and Hsp90 to form an Hsp90.Hop.hsp70 complex that acts as a machinery to open up the GR steroid binding site. Because purified Hsp90 and hsp70 are sufficient for some activation of GR steroid binding activity, some investigators have rejected any role for Hop in GR.Hsp90 heterocomplex assembly. Here, we counter that impression by showing that all of the Hop in reticulocyte lysate is present in an Hsp90.Hop.hsp70 complex with a stoichiometry of 2:1:1. The complex accounts for approximately 30% of the Hsp90 and approximately 9% of the hsp70 in lysate, and upon Sephacryl S-300 chromatography the GR.Hsp90 assembly activity resides in the peak containing Hop-bound Hsp90. Consistent with the notion that the two essential chaperones cooperate with each other to open up the steroid binding site, we also show that purified Hsp90 and hsp70 interact directly with each other to form weak Hsp90.hsp70 complexes with a stoichiometry of 2:1.
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stepwise assembly of a glucocorticoid receptor Hsp90 heterocomplex resolves two sequential atp dependent events involving first hsp70 and then Hsp90 in opening of the steroid binding pocket
Journal of Biological Chemistry, 2000Co-Authors: Yoshihiro Morishima, Patrick J. Murphy, Edwin R Sanchez, William B PrattAbstract:Abstract A system of five purified proteins that assembles stable glucocorticoid receptor (GR)-Hsp90 heterocomplexes has been reconstituted from reticulocyte lysate. Two proteins, Hsp90 and hsp70, are required for the activation of steroid binding activity that occurs with heterocomplex assembly, and three proteins, Hop, hsp40, p23, act as co-chaperones that enhance activation and assembly (Morishima, Y., Kanelakis, K. C., Silverstein, A. M., Dittmar, K. D., Estrada, L., and Pratt, W. B. (2000) J. Biol. Chem. 275, 6894–6900). Here we demonstrate that the first step in assembly is the ATP-dependent and hsp40 (YDJ-1)-dependent binding of hsp70 to the GR. After elimination of free hsp70, these preformed GR·hsp70 complexes can be activated to the steroid binding state by the hsp70 free assembly system in a second ATP-dependent step. Hsp90 is required for opening of the steroid binding pocket and is converted to its ATP-dependent conformation during this second step. We predict that hsp70 in its ATP-dependent conformation binds initially to the folded receptor and is then converted to the ADP-dependent form with high affinity for hydrophobic substrate. This conversion initiates the opening of the hydrophobic steroid binding pocket such that it can now accept the hydrophobic binding form of Hsp90, which in turn must be converted to its ATP-dependent conformation for the pocket to be accessible by steroid.
Stefan H Millson - One of the best experts on this subject based on the ideXlab platform.
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Chaperone ligand-discrimination by the TPR-Domain Protein Tah1
Biochemical Journal, 2008Co-Authors: Stefan H Millson, Cara K. Vaughan, Chao Zhai, Maruf M. U. Ali, Barry Panaretou, Peter W. Piper, Laurence H. Pearl, Chrisostomos ProdromouAbstract:Tah1 has been identified as a tetratricopeptide (TPR)-domain protein. TPR-domain proteins are involved in protein-protein interactions and a number have been characterized that interact either with Hsp70 or Hsp90, but a few can bind both chaperones. Independent studies suggest that Tah1 interacts with Hsp90, but whether it can also interact with Hsp70/Ssa1 has not been investigated. Amino acid sequence alignments suggest that Tah1 is most similar to the TPR2b-domain of Hop which when mutated reduces binding to both Hsp90 and Hsp70. Our alignments suggest that there are three TPR-domain motifs in Tah1 which is consistent with the architecture of the TPR2b-domain. We find that Tah1 is specific for Hsp90, able to bind tightly the yeast Hsp90, and the human Hsp90α and Hsp90β proteins, but not the yeast Hsp70, Ssa1 isoform. Tah1 acheives ligand discrimination by favourably binding the methionine residue in the conserved MEEVD-motif (Hsp90) and positively discriminating against the first valine residue in the VEEVD-motif (Ssa1). We also show that Tah1 can affect the ATPase activity of Hsp90, in common with some other TPR-domain proteins.
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Chaperone ligand-discrimination by the TPR-domain protein Tah1
The Biochemical journal, 2008Co-Authors: Stefan H Millson, Cara K. Vaughan, Chao Zhai, Maruf M. U. Ali, Barry Panaretou, Peter W. Piper, Laurence H. Pearl, Chrisostomos ProdromouAbstract:Tah1 [TPR (tetratricopeptide repeat)-containing protein associated with Hsp (heat-shock protein) 90] has been identified as a TPR-domain protein. TPR-domain proteins are involved in protein–protein interactions and a number have been characterized that interact either with Hsp70 or Hsp90, but a few can bind both chaperones. Independent studies suggest that Tah1 interacts with Hsp90, but whether it can also interact with Hsp70/Ssa1 has not been investigated. Amino-acid-sequence alignments suggest that Tah1 is most similar to the TPR2b domain of Hop (Hsp-organizing protein) which when mutated reduces binding to both Hsp90 and Hsp70. Our alignments suggest that there are three TPR-domain motifs in Tah1, which is consistent with the architecture of the TPR2b domain. In the present study we find that Tah1 is specific for Hsp90, and is able to bind tightly the yeast Hsp90, and the human Hsp90α and Hsp90β proteins, but not the yeast Hsp70 Ssa1 isoform. Tah1 acheives ligand discrimination by favourably binding the methionine residue in the conserved MEEVD motif (Hsp90) and positively discriminating against the first valine residue in the VEEVD motif (Ssa1). In the present study we also show that Tah1 can affect the ATPase activity of Hsp90, in common with some other TPR-domain proteins.
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expressed as the sole Hsp90 of yeast the α and β isoforms of human Hsp90 differ with regard to their capacities for activation of certain client proteins whereas only Hsp90β generates sensitivity to the Hsp90 inhibitor radicicol
FEBS Journal, 2007Co-Authors: Stefan H Millson, Barry Panaretou, Andrew W Truman, Attila Racz, James M Nuttall, Mehdi Mollapour, Csaba Soti, Peter W. PiperAbstract:Heat shock protein 90 (Hsp90) is a molecular chaperone required for the activity of many of the most important regulatory proteins of eukaryotic cells (the Hsp90 ‘clients’). Vertebrates have two isoforms of cytosolic Hsp90, Hsp90α and Hsp90β. Hsp90β is expressed constitutively to a high level in most tissues and is generally more abundant than Hsp90α, whereas Hsp90α is stress-inducible and overexpressed in many cancerous cells. Expressed as the sole Hsp90 of yeast, human Hsp90α and Hsp90β are both able to provide essential Hsp90 functions. Activations of certain Hsp90 clients (heat shock transcription factor, v-src) were more efficient with Hsp90α, rather than Hsp90β, present in the yeast. In contrast, activation of certain other clients (glucocorticoid receptor; extracellular signal-regulated kinase-5 mitogen-activated protein kinase) was less affected by the human Hsp90 isoform present in these cells. Remarkably, whereas expression of Hsp90β as the sole Hsp90 of yeast rendered cells highly sensitive to the Hsp90 inhibitor radicicol, comparable expression of Hsp90α did not. This raises the distinct possibility that, also for mammalian systems, alterations to the Hsp90α/Hsp90β ratio (as with heat shock) might be a significant factor affecting cellular susceptibility to Hsp90 inhibitors.
Masayoshi Kanisawa - One of the best experts on this subject based on the ideXlab platform.
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Expression of heat shock proteins in squamous cell carcinoma of the tongue: an immunohistochemical study
Journal of Oral Pathology & Medicine, 2007Co-Authors: Ryoichi Kawabe, Masamichi Hara, Yukiko Kurasono, Hitoshi Kitamura, Kiyohide Fujita, Masayoshi KanisawaAbstract:Ito T, Kawabe R, Kurasono Y, Hara M, Kitamura H, Fujita K, Kanisawa M: Expression of heat shock proteins in squamous cell carcinoma of the tongue: an immunohistochemical study. J Oral Pathol Med 1998; 27: 18–22. © Munksgaard, 1998. Twenty-four specimens of squamous cell carcinoma of the tongue were immuno-stained for heat shock proteins (HSPs) to reveal differences in stainability among normal epithelium, dysplasia and carcinoma and to clarify the prognostic significance of HSPs in comparison with survival period, clinical stage, lymph node metastasis, histological grade, and p53 immunostaining. Normal epithelium was positively stained in the suprabasal layer for HSP60 and HSP70, but was negative for HSP27 and Hsp90. Dysplastic lesions were positive for HSP27, HSP70 and Hsp90, but stained variously for HSP60. In squamous cell carcinoma, the cytoplasm of suprabasal tumor cells was often positive for HSP27 and Hsp90 (18/24, 17/24, respectively). Although HSP immunohistochemistry has revealed changes in HSP expression during tumorigenesis of squamous epithelium of the tongue, there was no correlation between HSP staining and survival period, stage, lymph node metastasis, histological grade or p53 immunostaining.
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Expression of heat shock proteins in squamous cell carcinoma of the tongue: an immunohistochemical study
Journal of Oral Pathology & Medicine, 2007Co-Authors: Ryoichi Kawabe, Masamichi Hara, Yukiko Kurasono, Hitoshi Kitamura, Kiyohide Fujita, Masayoshi KanisawaAbstract:Ito T, Kawabe R, Kurasono Y, Hara M, Kitamura H, Fujita K, Kanisawa M: Expression of heat shock proteins in squamous cell carcinoma of the tongue: an immunohistochemical study. J Oral Pathol Med 1998; 27: 18–22. © Munksgaard, 1998. Twenty-four specimens of squamous cell carcinoma of the tongue were immuno-stained for heat shock proteins (HSPs) to reveal differences in stainability among normal epithelium, dysplasia and carcinoma and to clarify the prognostic significance of HSPs in comparison with survival period, clinical stage, lymph node metastasis, histological grade, and p53 immunostaining. Normal epithelium was positively stained in the suprabasal layer for HSP60 and HSP70, but was negative for HSP27 and Hsp90. Dysplastic lesions were positive for HSP27, HSP70 and Hsp90, but stained variously for HSP60. In squamous cell carcinoma, the cytoplasm of suprabasal tumor cells was often positive for HSP27 and Hsp90 (18/24, 17/24, respectively). Although HSP immunohistochemistry has revealed changes in HSP expression during tumorigenesis of squamous epithelium of the tongue, there was no correlation between HSP staining and survival period, stage, lymph node metastasis, histological grade or p53 immunostaining.
Peter W. Piper - One of the best experts on this subject based on the ideXlab platform.
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Chaperone ligand-discrimination by the TPR-Domain Protein Tah1
Biochemical Journal, 2008Co-Authors: Stefan H Millson, Cara K. Vaughan, Chao Zhai, Maruf M. U. Ali, Barry Panaretou, Peter W. Piper, Laurence H. Pearl, Chrisostomos ProdromouAbstract:Tah1 has been identified as a tetratricopeptide (TPR)-domain protein. TPR-domain proteins are involved in protein-protein interactions and a number have been characterized that interact either with Hsp70 or Hsp90, but a few can bind both chaperones. Independent studies suggest that Tah1 interacts with Hsp90, but whether it can also interact with Hsp70/Ssa1 has not been investigated. Amino acid sequence alignments suggest that Tah1 is most similar to the TPR2b-domain of Hop which when mutated reduces binding to both Hsp90 and Hsp70. Our alignments suggest that there are three TPR-domain motifs in Tah1 which is consistent with the architecture of the TPR2b-domain. We find that Tah1 is specific for Hsp90, able to bind tightly the yeast Hsp90, and the human Hsp90α and Hsp90β proteins, but not the yeast Hsp70, Ssa1 isoform. Tah1 acheives ligand discrimination by favourably binding the methionine residue in the conserved MEEVD-motif (Hsp90) and positively discriminating against the first valine residue in the VEEVD-motif (Ssa1). We also show that Tah1 can affect the ATPase activity of Hsp90, in common with some other TPR-domain proteins.
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Chaperone ligand-discrimination by the TPR-domain protein Tah1
The Biochemical journal, 2008Co-Authors: Stefan H Millson, Cara K. Vaughan, Chao Zhai, Maruf M. U. Ali, Barry Panaretou, Peter W. Piper, Laurence H. Pearl, Chrisostomos ProdromouAbstract:Tah1 [TPR (tetratricopeptide repeat)-containing protein associated with Hsp (heat-shock protein) 90] has been identified as a TPR-domain protein. TPR-domain proteins are involved in protein–protein interactions and a number have been characterized that interact either with Hsp70 or Hsp90, but a few can bind both chaperones. Independent studies suggest that Tah1 interacts with Hsp90, but whether it can also interact with Hsp70/Ssa1 has not been investigated. Amino-acid-sequence alignments suggest that Tah1 is most similar to the TPR2b domain of Hop (Hsp-organizing protein) which when mutated reduces binding to both Hsp90 and Hsp70. Our alignments suggest that there are three TPR-domain motifs in Tah1, which is consistent with the architecture of the TPR2b domain. In the present study we find that Tah1 is specific for Hsp90, and is able to bind tightly the yeast Hsp90, and the human Hsp90α and Hsp90β proteins, but not the yeast Hsp70 Ssa1 isoform. Tah1 acheives ligand discrimination by favourably binding the methionine residue in the conserved MEEVD motif (Hsp90) and positively discriminating against the first valine residue in the VEEVD motif (Ssa1). In the present study we also show that Tah1 can affect the ATPase activity of Hsp90, in common with some other TPR-domain proteins.
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expressed as the sole Hsp90 of yeast the α and β isoforms of human Hsp90 differ with regard to their capacities for activation of certain client proteins whereas only Hsp90β generates sensitivity to the Hsp90 inhibitor radicicol
FEBS Journal, 2007Co-Authors: Stefan H Millson, Barry Panaretou, Andrew W Truman, Attila Racz, James M Nuttall, Mehdi Mollapour, Csaba Soti, Peter W. PiperAbstract:Heat shock protein 90 (Hsp90) is a molecular chaperone required for the activity of many of the most important regulatory proteins of eukaryotic cells (the Hsp90 ‘clients’). Vertebrates have two isoforms of cytosolic Hsp90, Hsp90α and Hsp90β. Hsp90β is expressed constitutively to a high level in most tissues and is generally more abundant than Hsp90α, whereas Hsp90α is stress-inducible and overexpressed in many cancerous cells. Expressed as the sole Hsp90 of yeast, human Hsp90α and Hsp90β are both able to provide essential Hsp90 functions. Activations of certain Hsp90 clients (heat shock transcription factor, v-src) were more efficient with Hsp90α, rather than Hsp90β, present in the yeast. In contrast, activation of certain other clients (glucocorticoid receptor; extracellular signal-regulated kinase-5 mitogen-activated protein kinase) was less affected by the human Hsp90 isoform present in these cells. Remarkably, whereas expression of Hsp90β as the sole Hsp90 of yeast rendered cells highly sensitive to the Hsp90 inhibitor radicicol, comparable expression of Hsp90α did not. This raises the distinct possibility that, also for mammalian systems, alterations to the Hsp90α/Hsp90β ratio (as with heat shock) might be a significant factor affecting cellular susceptibility to Hsp90 inhibitors.