Carta Acesso aberto Revisado por pares

Phosphotyrosine Confers Client Specificity to Hsp90

2010; Elsevier BV; Volume: 37; Issue: 3 Linguagem: Inglês

10.1016/j.molcel.2010.01.028

ISSN

1097-4164

Autores

Matthias P. Mayer,

Tópico(s)

Computational Drug Discovery Methods

Resumo

In this issue of Molecular Cell, Mollapour et al., 2010Mollapour M. Tsutsumi S. Donnelly A.C. Beebe K. Tokita M.J. Lee M.-J. Lee S. Morra G. Bourboulia D. Scroggins B.T. et al.Mol. Cell. 2010; 37 (this issue): 333-343Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar report a new tyrosine phosphorylation site in Hsp90, which is essential for Hsp90's interaction with a subset of its client proteins, notably protein kinases. In this issue of Molecular Cell, Mollapour et al., 2010Mollapour M. Tsutsumi S. Donnelly A.C. Beebe K. Tokita M.J. Lee M.-J. Lee S. Morra G. Bourboulia D. Scroggins B.T. et al.Mol. Cell. 2010; 37 (this issue): 333-343Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar report a new tyrosine phosphorylation site in Hsp90, which is essential for Hsp90's interaction with a subset of its client proteins, notably protein kinases. The molecular chaperone Hsp90, assisted by a number of cochaperones, forms dynamic complexes with some 200 client proteins, among them many signaling molecules controlling cell homeostasis, proliferation, differentiation, and cell death. Hsp90 clients are found in many pathways involved in malignant transformation, and activated variants of these clients are often much more dependent on Hsp90 than their wild-type counterparts. Since clients only become active after interaction with the Hsp90 machinery, it is not surprising that Hsp90 advanced to a prime target for anticancer therapy (Pearl et al., 2008Pearl L.H. Prodromou C. Workman P. Biochem. J. 2008; 410: 439-453Crossref PubMed Scopus (352) Google Scholar, Wandinger et al., 2008Wandinger S.K. Richter K. Buchner J. J. Biol. Chem. 2008; 283: 18473-18477Crossref PubMed Scopus (450) Google Scholar). Client activation by Hsp90 is believed to occur in all cases in a similar manner to that originally devised for steroid hormone receptors (Smith, 2000Smith D.F. Semin. Cell Dev. Biol. 2000; 11: 45-52Crossref PubMed Scopus (37) Google Scholar). Free steroid hormone receptor first interacts with the Hsp70 chaperone in an Hsp40-assisted process. Mediated by the adaptor protein Hop/Sti1, the early Hsp90-Hsp70-client complex assembles. Hsp70 and Hop/Sti1 are subsequently replaced by p23/Sba1 and immunophilins to yield the mature Hsp90-client complex. With a half-life of about 5 min, this complex decays and Hsp40 and Hsp70 may rebind the receptor. Steroid binding induces the exit from this cycle and leads to transcriptional activity of the hormone receptor. The cochaperone Cdc37 is required in addition for chaperoning protein kinases (Pearl, 2005Pearl L.H. Curr. Opin. Genet. Dev. 2005; 15: 55-61Crossref PubMed Scopus (182) Google Scholar). This activation cycle is coupled to the ATPase cycle of Hsp90, which is controlled by the cochaperones in the following way: ATP hydrolysis is inhibited by Hop/Sti1, p23/Sba1, and Cdc37 and stimulated by Aha1. Since ATP hydrolysis was shown to stimulate client release (Young and Hartl, 2000Young J.C. Hartl F.U. EMBO J. 2000; 19: 5930-5940Crossref PubMed Scopus (190) Google Scholar), Hop/Sti1 and Cdc37 are believed to favor client transfer from Hsp70 to Hsp90, and p23/Sba1 fosters the Hsp90-client complexes, while Aha1 accelerates complex dissociation. In addition, Hsp90 is regulated by posttranslational modifications including phosphorylation on serines, threonines, and tyrosines; acetylation of lysines; and S-nitrosylation of a single cysteine. Acetylation and S-nitrosylation lead to decreased activity of Hsp90 toward several different clients, suggesting a general deactivation (Retzlaff et al., 2009Retzlaff M. Stahl M. Eberl H.C. Lagleder S. Beck J. Kessler H. Buchner J. EMBO Rep. 2009; 10: 1147-1153Crossref PubMed Scopus (112) Google Scholar, Scroggins et al., 2007Scroggins B.T. Robzyk K. Wang D. Marcu M.G. Tsutsumi S. Beebe K. Cotter R.J. Felts S. Toft D. Karnitz L. et al.Mol. Cell. 2007; 25: 151-159Abstract Full Text Full Text PDF PubMed Scopus (312) Google Scholar). For phosphorylation the situation seems to be more complicated. Phosphorylation on two serine residues by casein kinase 2 seems to be necessary for activity, while hyperphosphorylation decreased the activity again (Miyata, 2009Miyata Y. Cell. Mol. Life Sci. 2009; 66: 1840-1849Crossref PubMed Scopus (84) Google Scholar). In addition, it was reported that a cycle of phosphorylation and dephosphorylation of Hsp90 is necessary for client binding and release (Zhao et al., 2001Zhao Y.G. Gilmore R. Leone G. Coffey M.C. Weber B. Lee P.W. J. Biol. Chem. 2001; 276: 32822-32827Crossref PubMed Scopus (88) Google Scholar). The question remains how global control of Hsp90, affecting its activity toward many clients in the same way, could be suitable for a chaperone with such a diverse clientele involved in signal transduction and regulatory circuits. In this issue of Molecular Cell, Neckers and coworkers (Mollapour et al., 2010Mollapour M. Tsutsumi S. Donnelly A.C. Beebe K. Tokita M.J. Lee M.-J. Lee S. Morra G. Bourboulia D. Scroggins B.T. et al.Mol. Cell. 2010; 37 (this issue): 333-343Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar) present an important addition to the complexity of the Hsp90 system. They identified a new tyrosine phosphorylation site in the N-terminal nucleotide-binding domain of Hsp90, the first for the yeast homolog Hsp82. The identified tyrosine is conserved in all eukaryotic Hsp90 proteins, and human Hsp90 can be phosphorylated at the same site. Swe1, the only true tyrosine-specific protein kinase in yeast and ortholog to mammalian Wee1, was found to phosphorylate the identified site in a cell-cycle-specific manner during S phase. Surprisingly, Mollapour et al., 2010Mollapour M. Tsutsumi S. Donnelly A.C. Beebe K. Tokita M.J. Lee M.-J. Lee S. Morra G. Bourboulia D. Scroggins B.T. et al.Mol. Cell. 2010; 37 (this issue): 333-343Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar found no sign of dephosphorylation. Instead, they show that Hsp90 is degraded in the yeast cytosol by the ubiquitin-proteasomal pathway and that tyrosine phosphorylation was a prerequisite for degradation. Thus, Hsp90 is phosphorylated in a cell-cycle-dependent manner, most likely in the nucleus, and subsequently degraded in the cytosol (Figure 1). What are the structural consequences of this modification? In the crystal structure of yeast Hsp82 in complex with AMPPNP and p23/Sba1, representing the ATP-bound prehydrolysis state, the identified tyrosine is at the interface of the dimerized N termini (Figure 2A ). In this position the tyrosine would not be accessible, and phosphorylation must happen in one of the open conformations known to exist for Hsp90 (Figure 2B). Furthermore, the close vicinity of the two tyrosines and the hydrophobic environment in the crystal structure would suggest that a phosphate group with its hydrophilic character and negative charges would disfavor such a conformation (Figure 2A, inset). Consequently, phosphotyrosine in this position should prevent N-terminal dimerization and reduce ATP hydrolysis, which in turn would affect client release (Young and Hartl, 2000Young J.C. Hartl F.U. EMBO J. 2000; 19: 5930-5940Crossref PubMed Scopus (190) Google Scholar). Neckers and coworkers show that replacement of this tyrosine by glutamate abrogates ATPase activity of Hsp90 and reduces N-terminal dimerization. Although it is not clear whether glutamate is a suitable surrogate for phosphotyrosine, these results are consistent with the structural analysis. What are the functional consequences? Phosphorylation at the identified tyrosine residue was not essential for viability of yeast but affected the interaction with certain cochaperones and a subset of client proteins. Physical interaction of Hop/Sti1 and Cdc37 with the nonphosphorylatable Hsp90 variant was not distinguishable from the interaction with the wild-type protein. In contrast, p23/Sba1 bound significantly less well and Aha1 did not bind at all to the phosphorylation minus variant. Most importantly, some clients, including several protein kinases and the heat shock factor, seem to depend on phosphorylation of the identified tyrosine, while glucocorticoid receptor (GR) and androgen receptor (AR) appear to be bound and activated independently of the tyrosine phosphorylation. This is the first demonstration that Hsp90 can be switched from an inactive to an active state for a subset of clients while remaining active for other clients. Therefore, the large number of Hsp90 molecules within a cell may constitute many different subpopulations specifically activated for a subset of clients in a timely and spatially regulated manner. Interestingly, the Hsp90 inhibitor geldanamycin bound more tightly to the nonphosphorylated form. Consequently, preventing phosphorylation should enhance the inhibitory effect of geldanamycin. In fact, Mollapour et al., 2010Mollapour M. Tsutsumi S. Donnelly A.C. Beebe K. Tokita M.J. Lee M.-J. Lee S. Morra G. Bourboulia D. Scroggins B.T. et al.Mol. Cell. 2010; 37 (this issue): 333-343Abstract Full Text Full Text PDF PubMed Scopus (129) Google Scholar show that in vivo knockdown of Wee1 or a Wee1 inhibitor potentiates the deleterious effect of geldanamycin on cell viability. Such a combination of drugs may widen the therapeutic window for Hsp90 inhibitors and reduce undesirable side effects. It remains to be shown how other Hsp90 inhibitors interact with the phosphorylated Hsp90 species. Swe1Wee1-Dependent Tyrosine Phosphorylation of Hsp90 Regulates Distinct Facets of Chaperone FunctionMollapour et al.Molecular CellFebruary 12, 2010In BriefSaccharomyces WEE1 (Swe1), the only “true” tyrosine kinase in budding yeast, is an Hsp90 client protein. Here we show that Swe1Wee1 phosphorylates a conserved tyrosine residue (Y24 in yeast Hsp90 and Y38 in human Hsp90α) in the N domain of Hsp90. Phosphorylation is cell-cycle associated and modulates the ability of Hsp90 to chaperone a selected clientele, including v-Src and several other kinases. Nonphosphorylatable mutants have normal ATPase activity, support yeast viability, and productively chaperone the Hsp90 client glucocorticoid receptor. Full-Text PDF Open Archive

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