Regulation of the Heat-shock Protein 70 Reaction Cycle by the Mammalian DnaJ Homolog, Hsp40

Minami, Yoshiko; Höhfeld, Jörg; Ohtsuka, Kenzo; Hartl, F. Ulrich

doi:10.1074/jbc.271.32.19617

Cited by 301 publications

(315 citation statements)

References 49 publications

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“…Several laboratories have found that the binding of unfolded proteins and peptides to Hsc70 is inhibited by DnaJ homologs (14 -16). On the other hand, studies on the refolding of several proteins by Hsc70 have strongly suggested that DnaJ homologs promote substrate binding to Hsc70 or at the very least are required for refolding to take place (8,17,18). Even in these studies, however, there is controversy as to whether ATP is always required for refolding as well as to whether Hsc70 and DnaJ alone are sufficient for refolding or other factors are also required.…”

mentioning

confidence: 83%

“…Much of the work on the mechanism of these partner proteins has been carried out with the Escherichia coli Hsc70 protein, DnaK, and its partner proteins, DnaJ and GrpE, which are required for numerous functions of DnaK. Thus far, GrpE homologs have only been found in eukaryotic mitochondria (8), but there are numerous eukaryotic DnaJ homologs that interact with Hsc70 (7,9,10). Some are membrane-bound, such as Sec63 in the endoplasmic reticulum and TiM44 in mitochondria, whereas others are soluble, occurring in the cytoplasm and in the lumens of the endoplasmic reticulum and mitochondria.…”

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confidence: 99%

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Interaction of Auxilin with the Molecular Chaperone, Hsc70

Jiang

Greener

Barouch

et al. 1997

Journal of Biological Chemistry

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We have studied the direct interaction of the constitutive isoform of Hsp70 (Hsc70) with the DnaJ homolog, auxilin, a cofactor that binds to clathrin-coated vesicles and is required for their uncoating by Hsc70. Auxilin caused a 5-fold increase in Hsc70 ATPase activity and a corresponding increase in steady-state levels of bound ADP; the dissociation constant for this effect was 0.6 M. Auxilin also induced polymerization of Hsc70 and bound to the resulting polymer at a 1:1 molar ratio; here too the dissociation constant was 0.6 M. Both this binding and polymerization required ATP; the Hsc70 depolymerized with a 4-min half-life when ATP was completely hydrolyzed to ADP. Although auxilin induces polymerization stoichiometrically and other DnaJ homologs induce polymerization catalytically, these data show that auxilin is similar to other DnaJ homologs in its ability to activate the Hsc70 ATPase activity, to polymerize Hsc70, and in the nucleotide dependence of this polymerization. Furthermore, the 70-amino acid J-domain of auxilin polymerized Hsc70 with the same nucleotide dependence as intact auxilin. Therefore, although only auxilin and not other DnaJ homologs support uncoating, our data suggest that various DnaJ homologs share a common mechanism of interaction with Hsc70, perhaps because their J-domains interact similarly with Hsc70.

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mentioning

confidence: 83%

mentioning

confidence: 99%

Interaction of Auxilin with the Molecular Chaperone, Hsc70

Jiang

Greener

Barouch

et al. 1997

Journal of Biological Chemistry

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“…The observations of Hernandez et al (21) with the PR suggest that YDJ-1 is bound directly to the receptor when it interacts as a co-chaperone with receptorbound hsp70. Inasmuch as hsp40/YDJ-1 promotes the ATPase activity of hsp70 (37) and hsp70 ATPase activity is required for both GR priming by hsp70 and the subsequent activation of the primed receptor complex by hsp90 (5,22), it is reasonable to propose that stimulation of hsp70 ATPase activity is the principal function of hsp40/YDJ-1 in opening of the steroid binding cleft. Although hsp40/YDJ-1 acts on the receptor-bound hsp70, it may also interact directly with the receptor.…”

Section: Fig 12mentioning

confidence: 99%

Visualization and Mechanism of Assembly of a Glucocorticoid Receptor·Hsp70 Complex That Is Primed for Subsequent Hsp90-dependent Opening of the Steroid Binding Cleft

Murphy

Morishima

Chen

et al. 2003

Journal of Biological Chemistry

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A minimal system of five proteins, hsp90, hsp70, Hop, hsp40, and p23, assembles glucocorticoid receptor (GR)⅐hsp90 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 GR⅐hsp70 complex that primes the receptor for subsequent ATPdependent 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 GR⅐hsp70 complexes by atomic force microscopy, and we find the most common stoichiometry to be 1:1, with some complexes of a size~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 GR⅐hsp90 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.Glucocorticoid receptors (GR) 1 are recovered from hormonefree cells as large multiprotein complexes containing a dimer of hsp90 (for review see Refs. 1 and 2). Hsp90 binds to the ligand binding domain (LBD) of the GR, and the steroid binding activity of the GR is absolutely hsp90-dependent (3, 4). When the GR is stripped of its associated hsp90, it immediately loses its ability to bind steroid, and steroid binding activity is regenerated when GR⅐hsp90 heterocomplexes are reformed by the hsp90/hsp70-based chaperone machinery (5, 6). The steroids bind deep in a hydrophobic cleft that appears to be collapsed in the absence of ligand, such that the LBD must change its conformation to allow entry of ligand (7). The hsp90/hsp70-dependent chaperone machinery carries out the ATP-dependent opening of the binding cleft in the GR LBD such that it can be accessed by steroid. In addition to opening the steroid binding cleft, formation of a complex with hsp90 is accompanied by conformational changes that increase the sensitivity of the GR LBD to attack by thiol-derivatizing agents and trypsin (8 -10).The multiprotein chaperone machinery that forms receptor⅐hsp90 heterocomplexes was originally identified in reticulocyte lysate (11,12). This machinery has been reconstituted (13), and a mixture of five purified proteins (hsp90, hsp70, 2 Hop, hsp40, and p23) is now used to achieve efficient receptor⅐hsp90 heterocomplex assembly (14, 15). The chaperones hsp90 and hsp70 are both essential for opening the steroid binding cleft in the GR LBD, and hsp40, Hop (hsp70/hsp90 organizing protein), and p23 act as co-chaperones to increase the rate or extent of GR⅐hsp90 heterocomplex assembly (16). Hop b...

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“…A polypeptide substrate commonly used to study Hsc70 function is firefly luciferase, which has a sensitive and reproducible enzymatic assay. Purified Hsc70 has been shown to assist luciferase refolding in numerous studies (Hohfeld et al, 1995;Minami et al, 1996;Hohfeld and Jentsch, 1997;Terada et al, 1997;Luders et al, 2000;Terada and Mori, 2000). However, in most cases the stress-inducible cochaperone Hsp40 was used, and a direct comparison between all three DJAs with Hsc70 has not yet been reported.…”

Section: Dja Activation Of Hsc70mentioning

confidence: 99%

“…Inhibition of Hsc70 binding should then impair the Tom70 docking step required for import. Alternatively, Hsp40 can activate substrate binding by Hsc70 without strongly interacting with substrate (Minami et al, 1996;Cintron and Toft, 2006). If Hsp40 or some mechanism other than the DJAs was sufficient for Hsc70 binding, the truncated DJAs should have less effect.…”

Section: Dja Interaction With Preproteinmentioning

confidence: 99%

Multiple 40-kDa Heat-Shock Protein Chaperones Function in Tom70-dependent Mitochondrial Import

Bhangoo

Tzankov

Fan

et al. 2007

MBoC

108

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Mitochondrial preproteins that are imported via the translocase of the mitochondrial outer membrane (Tom)70 receptor are complexed with cytosolic chaperones before targeting to the mitochondrial outer membrane. The adenine nucleotide transporter (ANT) follows this pathway, and its purified mature form is identical to the preprotein. Purified ANT was reconstituted with chaperones in reticulocyte lysate, and bound proteins were identified by mass spectrometry. In addition to 70-kDa heat-shock cognate protein (Hsc70) and 90-kDa heat-shock protein (Hsp90), a specific subset of cochaperones were found, but no mitochondria-specific targeting factors were found. Interestingly, three different Hsp40-related J-domain proteins were identified: DJA1, DJA2, and DJA4. The DJAs bound preproteins to different extents through their C-terminal regions. DJA dominant-negative mutants lacking the N-terminal J-domains impaired mitochondrial import. The mutants blocked the binding of Hsc70 to preprotein, but with varying efficiency. The DJAs also showed significant differences in activation of the Hsc70 ATPase and Hsc70-dependent protein refolding. In HeLa cells, the DJAs increased new protein folding and mitochondrial import, although to different extents. No single DJA was superior to the others in all aspects, but each had a profile of partial specialization. The Hsp90 cochaperones p23 and Aha1 also regulated Hsp90 -preprotein interactions. We suggest that multiple cochaperones with similar yet partially specialized properties cooperate in optimal chaperone-preprotein complexes.

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Regulation of the Heat-shock Protein 70 Reaction Cycle by the Mammalian DnaJ Homolog, Hsp40

Cited by 301 publications

References 49 publications

Interaction of Auxilin with the Molecular Chaperone, Hsc70

Interaction of Auxilin with the Molecular Chaperone, Hsc70

Visualization and Mechanism of Assembly of a Glucocorticoid Receptor·Hsp70 Complex That Is Primed for Subsequent Hsp90-dependent Opening of the Steroid Binding Cleft

Multiple 40-kDa Heat-Shock Protein Chaperones Function in Tom70-dependent Mitochondrial Import

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