HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 283, Issue 23, 15732-15739, June 6, 2008

This Article Full Text Full Text (PDF) All Versions of this Article: 283/23/15732    most recent M800728200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Savistchenko, J. Articles by Melki, R. Search for Related Content PubMed PubMed Citation Articles by Savistchenko, J. Articles by Melki, R. Social Bookmarking                      What's this?

Molecular Chaperones and the Assembly of the Prion Ure2p in Vitro^*

From the Laboratoire d'Enzymologie et Biochimie Structurales, CNRS, 91198 Gif-sur-Yvette Cedex, France

The protein Ure2 from Saccharomyces cerevisiae possesses prion properties at the origin of the [URE3] trait. In vivo, a high molecular weight form of inactive Ure2p is associated to [URE3]. The faithful and continued propagation of [URE3]is dependent on the expression levels of molecular chaperones from the Hsp100, -70, and -40 families; however, so far, their role is not fully documented. Here we investigate the effects of molecular chaperones from the Hsp40, Hsp70, Hsp90, and Hsp100 families and the chaperonin CCT/Tric on the assembly of full-length Ure2p. We show that Hsp104p greatly stimulates Ure2p aggregation, whereas Ssa1p, Ydj1p, Sis1p, and Hsp82p inhibit aggregation to different extents. The nature of the high molecular weight Ure2p species that forms in the presence of the different molecular chaperones and their nucleotide dependence is described. We show that Hsp104p favors the aggregation of Ure2p into non-fibrillar high molecular weight particles, whereas Ssa1p, Ydj1p, Sis1p, and Hsp82p sequester Ure2p in spherical oligomers. Using fluorescently labeled full-length Ure2p and Ure2p-(94–354) and fluorescence polarization, we show that Ssa1p binding to Ure2p is ATP-dependent, whereas that of Hsp104p is not. We also show that Ssa1p preferentially interacts with the N-terminal domain of Ure2p that is critical for prion propagation, whereas Ydj1p preferentially interacts with the C-terminal domain of the protein, and we discuss the significance of this observation. Finally, the affinities of Ssa1p, Ydj1p, and Hsp104p for Ure2p are determined. Our in vitro observations bring new insight into the mechanism by which molecular chaperones influence the propagation of [URE3].

Received for publication, January 28, 2008 , and in revised form, March 24, 2008.

^* This work was supported by the French Ministry of Education, Research and Technology through the Groupement d'Intérêt Scientifique Prion, the CNRS, the Fondation pour la Recherche Médicale, and the ANR. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement"in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Present address: Lilly UK, Erl Wood Manor, Windlesham, Surrey, GU20 6PH, UK.

² To whom correspondence should be addressed. Tel.: 33-169823503; Fax: 33-169823129; E-mail: melki{at}lebs.cnrs-gif.fr.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				P. Kota, D. W. Summers, H.-Y. Ren, D. M. Cyr, and N. V. Dokholyan Identification of a consensus motif in substrates bound by a Type I Hsp40 PNAS, July 7, 2009; 106(27): 11073 - 11078. [Abstract] [Full Text] [PDF]

				D. Sharma, R. F. Stanley, and D. C. Masison Curing of Yeast [URE3] Prion by the Hsp40 Cochaperone Ydj1p Is Mediated by Hsp70 Genetics, January 1, 2009; 181(1): 129 - 137. [Abstract] [Full Text] [PDF]