Prion generation in vitro:: amyloid of Ure2p is infectious

被引:188
作者
Brachmann, A
Baxa, U
Wickner, RB
机构
[1] NIDDKD, Lab Biochem & Genet, NIH, Bethesda, MD 20892 USA
[2] NIAMSD, Struct Biol Lab, NIH, Bethesda, MD 20892 USA
关键词
amyloid; prion transformation; prion variants; URE3;
D O I
10.1038/sj.emboj.7600772
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
[URE3] is a prion ( infectious protein) of the Ure2 protein of yeast. In vitro, Ure2p can form amyloid filaments, but direct evidence that these filaments constitute the infectious form is still missing. Here we demonstrate that recombinant Ure2p converted into amyloid can infect yeast cells lacking the prion. Infection produced a variety of [ URE3] variants. Extracts of [ URE3] strains, as well as amyloid of Ure2p formed in an extract-primed reaction could transmit to uninfected cells the [ URE3] variant present in the cells from which the extracts were made. Infectivity and determinant of [ URE3] variants resided within the N-terminal 65 amino acids of Ure2p. Notably, we could show that amyloid filaments of recombinant Ure2p are nearly as infectious per mass of Ure2p as extracts of [ URE3] strains. Sizing experiments indicated that infectious particles in vitro and in vivo were >20 nm in diameter, suggesting that they were amyloid filaments and not smaller oligomeric structures. Our data indicate that there is no substantial difference between filaments formed in vivo and in vitro.
引用
收藏
页码:3082 / 3092
页数:11
相关论文
共 46 条
[1]   GENETIC ASPECTS OF [URE3], A NON-MITOCHONDRIAL, CYTOPLASMICALLY INHERITED MUTATION IN YEAST [J].
AIGLE, M ;
LACROUTE, F .
MOLECULAR & GENERAL GENETICS, 1975, 136 (04) :327-335
[2]  
Amberg D.C., 2005, METHODS YEAST GENETI
[3]   Filaments of the Ure2p prion protein have a cross-β core structure [J].
Baxa, U ;
Cheng, NQ ;
Winkler, DC ;
Chiu, TK ;
Davies, DR ;
Sharma, D ;
Inouye, H ;
Kirschner, DA ;
Wickner, RB ;
Steven, AC .
JOURNAL OF STRUCTURAL BIOLOGY, 2005, 150 (02) :170-179
[4]   Architecture of Ure2p prion filaments - The N-terminal domains form a central core fiber [J].
Baxa, U ;
Taylor, KL ;
Wall, JS ;
Simon, MN ;
Cheng, NQ ;
Wickner, RB ;
Steven, AC .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2003, 278 (44) :43717-43727
[5]   Mechanism of inactivation on prion conversion of the Saccharomyces cerevisiae Ure2 protein [J].
Baxa, U ;
Speransky, V ;
Steven, AC ;
Wickner, RB .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2002, 99 (08) :5253-5260
[6]   BIOCHEMICAL AND PHYSICAL-PROPERTIES OF THE PRION PROTEIN FROM 2 STRAINS OF THE TRANSMISSIBLE MINK ENCEPHALOPATHY AGENT [J].
BESSEN, RA ;
MARSH, RF .
JOURNAL OF VIROLOGY, 1992, 66 (04) :2096-2101
[7]   Structural characterization of the fibrillar form of the yeast Saccharomyces cerevisiae prion Ure2p [J].
Bousset, L ;
Redeker, V ;
Decottignies, P ;
Dubois, S ;
Le Maréchal, P ;
Melki, R .
BIOCHEMISTRY, 2004, 43 (17) :5022-5032
[8]   The yeast prion Ure2p retains its native α-helical conformation upon assembly into protein fibrils in vitro [J].
Bousset, L ;
Thomson, NH ;
Radford, SE ;
Melki, R .
EMBO JOURNAL, 2002, 21 (12) :2903-2911
[9]   TSE strain variation [J].
Bruce, ME .
BRITISH MEDICAL BULLETIN, 2003, 66 :99-108
[10]   Protofibrils, pores, fibrils, and neurodegeneration: Separating the responsible protein aggregates from the innocent bystanders [J].
Caughey, B ;
Lansbury, PT .
ANNUAL REVIEW OF NEUROSCIENCE, 2003, 26 :267-298