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1.
The protein-remodeling factor Hsp104 governs inheritance of [PSI+], a yeast prion formed by self-perpetuating amyloid conformers of the translation termination factor Sup35. Perplexingly, either excess or insufficient Hsp104 eliminates [PSI+]. In vitro, at low concentrations, Hsp104 catalyzed the formation of oligomeric intermediates that proved critical for the nucleation of Sup 35 fibrillization de novo and displayed a conformation common among amyloidogenic polypeptides. At higher Hsp104 concentrations, amyloidogenic oligomerization and contingent fibrillization were abolished. Hsp104 also disassembled mature fibers in a manner that initially exposed new surfaces for conformational replication but eventually exterminated prion conformers. These Hsp104 activities differed in their reaction mechanism and can explain [PSI+] inheritance patterns. 相似文献
2.
According to the prion hypothesis, atypical phenotypes arise when a prion protein adopts an alternative conformation and persist when that form assembles into self-replicating aggregates. Amyloid formation in vitro provides a model for this protein-misfolding pathway, but the mechanism by which this process interacts with the cellular environment to produce transmissible phenotypes is poorly understood. Using the yeast prion Sup35/[PSI(+)], we found that protein conformation determined the size distribution of aggregates through its interactions with a molecular chaperone. Shifts in this range created variations in aggregate abundance among cells because of a size threshold for transmission, and this heterogeneity, along with aggregate growth and fragmentation, induced age-dependent fluctuations in phenotype. Thus, prion conformations may specify phenotypes as population averages in a dynamic system. 相似文献
3.
Nucleated conformational conversion and the replication of conformational information by a prion determinant 总被引：2，自引：0，他引：2
Serio TR Cashikar AG Kowal AS Sawicki GJ Moslehi JJ Serpell L Arnsdorf MF Lindquist SL 《Science (New York, N.Y.)》2000,289(5483):1317-1321
Prion proteins can serve as genetic elements by adopting distinct physical and functional states that are self-perpetuating and heritable. The critical region of one prion protein, Sup35, is initially unstructured in solution and then forms self-seeded amyloid fibers. We examined in vitro the mechanism by which this state is attained and replicated. Structurally fluid oligomeric complexes appear to be crucial intermediates in de novo amyloid nucleus formation. Rapid assembly ensues when these complexes conformationally convert upon association with nuclei. This model for replicating protein-based genetic information, nucleated conformational conversion, may be applicable to other protein assembly processes. 相似文献
4.
Taylor KL Cheng N Williams RW Steven AC Wickner RB 《Science (New York, N.Y.)》1999,283(5406):1339-1343
The [URE3] non-Mendelian genetic element of Saccharomyces cerevisiae is an infectious protein (prion) form of Ure2p, a regulator of nitrogen catabolism. Here, synthetic Ure2p1-65 were shown to polymerize to form filaments 40 to 45 angstroms in diameter with more than 60 percent beta sheet. Ure2p1-65 specifically induced full-length native Ure2p to copolymerize under conditions where native Ure2p alone did not polymerize. Like Ure2p in extracts of [URE3] strains, these 180- to 220-angstrom-diameter filaments were protease resistant. The Ure2p1-65-Ure2p cofilaments could seed polymerization of native Ure2p to form thicker, less regular filaments. All filaments stained with Congo Red to produce the green birefringence typical of amyloid. This self-propagating amyloid formation can explain the properties of [URE3]. 相似文献
5.
Creating a protein-based element of inheritance 总被引：1，自引：0，他引：1
Proteins capable of self-perpetuating changes in conformation and function (known as prions) can serve as genetic elements. To test whether novel prions could be created by recombinant methods, a yeast prion determinant was fused to the rat glucocorticoid receptor. The fusion protein existed in different heritable functional states, switched between states at a low spontaneous rate, and could be induced to switch by experimental manipulations. The complete change in phenotype achieved by transferring a prion determinant from one protein to another confirms the protein-only nature of prion inheritance and establishes a mechanism for engineering heritable changes in phenotype that should be broadly applicable. 相似文献
6.
A yeast prion, Mod5, promotes acquired drug resistance and cell survival under environmental stress 总被引：1，自引：0，他引：1
Prion conversion from a soluble protein to an aggregated state may be involved in the cellular adaptation of yeast to the environment. However, it remains unclear whether and how cells actively use prion conversion to acquire a fitness advantage in response to environmental stress. We identified Mod5, a yeast transfer RNA isopentenyltransferase lacking glutamine/asparagine-rich domains, as a yeast prion protein and found that its prion conversion in yeast regulated the sterol biosynthetic pathway for acquired cellular resistance against antifungal agents. Furthermore, selective pressure by antifungal drugs on yeast facilitated the de novo appearance of Mod5 prion states for cell survival. Thus, phenotypic changes caused by active prion conversion under environmental selection may contribute to cellular adaptation in living organisms. 相似文献
7.
Klöhn PC Farmer M Linehan JM O'Malley C Fernandez de Marco M Taylor W Farrow M Khalili-Shirazi A Brandner S Collinge J 《Science (New York, N.Y.)》2012,335(6064):52
Intraperitoneal administration of ICSM18 and 35, monoclonal antibodies against prion protein (PrP), has been shown to significantly delay the onset of prion disease in mice, and humanized versions are candidate therapeutics for prion and Alzheimer's diseases. However, a previous report of severe and widespread apoptosis after intracerebral injection of anti-PrP monoclonal antibodies raised concerns about such therapy and led to an influential model of prion neurotoxicity via cross-linking of cell surface PrP by disease-related PrP aggregates. In extensive studies including ICSM18 and 35, fully humanized ICSM18, and the previously reported proapoptotic antibodies, we found no evidence of apoptosis, thereby questioning this model of prion neurotoxicity. 相似文献
8.
A rare conformation of the prion protein, PrPSc, is found only in mammals with transmissible prion diseases and represents either the infectious agent itself or a major component of it. The mechanism for initiating PrPSc formation is unknown. We report that PrP retrogradely transported out of the endoplasmic reticulum produced both amorphous aggregates and a PrPSc-like conformation in the cytosol. The distribution between these forms correlated with the rate of appearance in the cytosol. Once conversion to the PrPSc-like conformation occurred, it was sustained. Thus, PrP has an inherent capacity to promote its own conformational conversion in mammalian cells. These observations might explain the origin of PrPSc. 相似文献
9.
Changes in prion protein (PrP) folding are associated with fatal neurodegenerative disorders, but the neurotoxic species is unknown. Like other proteins that traffic through the endoplasmic reticulum, misfolded PrP is retrograde transported to the cytosol for degradation by proteasomes. Accumulation of even small amounts of cytosolic PrP was strongly neurotoxic in cultured cells and transgenic mice. Mice developed normally but acquired severe ataxia, with cerebellar degeneration and gliosis. This establishes a mechanism for converting wild-type PrP to a highly neurotoxic species that is distinct from the self-propagating PrP(Sc) isoform and suggests a potential common framework for seemingly diverse PrP neurodegenerative disorders. 相似文献
10.
Tuite MF 《Science (New York, N.Y.)》2000,289(5479):556-557
Ever since Prusiner first proposed his radical "protein-only" hypothesis to explain how certain infectious proteins (prions) are transmitted from one mammal to another in the absence of DNA or RNA, scientists have been trying to prove him right (or wrong). The study of mammalian prions, such as those causing Creutzfeldt-Jakob disease in humans, scrapie in sheep and mad cow disease in cattle, has been slow to yield answers. However, as Tuite discusses in his Perspective, the Sup35p and Ure2p proteins of yeast that exist in both normal and infectious forms are providing evidence that the "protein-only" hypothesis may be right (Sparrer et al.). 相似文献
11.
K K Hsiao M Scott D Foster D F Groth S J DeArmond S B Prusiner 《Science (New York, N.Y.)》1990,250(4987):1587-1590
Transgenic mice were created to assess genetic linkage between Gerstmann-Str?ussler-Scheinker syndrome and a leucine substitution at codon 102 of the human prion protein gene. Spontaneous neurologic disease with spongiform degeneration and gliosis similar to that in mouse scrapie developed at a mean age of 166 days in 35 mice expressing mouse prion protein with the leucine substitution. Thus, many of the clinical and pathological features of Gerstmann-Str?ussler-Scheinker syndrome are reproduced in transgenic mice containing a prion protein with a single amino acid substitution, illustrating that a neurodegenerative process similar to a human disease can be genetically modeled in animals. 相似文献
12.
[目的]明确酵母突变株的最优富硒条件。[方法]以经紫外线诱变筛选所得的突变酵母菌为出发菌种,采用培养时间、发酵培养基中硒浓度、接种量的单因素试验,研究最优富硒条件。[结果]发酵初期酵母中的硒含量随培养时间的延长逐渐增加,培养35h时,酵母中的硒含量最大。随着培养基中硒浓度的增加,酵母菌的生物量呈减少趋势,硒含量呈增加趋势。综合考虑确定培养基中硒浓度为20μg/ml。随着接种量的增加,富硒酵母的产量逐渐增大,接种量为20%时有所下降,接种量10%时硒含量最大。[结论]突变酵母菌最优的富硒条件是发酵培养时间35h,培养基含硒量20μg/ml,接种量10%,此时生物量提高了58.00%,硒含量提高了74.87%,达1343mg/kg。 相似文献
13.
14.
Reversible conversion of monomeric human prion protein between native and fibrilogenic conformations
Jackson GS Hosszu LL Power A Hill AF Kenney J Saibil H Craven CJ Waltho JP Clarke AR Collinge J 《Science (New York, N.Y.)》1999,283(5409):1935-1937
Prion propagation involves the conversion of cellular prion protein (PrPC) into a disease-specific isomer, PrPSc, shifting from a predominantly alpha-helical to beta-sheet structure. Here, conditions were established in which recombinant human PrP could switch between the native alpha conformation, characteristic of PrPC, and a compact, highly soluble, monomeric form rich in beta structure. The soluble beta form (beta-PrP) exhibited partial resistance to proteinase K digestion, characteristic of PrPSc, and was a direct precursor of fibrillar structures closely similar to those isolated from diseased brains. The conversion of PrPC to beta-PrP in suitable cellular compartments, and its subsequent stabilization by intermolecular association, provide a molecular mechanism for prion propagation. 相似文献
15.
Petkova AT Leapman RD Guo Z Yau WM Mattson MP Tycko R 《Science (New York, N.Y.)》2005,307(5707):262-265
Amyloid fibrils commonly exhibit multiple distinct morphologies in electron microscope and atomic force microscope images, often within a single image field. By using electron microscopy and solid-state nuclear magnetic resonance measurements on fibrils formed by the 40-residue beta-amyloid peptide of Alzheimer's disease (Abeta(1-40)), we show that different fibril morphologies have different underlying molecular structures, that the predominant structure can be controlled by subtle variations in fibril growth conditions, and that both morphology and molecular structure are self-propagating when fibrils grow from preformed seeds. Different Abeta(1-40) fibril morphologies also have significantly different toxicities in neuronal cell cultures. These results have implications for the mechanism of amyloid formation, the phenomenon of strains in prion diseases, the role of amyloid fibrils in amyloid diseases, and the development of amyloid-based nano-materials. 相似文献
16.
[目的]揭示SGR基因的作用机理。[方法]构建水稻SGR基因在酵母双杂交体系中的诱饵载体,并对其表达进行鉴定,将目的基因SGR与诱饵质粒载体pGBKT7通过双酶切定向重组构建诱饵质粒pGBKT7-SGR,将pGBKT7-SGR转入酵母菌株Y187中,利用Western印迹法检测其在酵母中的表达,通过缺陷性培养基培养进行自激活检测。[结果]将鉴定正确的重组质粒测序结果与SGR基因比较,序列完全一致,阅读框分析正确。载体pGBKT7-SGR在酵母菌株Y187中可以正确表达出融合蛋白。重组诱饵pGBKT7-SGR质粒对酵母无毒性,其表达产物不能激活酵母菌株Y187的营养缺陷型报告基因。[结论]该重组诱饵质粒可用于酵母双杂交体系,该研究为从cDNA文库筛选水稻诱饵蛋白SGR的互作蛋白奠定了基础。 相似文献
17.
Heikenwalder M Zeller N Seeger H Prinz M Klöhn PC Schwarz P Ruddle NH Weissmann C Aguzzi A 《Science (New York, N.Y.)》2005,307(5712):1107-1110
Prions typically accumulate in nervous and lymphoid tissues. Because proinflammatory cytokines and immune cells are required for lymphoid prion replication, we tested whether inflammatory conditions affect prion pathogenesis. We administered prions to mice with five inflammatory diseases of the kidney, pancreas, or liver. In all cases, chronic lymphocytic inflammation enabled prion accumulation in otherwise prion-free organs. Inflammatory foci consistently correlated with lymphotoxin up-regulation and ectopic induction of FDC-M1+ cells expressing the normal cellular prion protein PrPC. By contrast, inflamed organs of mice lacking lymphotoxin-alpha or its receptor did not accumulate the abnormal isoform PrPSc, nor did they display infectivity upon prion inoculation. By expanding the tissue distribution of prions, chronic inflammatory conditions may act as modifiers of natural and iatrogenic prion transmission. 相似文献
18.
构建了绵羊朊蛋白的原核表达载体,获得了高纯度的融合表达蛋白。并在热力学因素的作用下,研究了重组绵羊朊蛋白的构象转化。以基因型为ARQ/ARQ蒙古绵羊的血液DNA为模板,利用DNA重组技术,将绵羊朊蛋白正常成熟蛋白基因OvPrP插入表达载体pET30a,在大肠杆菌BL2l(DE3)中高效表达,获得的表达产物以包涵体形式存在,并对其进行纯化和复性。超滤浓缩后浓度约为0.5 mg/mL的OvPrP97-234,进行热力学处理,利用远紫外线圆二色谱(CD)分析热力学处理前、后蛋白的二级结构的变化,同时,对热力学处理前、后的蛋白进行了蛋白酶K抗性的检测,并对其高级结构进行了预测。结果表明:获得的表达产物经SDS-PAGE分析可见分子量为16kD的蛋白条带,Western-blotting的鉴定证实了所获得的蛋白是特异性的朊蛋白。经Jascow32软件分析,测得天然构象的OvPrP97-234的二级结构含量为:α螺旋为28.8%、β折叠为0%、转角和无规卷曲为71.1%,无蛋白酶K的抗性。经过热力学处理之后,OvPrP97-234的二级结构含量为:α螺旋为19.3%,β折叠为36.9%,转角和无规卷曲为43.8%,有一定... 相似文献
19.
Hu J Shibata Y Voss C Shemesh T Li Z Coughlin M Kozlov MM Rapoport TA Prinz WA 《Science (New York, N.Y.)》2008,319(5867):1247-1250
The tubular structure of the endoplasmic reticulum (ER) appears to be generated by integral membrane proteins, the reticulons and a protein family consisting of DP1 in mammals and Yop1p in yeast. Here, individual members of these families were found to be sufficient to generate membrane tubules. When we purified yeast Yop1p and incorporated it into proteoliposomes, narrow tubules (approximately 15 to 17 nanometers in diameter) were generated. Tubule formation occurred with different lipids; required essentially only the central portion of the protein, including its two long hydrophobic segments; and was prevented by mutations that affected tubule formation in vivo. Tubules were also formed by reconstituted purified yeast Rtn1p. Tubules made in vitro were narrower than normal ER tubules, due to a higher concentration of tubule-inducing proteins. The shape and oligomerization of the "morphogenic" proteins could explain the formation of the tubular ER. 相似文献
20.
The passage of large-sized herpesviral capsids through the nuclear lamina and the inner nuclear membrane to leave the nucleus requires a dissolution of the nuclear lamina. Here, we report on the functions of M50/p35, a beta-herpesviral protein of murine cytomegalovirus. M50/p35 inserts into the inner nuclear membrane and is aggregated by a second viral protein, M53/p38, to form the capsid docking site. M50/p35 recruits the cellular protein kinase C for phosphorylation and dissolution of the nuclear lamina, suggesting that herpesviruses target a critical element of nuclear architecture. 相似文献