Chaperone-amyloid interactions

伴侣-淀粉样蛋白相互作用

• 批准号: 8553390
• 负责人: Daniel Masison
• 金额: $ 7.45万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8553390
• 关键词: Affect; Amyloid; Binding; Biochemical; C-terminal; Cells; Disease; Genetic; Glutamine; Goals; In Vitro; Kinetics; Mediating; Methods; Molecular; Molecular Chaperones; Mutation; N-terminal; Phenotype; Prions; Property; Protein Structure Initiative; Proteins; Structure; Toxic effect; Variant; Work; Yeasts; amyloid formation; falls; human Huntingtin protein; in vivo; interest; mutant; overexpression; polyglutamine; polyproline; sup35

Expressing proteins with high propensity to from amyloid, such as disease-associated huntingtin (Htt) fragments with 103 glutamines (HttQ103), is toxic in yeast containing either the PIN+ prion, which is the amyloid form of Rnq1, or PSI+ prion, which is the amyloid form of Sup35. In an effort to understand this toxicity we examined in detail the effects of expressing various Htt fragments in cells propagating different prion variants. HttQP103, which has a polyproline region at the C-terminal end of the polyQ repeat region, is significantly more toxic in yeast with PSI+ than PIN+. This toxicity observed in cells with a strong PSI+ variant was not seen in a weak PSI+ variant, which has more soluble Sup35 than the strong variant. Additionally, expression of Sup35p MC region, which remains soluble in PSI+ yeast because it lacks the N-terminal prion domain, almost completely rescued HttQP103 toxicity. Overexpressing the essential Sup45p, which binds to and functions with Sup35p, did not restore viability. Therefore, the toxicity of HttQP103 in yeast containing the PSI+ prion is primarily due to sequestration of the essential Sup35 protein. We are also continuing studies of how alterations in primary structure of prion proteins can affect chaperone interactions. We have been isolating mutants of prion proteins that fall into two major classes, those that interfere with propagation of prions formed of wild type proteins and those that don't. Both types of mutants contain subclasses that, in the absence of wild type protein, either form prions with weakened phenotypes or are unable to form prions. We are using genetic and biochemical methods to determine if the observed effects on prion phenotype are due to differences in how the prion proteins self-associate or interact with various chaperones. Our preliminary work suggests that the prion protein mutations can influence the propagation of amyloid directly, or the interaction of chaperones that in turn affect how efficiently the prions propagate.

表达具有淀粉样蛋白高倾向的蛋白质，例如疾病相关的亨廷顿蛋白（HTT）片段，具有103个谷氨酰胺（HTTQ103），在酵母中有毒，其中含有PIN+ PRION，含有PIN+ PRION，这是RNQ1或PSI+ Prion+ PSI+ Prion的淀粉样蛋白形式，或者是PSI+ Prion的淀粉样蛋白，这是Sup3的淀粉样蛋白。为了理解这种毒性，我们详细研究了表达各种HTT片段在传播不同prion变体的细胞中的影响。 HTTQP103在PolyQ重复区域的C末端具有多生区域，在PSI+的酵母中毒性明显高于PIN+。在弱的PSI+变体中未看到这种毒性在具有强PSI+变体的细胞中观察到的毒性，该变体的可溶性sup35比强的变体更可溶。此外，Sup35p MC区域的表达（由于缺乏N末端prion域缺乏）而溶于PSI+酵母菌中，几乎完全挽救了HTTQP103的毒性。与SUP35P结合并函数的Essential Sup45p过表达并未恢复生存能力。因此，HTTQP103在包含PSI+ Prion的酵母中的毒性主要是由于必需SUP35蛋白的隔离。 我们还在继续研究prion蛋白主要结构的变化如何影响伴侣的相互作用。我们一直在隔离落入两个主要类别的prion蛋白的突变体，这些类别会干扰由野生型蛋白质和不繁殖的prions的传播。两种类型的突变体都包含子类，在没有野生型蛋白质的情况下，要么形成弱的表型prions，要么无法形成王室。我们使用遗传和生化方法来确定观察到的对prion表型的影响是否是由于prion蛋白如何自我缔合或与各种伴侣相互作用的差异。我们的初步工作表明，prion蛋白突变可以直接影响淀粉样蛋白的传播，或者会影响伴侣的相互作用，而伴侣的相互作用反过来影响王室的传播效率。