Fernandez-Bellot E, Guillemet E, Cullin C
Centre de Génétique Moléculaire, Centre National de la Recherche Scientifique, Avenue de la Terrasse, 91198 Gif-sur-Yvette Cedex, France.
EMBO J. 2000 Jul 3;19(13):3215-22. doi: 10.1093/emboj/19.13.3215.
The non-Mendelian element [URE3] of yeast is considered to be a prion form of the Ure2 protein. The [URE3] phenotype occurs at a frequency of 10(-5) in haploid yeast strains, is reversible, and its frequency is increased by overexpressing the URE2 gene. We created a new mutant of the Ure2 protein, called H2p, which results in a 1000-fold increase in the rate of [URE3] occurrence. To date, only the overexpression of various C-terminal truncated mutants of Ure2p gives rise to a comparable level. The h2 allele is, thus, the first characterized URE2 allele that induces prion formation when expressed at a low level. By shuffling mutated and wild-type domains of URE2, we also created the first mutant Ure2 protein that is functional and induces prion formation. We demonstrate that the domains of URE2 function synergistically in cis to induce [URE3] formation, which highlights the importance of intramolecular interactions in Ure2p folding. Additionally, we show using a green fluorescent protein (GFP) fusion protein that the h2 allele exhibits numerous filiform structures that are not generated by the wild-type protein.
酵母的非孟德尔遗传元件[URE3]被认为是Ure2蛋白的一种朊病毒形式。[URE3]表型在单倍体酵母菌株中的出现频率为10^(-5)，具有可逆性，并且通过过表达URE2基因可提高其频率。我们创建了一种新的Ure2蛋白突变体，称为H2p，它使[URE3]的出现率增加了1000倍。迄今为止，只有Ure2p各种C端截短突变体的过表达能产生类似水平。因此，h2等位基因是第一个在低水平表达时诱导朊病毒形成的已鉴定URE2等位基因。通过对URE2的突变和野生型结构域进行改组，我们还创建了第一个具有功能并能诱导朊病毒形成的突变Ure2蛋白。我们证明URE2的结构域在顺式作用中协同发挥功能以诱导[URE3]形成，这突出了分子内相互作用在Ure2p折叠中的重要性。此外，我们使用绿色荧光蛋白（GFP）融合蛋白表明，h2等位基因表现出许多野生型蛋白不会产生的丝状结构。
