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The [URE3] yeast prion: from genetics to biochemistry
1Centre de Genetique Moleculaire, Centre National de la Recherche Scientifique, Gif-sur-Yvette Cedex, 91198, France. anton.komar@cgm. cnrs-gif.fr.
Biochemistry. Biokhimiia
|January 29, 2000
Summary
The yeast prion [URE3], an altered Ure2 protein, can self-assemble into various structures in vitro, similar to other prions. Ure2p aggregation in vivo doesn't always result in the [URE3] phenotype, suggesting complex prion formation mechanisms.
Area of Science:
- Biochemistry
- Molecular Biology
- Yeast Genetics
Background:
- The [URE3] element in Saccharomyces cerevisiae is a non-Mendelian genetic factor, representing an altered prion form of the Ure2 protein.
- Prions are proteins that can adopt alternative conformations, leading to heritable traits without changes in DNA sequence.
Purpose of the Study:
- To investigate the in vitro assembly properties of recombinant Ure2 protein.
- To delineate the structural domains of Ure2 protein and their interactions.
- To explore the relationship between Ure2p aggregation and the [URE3] prion phenotype.
Main Methods:
- In vitro protein assembly assays (dimers, tetramers, octamers, fibrils).
- Computational, genetic, biochemical, and structural analyses to map protein domains.
- Two-hybrid system and affinity binding experiments for in vivo and in vitro interaction studies.
- Monitoring Ure2p aggregation using Ure2p-GFP fusion and fluorescence.
Main Results:
- Recombinant Ure2p self-assembles in vitro into soluble oligomers and insoluble fibrils, mirroring mechanisms of other prions.
- A new boundary between prion-forming and catalytic domains was identified at Met-94, with the C-terminal region (94-354) being folded and the N-terminal region (1-94) unstructured.
- The N-terminal and C-terminal domains interact both in vivo and in vitro.
- The catalytic C-terminal domain can be synthesized independently via internal ribosome binding.
- Ure2p aggregation in vivo does not invariably lead to the [URE3] phenotype.
Conclusions:
- Yeast prion formation, exemplified by [URE3], involves complex self-assembly mechanisms potentially shared across different prions.
- The distinct structural domains of Ure2 protein play crucial roles in its folding, interaction, and potential prion propagation.
- Ure2p aggregation is a necessary but not sufficient condition for the [URE3] prion state, indicating regulatory mechanisms governing prion establishment.