Programme 5

Structural studies on the mechanisms of prion-like protein assemblies by using yeast model

Background
The yeast model has been widely used for research on aggregation properties of peptides and proteins associated with neurodegenerative diseases. Dozens of proteins from the yeast Saccharomyces cerevisiae, including Ure2p, Sup35p and Rnq1p, exhibit “prion properties”. Like mammalian prions, yeast prions propagate by a protein-only assembly process, and possess strain diversity through distinct conformers.  Distinct conformations of Ure2p and Sup35p amyloids give rise to different phenotypes of yeast that result in the various yeast prion strains. Though in vitro assembled fibrils by prion domains of Sup35p and Ure2p have been reported to adopt in-register parallel β-sheet structure, the structures of full-length amyloid fibrils of Ure2p and Sup35p from yeast cells remain to be determined. The aim of this project is to study how distinct structures encipher different prion strains by determining the cryo-EM structures of yeast prions which are formed by assembly of Ure2p and Sup35p from yeast cells.

Rotation project
During the 3-month rotation, the student will learn yeast prion strain characterizations, yeast culture, the methods for purification of yeast prions from yeast cells and check the yeast prions by negative-stain electron microscope. This will arrow the students to learn multiple biochemical and biophysical techniques and getting ready for cryo-EM high resolution structural studies.

PhD project
The PhD studies will optimize the purification methods of different prion strains and load the samples for Cryo-EM high resolution structural studies. By comparing the structures from different prion strains, coupled with site-directed mutagenesis and yeast reporter methods, we will study why different structures were formed, how distinct structures encipher prion strains, as well as the effects of other regulation factors like PTMs on the yeast prion structures.