The yeast prion [PSI(+)] is a protein-based heritable element, in which aggregates of Sup35 protein are transmitted to daughter cells in a non-Mendelian manner. To elucidate the mechanism of the transmission, we have developed methods to directly analyse the dynamics of Sup35 fused with GFP in single mother-daughter pairs. As it is known that the treatment of yeast cells with guanidine hydrochloride (GuHCl) cures [PSI(+)] by perturbing Hsp104, a prion-remodelling factor, we analysed the diffusion profiles of Sup35-GFP in GuHCl-treated [PSI(+)] cells using fluorescence correlation spectroscopy (FCS). FCS analyses revealed that Sup35-GFP diffusion in the daughter cells was faster; that is, the Sup35-GFP particle was smaller, than that in the mother [PSI(+)] cells, and it eventually reached the diffusion profiles in [psi(-)] cells. We then analysed the flux of Sup35-GFP oligomers from mother to daughter [PSI(+)] cells in the presence of GuHCl, using a modified fluorescent recovery after photobleaching technique, and found that the flux of the diffuse oligomers was completely inhibited. The noninvasive methods described here can be applied to other protein-based transmissible systems inside living cells.

• PMID: 19674118
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Evaluation Study | Journal Article | Research Support, Non-U.S. Gov't

Authors

Kawai-Noma S, Pack CG, Tsuji T, Kinjo M, Taguchi H

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