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Shigeko Kawai-Noma¹, Chan-Gi Pack^2,3, Toshikazu Tsuji¹, Masataka Kinjo² and Hideki Taguchi^1,*

¹ Department of Medical Genome Sciences, Graduate School of Frontier Sciences, University of Tokyo, FSB401, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan
² Laboratory of Molecular Cell Dynamics, Graduate School of Life Sciences, Hokkaido University, Sapporo 001-0021, Japan
³ Cellular Systems Modeling Team, RIKEN Advanced Science Institute, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan

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.

These authors contributed equally.

^* taguchi{at}k.u-tokyo.ac.jp