Figure 7.

Ydj1 overexpression blocks Hsp104‐mediated curing of strong [PSI ⁺]^STR in a manner unrelated to changes in the expression of other relevant chaperones.

A. [PSI ⁺]^STR cells were transformed first by empty vector or plasmids overexpressing various J‐proteins, followed by a subsequent transformation with plasmid overexpressing Hsp104 driven by either the GPD (left) or TEF (right), promoter. Color phenotype assays are shown for representative transformants (n ≥ 10).

B. Lysates of representative cells from panel A were resolved by SDD‐AGE and subjected to immunoblot analysis using an antibody specific to Sup35. Dotted lines separate lanes taken from different parts of the same gel.

C. Lysates of a wild‐type strain, or strains overexpressing Hsp104, or both Hsp104 and Ydj1, were resolved by SDS‐PAGE and subjected to immunoblot analysis using antibodies specific for Hsp104, Ydj1, Sis1 or Ssa1–4.

D. Lysates of a wild‐type strain or sis1‐Δ cells containing plasmids expressing truncated Sis1 were resolved on SDS‐PAGE and subjected to immunoblot analysis using an antibody specific to Ydj1. Antibody specific for Ssc1 was used as a loading control.

E. Lysates of a wild‐type strain and strains lacking Apj1 but overexpressing Sis1 or Apj1 were resolved and visualized as in panel D.

F. Lysates of wild‐type and apj1‐Δ strains as well as strains expressing truncated versions of Sis1 from a plasmid in place of endogenous Sis1 or overexpressing Ydj1 were resolved on SDS‐PAGE and subjected to immunoblot analysis using an antibody specific to Apj1. Antibody specific for Ssc1 was used as a loading control.