Sis1 over-expression and Rnq1 amyloidogenic regions influence the formation of [PSI+].

(A) Over-expression of Sis1 enhances the spontaneous formation of [PSI+] in [RNQ+] cells. [rnq−] [psi−] and [RNQ+] [psi−] cells were transformed with a Sis1 over-expressing plasmid (Sis1 OE) or an empty vector control (EV). Cultures were plated on medium that selected for [PSI+] cells as well as a non-selective medium to determine the total number of cells plated. Averages and error bars representing standard error of the mean were calculated from at least three independent experiments. Statistical significance was assessed using Student's t-test (*p<0.05, **p<0.01). (B–E) Cells expressing WT Rnq1 or the indicated mutant Rnq1 were transformed with a plasmid to over-express Sup35. [PSI+] induction was monitored by spotting five-fold serial dilutions of normalized cell numbers on SD-ade and SD-ade-his. Representative spottings are shown of cells expressing the mutants (B) A1, (C) A7, or (D) A9 in comparison to expression of WT Rnq1. (E) Summary of data showing how disruption of the amyloidogenic regions by alanine mutations affects [PSI+] induction in each of the [RNQ+] variants. Mutants were categorized as having no effect (white), <5-fold increase (green) or decrease (orange) in [PSI+] induction, or >5-fold increase (blue) or decrease (red) in [PSI+] induction. Black boxes indicate that [RNQ+] was eliminated, while black hash marks indicate that [RNQ+] propagation was altered (see Figure 4). Data are summarized from at least five independent experiments.