Hsp26 and Hsp42 synergize to inhibit spontaneous Sup35 prionogenesis.

<p>(A, B) NM (5 µM) was incubated at 25°C with agitation for 6 h in the presence of increasing concentrations of BSA, Hsp26, Hsp42, or Hsp26 and Hsp42 (0–5 µM). For the mixture of Hsp26 and Hsp42, a 1∶1 ratio was employed. Thus, a concentration of 2 µM on the <i>x</i>-axis reflects 1 µM Hsp26 and 1 µM Hsp42. Fibrillization was measured by Thioflavin-T (ThT) fluorescence (A) or by determining the amount of SDS-resistant NM (B). Values represent means±SD (<i>n</i> = 3). (C) NM (5 µM) was assembled at 25°C with agitation for 6 h in the absence or presence of Hsp26 (0.6 µM or 3 µM), Hsp42 (0.6 µM or 3 µM), or Hsp26 and Hsp42 (0.3 µM or 1.5 µM of each). Reaction products were concentrated and transformed into [<i>psi</i>⁻] cells. No NM and soluble NM served as negative controls. The proportion of [<i>PSI</i>⁺] colonies was then determined. Values represent means±SD (<i>n</i> = 3). (D) Sup35 (5 µM) was incubated at 25°C with agitation for 6 h in the presence of increasing concentrations of BSA, Hsp26, Hsp42, or Hsp26 and Hsp42 (0–5 µM). For the mixture of Hsp26 and Hsp42, a 1∶1 ratio was employed. Thus, a concentration of 2 µM on the <i>x</i>-axis reflects 1 µM Hsp26 and 1 µM Hsp42. Fibrillization was measured by ThT fluorescence. Values represent means±SD (<i>n</i> = 3).