FIGURE 1: Effect of genetic modifiers of toxic HDEL-Aβ42 on Aβ42-RF oligomer formation and Aβ42-MRF aggregates into toxic SDS-stable oligomers in yeast.

(A) Overexpressed Yap1802 but not other previously described [24] HDEL-Aβ42 modifiers dramatically enhanced the translational termination factor activity of Aβ42-RF. Aβ42-RF expressing cells (CUP1::Aβ42-RF) lacking chromosomal SUP35 were transformed with a plasmid carrying the indicated gene under control of the GAL1 promoter. Ten-fold serial dilutions of transformants are shown on plasmid selective non-inducing media (SD-Ura); galactose plasmid selective media (SGal-Ura) to turn on each gene (↑ORF); +Ade medium containing copper to overexpress Aβ42-RF (↑ORF ↑Aβ42-RF); the identical -Ade medium SGal-Ura-Ade+Cu2+ (-Ade ↑ORF ↑Aβ42-RF) to determine translational termination factor activity as read-through of the ade1-14 nonsense mutant. The Aβ42-RF overexpressed in cells was shown to have reduced translational termination factor activity as cells grew on -Ade due to aggregation of the fusion protein into small oligomers (Aβ42-RF-e. v.). However, the translation termination factor activity was retained in yeast cells overexpressing Aβ42m2-RF (Aβ42 aggregation-deficient mutant) or YAP1802, due to the absence of oligomer formation, resulting in reduced growth on -Ade (Aβ42m2-RF-e. v.).

(B) Yap1802 suppression of Aβ42-RF oligomerization by immunoblot analysis. Total cell lysates were prepared from 2 independent transformants of an Aβ42-RF strain carrying YAP1802 (+) or an empty vector (-) plasmid. Both Aβ42-RF and YAP1802 were overexpressed (SGal-Ura-Ade+Cu2+) prior to lysis. Immunoblots were probed with anti-Sup35 RF to evaluate the level of oligomers and monomers. ↑Yap1802 indicates overexpression of YAP1802. The identification of bands as Aβ42-RF dimers and monomers was determined by the estimated sizes of the bands in the immunoblot.

By continuing to use the site, you agree to the use of cookies. more information

The cookie settings on this website are set to "allow cookies" to give you the best browsing experience possible. If you continue to use this website without changing your cookie settings or you click "Accept" below then you are consenting to this. Please refer to our "privacy statement" and our "terms of use" for further information.

Close