Enabling stop codon read-through translation in bacteria as a probe for amyloid aggregation

9 p.-5 fig.Amyloid aggregation of the eukaryotic translation terminator eRF3/Sup35p, the [PSI+] prion, empowers yeast ribosomes to read-through UGA stop codons. No similar functional prion, skipping a stop codon, has been found in Escherichia coli, a fact possibly due to the efficient back-up systems found in bacteria to rescue non-stop complexes. Here we report that engineering hydrophobic amyloidogenic repeats from a synthetic bacterial prion-like protein (RepA-WH1) into the E. coli releasing factor RF1 promotes its aggregation and enables ribosomes to continue with translation through a premature UAG stop codon located in a β-galactosidase reporter. To our knowledge, intended aggregation of a termination factor is a way to overcome the bacterial translation quality checkpoint that had not been reported so far. We also show the feasibility of using the amyloidogenic RF1 chimeras as a reliable, rapid and cost-effective system to screen for molecules inhibiting intracellular protein amyloidogenesis in vivo, by testing the effect on the chimeras of natural polyphenols with known anti-amyloidogenic properties. Resveratrol exhibits a clear amyloid-solubilizing effect in this assay, showing no toxicity to bacteria or interference with the enzymatic activity of β-galactosidase.This work has been financed with grants (CSD2009-00088, BIO2012-30852 and BIO2015-68730-R) from Spanish AEI and UE-FEDER. R.G. is a member of the CIB-CSIC Intramural Research Program “Macromolecular Machines for Better Life” (MACBET).Peer reviewe