Aggregation and toxicity in bacteria of Rep-WH1 prion-like proteins from the phytopathogen Xylella fastidiosa

Xylella fastidiosa is a global bacterial phytopathogen that colonizes the xylem vessels, leading to their occlusion and thus to plant death. The expression of the amyloidogenic, prion-like WH1 domain of the replication protein RepA from the Pseudomonas savastanoi plasmid pPS10 severely damages Escherichia coli cells. Here, we characterize the WH1 domains in the Rep proteins from the X. fastidiosa plasmids pXF51 and pXF64. The expression of these Rep-WH1 proteins, as happens with the archetypical RepA-WH1 protein from pPS10, decreased the culturability of E. coli in vitro, promoting dormancy. Expression of pXF51 Rep-WH1 causes the conversion of the bacillary cells into large cocci in which this protein forms a hydrogel, whereas pXF64 Rep-WH1 generates compact amyloid-like aggregates, as do those for pPS10 RepA-WH1, thus defining distinct protein-specific phase separations. In X. fastidiosa, the Rep-WH1 proteins also reduce culturability by altering the integrity of the cell envelope, driving bacterial cells into dormancy, while co-aggregation of a subset of the X. fastidiosa host proteins probably contribute to reducing bacterial virulence in plants. These findings open a way to the exploitation of the intrinsic amyloidogenic potential of endogenous plasmid-encoded Rep proteins for the control of X. fastidiosa, aiming to reduce its economic and environmental impact.IMPORTANCEXylella fastidiosa is a bacterial pathogen that causes the withering of many woody plants. Currently, there is no efficient way to eradicate this phytopathogen, which menaces crops and natural environments worldwide. In this study, we show that the expression in X. fastidiosa and E. coli of Rep-WH1 proteins, originally encoded by plasmids found either in X. fastidiosa or in Pseudomonas, leads to their aggregation as prions (i.e., transmissible protein particles) that impact bacterial cells by significantly reducing their fitness and, potentially, their virulence. These findings on prion-like Rep-WH1 proteins may constitute the grounds for the development of an efficient weapon against X. fastidiosa.