TALE-Like Effectors Are an Ancestral Feature of the Ralstonia solanacearum Species Complex and Converge in DNA Targeting Specificity

Abstract :

Ralstonia solanacearum, a species complex of bacterial plant pathogens divided intofour monophyletic phylotypes, causes plant diseases in tropical climates around theworld. Some strains exhibit a broad host range on solanaceous hosts, while othersare highly host-specific as for example some banana-pathogenic strains. Previous studies showed that transcription activator-like (TAL) effectors from Ralstonia, termedRipTALs, are capable of activating reporter genes in planta, if these are precededby a matching effector binding element (EBE). RipTALs target DNA via their centralrepeat domain (CRD), where one repeat pairs with one DNA-base of the given EBE.The repeat variable diresidue dictates base repeat specificity in a predictable fashion,known as the TALE code. In this work, we analyze RipTALs across all phylotypes ofthe Ralstonia solanacearum species complex. We find that RipTALs are prevalent inphylotypes I and IV but absent from most phylotype III and II strains (10/12, 8/14, 1/24,and 1/5 strains contained a RipTAL, respectively). RipTALs originating from strains ofthe same phylotype show high levels of sequence similarity (>98%) in the N-terminaland C-terminal regions, while RipTALs isolated from different phylotypes show 47–91% sequence similarity in those regions, giving rise to four RipTAL classes. We showthat, despite sequence divergence, the base preference for guanine, mediated by theN-terminal region, is conserved across RipTALs of all classes. Using the number andorder of repeats found in the CRD, we functionally sub-classify RipTALs, introduce anew simple nomenclature, and predict matching EBEs for all seven distinct RipTALsidentified. We experimentally study RipTAL EBEs and uncover that some RipTALs areable to target the EBEs of other RipTALs, referred to as cross-reactivity. In particular,RipTALs from strains with a broad host range on solanaceous hosts cross-react on eachother’s EBEs. Investigation of sequence divergence between RipTAL repeats allows for areconstruction of repeat array biogenesis, for example through slipped strand mispairingor gene conversion. Using these studies we show how RipTALs of broad host rangestrains evolved convergently toward a shared target sequence. Finally, we discuss thedifferences between TALE-likes of plant pathogens in the context of disease ecology.