Ehrlichia chaffeensis Type 1 Secreted Effector TRP47 is a Novel SUMOylated Nucleomodulin

Abstract

Ehrlichia chaffeensis secretes immunodominant and immunoprotective protein effectors into the host cell using a type I secretion system. The best-characterized of these effectors are the tandem repeat proteins (TRPs). In this study, E. chaffeensis TRP47 was identified as a target for host cell SUMOylation and as a novel nucleomodulin with DNA-binding activity and host nuclear localization. Using a microfluidic chip peptide array, a SUMOylation site in the TRP47 N-terminal domain was identified. A SUMO-2 modification was further confirmed using an in vitro SUMOylation assay with recombinant TRP47, and native polySUMOylated TRP47 was immunoprecipitated from E. chaffeensis-infected cell lysates. TRP47 was detected in host cell nuclei primarily at 24 and 48 hours post-infection by immunofluorescence microscopy. A domain in the N-terminal portion of the protein containing a variant MYND-binding motif was identified as the region responsible for TRP47 nuclear localization using ectopically expressed GFP-tagged TRP47 truncation constructs. Recombinant TRP47 also binds human genomic DNA in an electrophoretic mobility shift assay. Taken together, these data demonstrate that E. chaffeensis TRP47 is a nucleomodulin with DNA-binding activity and a non-canonical nuclear localization signal, and is post-translationally modified by SUMOylation, which is a novel modification among bacteria.