Abstract
Ehrlichia chaffeensis is an obligately intracellular bacterium that establishes infection in mononuclear phagocytes through largely undefined reprogramming strategies including modulation of host gene transcription. In this study, we demonstrate that the E. chaffeensis effector TRP47 enters the host cell nucleus and binds regulatory regions of host genes relevant to infection. TRP47 was observed in the nucleus of E. chaffeensis-infected host cells, and nuclear localization was dependent on a variant MYND-binding domain. An electrophoretic mobility shift assay (EMSA) demonstrated that TRP47 directly binds host DNA via its tandem repeat domain. Utilizing chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) with E. chaffeensis-infected cells, TRP47 was found to bind at multiple sites in the human genome (n = 2,051 at p < 10−30). Ontology analysis identified genes involved in functions such as immune response, cytoskeletal organization, and signal transduction. TRP47-bound genes included RNA-coding genes, many of these linked to cell proliferation or apoptosis. Comparison of TRP47 binding sites with those of previously-identified E. chaffeensis nucleomodulins identified multiple genes and gene functional categories in common including intracellular transport, cell signaling, and transcriptional regulation. Further, motif analysis followed by EMSA with synthetic oligonucleotides containing discovered motifs revealed a conserved TRP47 DNA-binding motif. This study reveals that TRP47 is a nucleomodulin that enters the nucleus via a MYND-binding domain and appears to play a role in host cell reprogramming by regulation of transcription.
Highlights
Ehrlichia chaffeensis is an obligately intracellular bacterium and the etiologic agent of human monocytic ehrlichiosis (HME), a life-threatening NIAID emerging tick-borne zoonosis
Recent studies demonstrated that E. chaffeensis effectors Ank200, TRP120, and TRP32 act as nucleomodulins that target genes involved in processes known or reasonably assumed to have an impact on ehrlichial survival
The results of this study suggest that TRP47 has similar nucleomodulin activity
Summary
Ehrlichia chaffeensis is an obligately intracellular bacterium and the etiologic agent of human monocytic ehrlichiosis (HME), a life-threatening NIAID emerging tick-borne zoonosis. E. chaffeensis preferentially infects mononuclear phagocytes and subverts host defenses by direct activation of cellular pathways and through other complex molecular mechanisms that involve type 1-secreted ankyrin and tandem repeat protein (TRP) effectors. Previous studies have demonstrated that TRP effectors interact with a diverse array of host proteins involved in biological processes such as cell signaling, intracellular transport, metabolism, protein post-translational modification, apoptosis, and regulation of transcription [2]. E. chaffeensis appears to target host gene transcription to manipulate apoptosis, cell differentiation, signal transduction, immune response, metabolism, and intracellular transport during infection [3,4]; the mechanisms by which E. chaffeensis alters transcription are not fully understood
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