Ehrlichia chaffeensis TRP32 is a nucleomodulin that directly regulates expression of host genes governing differentiation and proliferation

Tierra R. Farris, Paige S. Dunphy, Bing Zhu, Clayton E. Kibler, Jere McBride

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Ehrlichia chaffeensis is an obligately intracellular bacterium that reprograms the mononuclear phagocyte through diverse effector- host interactions to modulate numerous host cell processes, including transcription. In a previous study, we reported that E. chaffeensis TRP32, a type 1 secreted effector, interacts with multiple host nucleus-associated proteins and also autoactivates reporter gene expression in yeast. In this study, we demonstrate that TRP32 is a nucleomodulin that binds host DNA and alters host gene transcription. TRP32 enters the host cell nucleus via a noncanonical translocation mechanism that involves phosphorylation of Y179 located in a C-terminal trityrosine motif. Both genistein and mutation of Y179 inhibited TRP32 nuclear entry. An electromobility shift assay (EMSA) demonstrated TRP32 host DNA binding via its tandem repeat domain. TRP32 DNA-binding and motif preference were further confirmed by supershift assays, as well as competition and mutant probe analyses. Using chromatin immunoprecipitation with next-generation sequencing (ChIP-seq), we determined that TRP32 binds a G-rich motif primarily within±500 bp of the gene transcription start site. An ontology analysis identified genes involved in processes such as immune cell differentiation, chromatin remodeling, and RNA transcription and processing as primary TRP32 targets. TRP32- bound genes (n=1,223) were distributed on all chromosomes and included several global regulators of proliferation and inflammation such as those encoding FOS, JUN, AKT3, and NRAS and noncoding RNA genes microRNA 21 (miRNA 21) and miRNA 142. TRP32 target genes were differentially regulated during infection, the majority of which were repressed, and direct repression/activation of these genes by TRP32 was confirmed in vitro with a cellular luciferase reporter assay.

Original languageEnglish (US)
Pages (from-to)3182-3194
Number of pages13
JournalInfection and Immunity
Volume84
Issue number11
DOIs
StatePublished - 2016

Fingerprint

Ehrlichia chaffeensis
Gene Expression
Genes
MicroRNAs
Untranslated RNA
Nucleotide Motifs
Tandem Repeat Sequences
Chromatin Assembly and Disassembly
Genistein
Chromatin Immunoprecipitation
Transcription Initiation Site
DNA
Phagocytes
Luciferases
Cell Nucleus
Reporter Genes
Transcriptional Activation
Cell Differentiation
Chromosomes
Yeasts

ASJC Scopus subject areas

  • Parasitology
  • Microbiology
  • Immunology
  • Infectious Diseases

Cite this

Ehrlichia chaffeensis TRP32 is a nucleomodulin that directly regulates expression of host genes governing differentiation and proliferation. / Farris, Tierra R.; Dunphy, Paige S.; Zhu, Bing; Kibler, Clayton E.; McBride, Jere.

In: Infection and Immunity, Vol. 84, No. 11, 2016, p. 3182-3194.

Research output: Contribution to journalArticle

Farris, Tierra R. ; Dunphy, Paige S. ; Zhu, Bing ; Kibler, Clayton E. ; McBride, Jere. / Ehrlichia chaffeensis TRP32 is a nucleomodulin that directly regulates expression of host genes governing differentiation and proliferation. In: Infection and Immunity. 2016 ; Vol. 84, No. 11. pp. 3182-3194.
@article{2f2a9d6252db49a8a2a89fd10c0b982b,
title = "Ehrlichia chaffeensis TRP32 is a nucleomodulin that directly regulates expression of host genes governing differentiation and proliferation",
abstract = "Ehrlichia chaffeensis is an obligately intracellular bacterium that reprograms the mononuclear phagocyte through diverse effector- host interactions to modulate numerous host cell processes, including transcription. In a previous study, we reported that E. chaffeensis TRP32, a type 1 secreted effector, interacts with multiple host nucleus-associated proteins and also autoactivates reporter gene expression in yeast. In this study, we demonstrate that TRP32 is a nucleomodulin that binds host DNA and alters host gene transcription. TRP32 enters the host cell nucleus via a noncanonical translocation mechanism that involves phosphorylation of Y179 located in a C-terminal trityrosine motif. Both genistein and mutation of Y179 inhibited TRP32 nuclear entry. An electromobility shift assay (EMSA) demonstrated TRP32 host DNA binding via its tandem repeat domain. TRP32 DNA-binding and motif preference were further confirmed by supershift assays, as well as competition and mutant probe analyses. Using chromatin immunoprecipitation with next-generation sequencing (ChIP-seq), we determined that TRP32 binds a G-rich motif primarily within±500 bp of the gene transcription start site. An ontology analysis identified genes involved in processes such as immune cell differentiation, chromatin remodeling, and RNA transcription and processing as primary TRP32 targets. TRP32- bound genes (n=1,223) were distributed on all chromosomes and included several global regulators of proliferation and inflammation such as those encoding FOS, JUN, AKT3, and NRAS and noncoding RNA genes microRNA 21 (miRNA 21) and miRNA 142. TRP32 target genes were differentially regulated during infection, the majority of which were repressed, and direct repression/activation of these genes by TRP32 was confirmed in vitro with a cellular luciferase reporter assay.",
author = "Farris, {Tierra R.} and Dunphy, {Paige S.} and Bing Zhu and Kibler, {Clayton E.} and Jere McBride",
year = "2016",
doi = "10.1128/IAI.00657-16",
language = "English (US)",
volume = "84",
pages = "3182--3194",
journal = "Infection and Immunity",
issn = "0019-9567",
publisher = "American Society for Microbiology",
number = "11",

}

TY - JOUR

T1 - Ehrlichia chaffeensis TRP32 is a nucleomodulin that directly regulates expression of host genes governing differentiation and proliferation

AU - Farris, Tierra R.

AU - Dunphy, Paige S.

AU - Zhu, Bing

AU - Kibler, Clayton E.

AU - McBride, Jere

PY - 2016

Y1 - 2016

N2 - Ehrlichia chaffeensis is an obligately intracellular bacterium that reprograms the mononuclear phagocyte through diverse effector- host interactions to modulate numerous host cell processes, including transcription. In a previous study, we reported that E. chaffeensis TRP32, a type 1 secreted effector, interacts with multiple host nucleus-associated proteins and also autoactivates reporter gene expression in yeast. In this study, we demonstrate that TRP32 is a nucleomodulin that binds host DNA and alters host gene transcription. TRP32 enters the host cell nucleus via a noncanonical translocation mechanism that involves phosphorylation of Y179 located in a C-terminal trityrosine motif. Both genistein and mutation of Y179 inhibited TRP32 nuclear entry. An electromobility shift assay (EMSA) demonstrated TRP32 host DNA binding via its tandem repeat domain. TRP32 DNA-binding and motif preference were further confirmed by supershift assays, as well as competition and mutant probe analyses. Using chromatin immunoprecipitation with next-generation sequencing (ChIP-seq), we determined that TRP32 binds a G-rich motif primarily within±500 bp of the gene transcription start site. An ontology analysis identified genes involved in processes such as immune cell differentiation, chromatin remodeling, and RNA transcription and processing as primary TRP32 targets. TRP32- bound genes (n=1,223) were distributed on all chromosomes and included several global regulators of proliferation and inflammation such as those encoding FOS, JUN, AKT3, and NRAS and noncoding RNA genes microRNA 21 (miRNA 21) and miRNA 142. TRP32 target genes were differentially regulated during infection, the majority of which were repressed, and direct repression/activation of these genes by TRP32 was confirmed in vitro with a cellular luciferase reporter assay.

AB - Ehrlichia chaffeensis is an obligately intracellular bacterium that reprograms the mononuclear phagocyte through diverse effector- host interactions to modulate numerous host cell processes, including transcription. In a previous study, we reported that E. chaffeensis TRP32, a type 1 secreted effector, interacts with multiple host nucleus-associated proteins and also autoactivates reporter gene expression in yeast. In this study, we demonstrate that TRP32 is a nucleomodulin that binds host DNA and alters host gene transcription. TRP32 enters the host cell nucleus via a noncanonical translocation mechanism that involves phosphorylation of Y179 located in a C-terminal trityrosine motif. Both genistein and mutation of Y179 inhibited TRP32 nuclear entry. An electromobility shift assay (EMSA) demonstrated TRP32 host DNA binding via its tandem repeat domain. TRP32 DNA-binding and motif preference were further confirmed by supershift assays, as well as competition and mutant probe analyses. Using chromatin immunoprecipitation with next-generation sequencing (ChIP-seq), we determined that TRP32 binds a G-rich motif primarily within±500 bp of the gene transcription start site. An ontology analysis identified genes involved in processes such as immune cell differentiation, chromatin remodeling, and RNA transcription and processing as primary TRP32 targets. TRP32- bound genes (n=1,223) were distributed on all chromosomes and included several global regulators of proliferation and inflammation such as those encoding FOS, JUN, AKT3, and NRAS and noncoding RNA genes microRNA 21 (miRNA 21) and miRNA 142. TRP32 target genes were differentially regulated during infection, the majority of which were repressed, and direct repression/activation of these genes by TRP32 was confirmed in vitro with a cellular luciferase reporter assay.

UR - http://www.scopus.com/inward/record.url?scp=84994680377&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84994680377&partnerID=8YFLogxK

U2 - 10.1128/IAI.00657-16

DO - 10.1128/IAI.00657-16

M3 - Article

VL - 84

SP - 3182

EP - 3194

JO - Infection and Immunity

JF - Infection and Immunity

SN - 0019-9567

IS - 11

ER -