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Histone methylation by NUE, a novel nuclear effector of the intracellular pathogen Chlamydia trachomatis.

ABSTRACT: Sequence analysis of the genome of the strict intracellular pathogen Chlamydia trachomatis revealed the presence of a SET domain containing protein, proteins that primarily function as histone methyltransferases. In these studies, we demonstrated secretion of this protein via a type III secretion mechanism. During infection, the protein is translocated to the host cell nucleus and associates with chromatin. We therefore named the protein nuclear effector (NUE). Expression of NUE in mammalian cells by transfection reconstituted nuclear targeting and chromatin association. In vitro methylation assays confirmed NUE is a histone methyltransferase that targets histones H2B, H3 and H4 and itself (automethylation). Mutants deficient in automethylation demonstrated diminished activity towards histones suggesting automethylation functions to enhance enzymatic activity. Thus, NUE is secreted by Chlamydia, translocates to the host cell nucleus and has enzymatic activity towards eukaryotic substrates. This work is the first description of a bacterial effector that directly targets mammalian histones.

SUBMITTER: Pennini ME

PROVIDER: S-EPMC2904774 | biostudies-literature | 2010 Jul

REPOSITORIES: biostudies-literature

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Histone methylation by NUE, a novel nuclear effector of the intracellular pathogen Chlamydia trachomatis.

Pennini Meghan E ME Perrinet Stéphanie S Dautry-Varsat Alice A Subtil Agathe A

PLoS pathogens 20100715 7

Sequence analysis of the genome of the strict intracellular pathogen Chlamydia trachomatis revealed the presence of a SET domain containing protein, proteins that primarily function as histone methyltransferases. In these studies, we demonstrated secretion of this protein via a type III secretion mechanism. During infection, the protein is translocated to the host cell nucleus and associates with chromatin. We therefore named the protein nuclear effector (NUE). Expression of NUE in mammalian cel ...[more]

PMID: 20657819

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Project description:Background: Chlamydia trachomatis is a prolific human pathogen that can cause serious long-term conditions if left untreated. Recent developments in Chlamydia genetics have opened the door to conducting targeted and random mutagenesis experiments to identify gene function. In the present study, an inducible transposon mutagenesis approach was developed for C. trachomatis using a self-replicating vector to deliver the transposon-transposase cassette - a significant step towards our ultimate aim of achieving saturation mutagenesis of the Chlamydia genome. Methods: The low transformation efficiency of C. trachomatis necessitated the design of a self-replicating vector carrying the transposon mutagenesis cassette (i.e. the Himar-1 transposon containing the beta lactamase gene as well as a hyperactive transposase gene under inducible control of the tet promoter system with the addition of a riboswitch). Chlamydia transformed with this vector (pSW2-RiboA-C9Q) were induced at 24 hours post-infection. Through dual control of transcription and translation, basal expression of transposase was tightly regulated to stabilise the plasmid prior to transposition. Results: Here we present the preliminary sequencing results of transposon mutant pools of both C. trachomatis biovars, using two plasmid-free representatives: urogenital strain C. trachomatis SWFP- and the lymphogranuloma venereum isolate L2(25667R). DNA sequencing libraries were generated and analysed using Oxford Nanopore Technologies' MinION technology. This enabled 'proof of concept' for the methods as an initial low-throughput screen of mutant libraries; the next step is to employ high throughput sequencing to assess saturation mutagenesis. Conclusions: This significant advance provides an efficient method for assaying C. trachomatis gene function and will enable the identification of the essential gene set of C. trachomatis. In the long-term, the methods described herein will add to the growing knowledge of chlamydial infection biology leading to the discovery of novel drug or vaccine targets.

Dataset Information

Histone methylation by NUE, a novel nuclear effector of the intracellular pathogen Chlamydia trachomatis.

Publications

Histone methylation by NUE, a novel nuclear effector of the intracellular pathogen Chlamydia trachomatis.

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