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The transcriptional repressor EUO regulates both subsets of Chlamydia late genes.

ABSTRACT: The pathogenic bacterium Chlamydia replicates in a eukaryotic host cell via a developmental cycle marked by temporal waves of gene expression. We have previously shown that late genes transcribed by the major chlamydial RNA polymerase, ?(66) RNA polymerase, are regulated by a transcriptional repressor EUO. We now report that EUO also represses promoters for a second subset of late genes that are transcribed by an alternative polymerase called ?(28) RNA polymerase. EUO bound in the vicinity of six ?(28) -dependent promoters and inhibited transcription of each promoter. We used a mutational analysis to demonstrate that the EUO binding site functions as an operator that is necessary and sufficient for EUO-mediated repression of ?(28) -dependent transcription. We also verified specific binding of EUO to ?(66) -dependent and ?(28) -dependent promoters with a DNA immunoprecipitation assay. These findings support a model in which EUO represses expression of both ?(66) -dependent and ?(28) -dependent late genes. We thus propose that EUO is the master regulator of late gene expression in the chlamydial developmental cycle.

SUBMITTER: Rosario CJ

PROVIDER: S-EPMC4428689 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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The transcriptional repressor EUO regulates both subsets of Chlamydia late genes.

Rosario Christopher J CJ Hanson Brett R BR Tan Ming M

Molecular microbiology 20141016 4

The pathogenic bacterium Chlamydia replicates in a eukaryotic host cell via a developmental cycle marked by temporal waves of gene expression. We have previously shown that late genes transcribed by the major chlamydial RNA polymerase, σ(66) RNA polymerase, are regulated by a transcriptional repressor EUO. We now report that EUO also represses promoters for a second subset of late genes that are transcribed by an alternative polymerase called σ(28) RNA polymerase. EUO bound in the vicinity of si ...[more]

PMID: 25250726

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Project description:A critical step in intracellular Chlamydia infection is the production of infectious progeny through the expression of late genes. This differentiation step involves conversion from a reticulate body (RB), which is the replicating form of the bacterium, into an elementary body (EB), which is the developmental form that spreads the infection to a new host cell. EUO is an important chlamydial transcription factor that controls the expression of late genes, but the mechanisms that regulate EUO are not known. We report that a plasmid-encoded protein, Pgp4, enhanced the repressor activity of EUO. Pgp4 did not function as a transcription factor because it did not bind or directly modulate transcription of its target promoters. Instead, Pgp4 increased the ability of EUO to bind and repress EUO-regulated promoters in vitro and physically interacted with EUO in pulldown assays with recombinant proteins. We detected earlier onset of EUO-dependent late gene expression by immunofluorescence microscopy in Pgp4-deficient C. trachomatis and C. muridarum strains. In addition, the absence of Pgp4 led to earlier onset of RB-to-EB conversion in C. muridarum These data support a role for Pgp4 as a negative regulator of chlamydial transcription that delays late gene expression. Our studies revealed that Pgp4 also has an EUO-independent function as a positive regulator of chlamydial transcription.IMPORTANCEChlamydia trachomatis is an important human pathogen that causes more than 150 million active cases of genital and eye infection in the world. This obligate intracellular bacterium produces infectious progeny within an infected human cell through the expression of late chlamydial genes. We showed that the ability of a key chlamydial transcription factor, EUO, to repress late genes was enhanced by a plasmid-encoded protein, Pgp4. In addition, studies with Chlamydia Pgp4-deficient strains provide evidence that Pgp4 delays late gene expression in infected cells. Thus, Pgp4 is a novel regulator of late gene expression in Chlamydia through its ability to enhance the repressor function of EUO.

Project description:Chlamydia trachomatis is an obligate intracellular bacterium whose unique developmental cycle consists of an infectious elementary body and a replicative reticulate body. Progression of this developmental cycle requires temporal control of the transcriptome. In addition to the three chlamydial sigma factors (σ66, σ28, and σ54) that recognize promoter sequences of genes, chlamydial transcription factors are expected to play crucial roles in transcriptional regulation. Here, we investigate the function of GrgA, a Chlamydia-specific transcription factor, in C. trachomatis transcriptomic expression. We show that 10 to 30 min of GrgA overexpression induces 13 genes, which likely comprise the direct regulon of GrgA. Significantly, σ66-dependent genes that code for two important transcription repressors are components of the direct regulon. One of these repressors is Euo, which prevents the expression of late genes during early phases. The other is HrcA, which regulates molecular chaperone expression and controls stress response. The direct regulon also includes a σ28-dependent gene that codes for the putative virulence factor PmpI. Furthermore, overexpression of GrgA leads to decreased expression of almost all tRNAs. Transcriptomic studies suggest that GrgA, Euo, and HrcA have distinct but overlapping indirect regulons. These findings, together with temporal expression patterns of grgA, euo, and hrcA, indicate that a transcriptional regulatory network of these three transcription factors plays critical roles in C. trachomatis growth and development. IMPORTANCE Chlamydia trachomatis is the most prevalent sexually transmitted bacterial pathogen worldwide and is a leading cause of preventable blindness in underdeveloped areas as well as some developed countries. Chlamydia carries genes that encode a limited number of known transcription factors. While Euo is thought to be critical for early chlamydial development, the functions of GrgA and HrcA in the developmental cycle are unclear. Activation of euo and hrcA immediately following GrgA overexpression indicates that GrgA functions as a master transcriptional regulator. In addition, by broadly inhibiting tRNA expression, GrgA serves as a key regulator of chlamydial protein synthesis. Furthermore, by upregulating pmpI, GrgA may act as an upstream virulence determinant. Finally, genes coregulated by GrgA, Euo, and HrcA likely play critical roles in chlamydial growth and developmental control.

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The transcriptional repressor EUO regulates both subsets of Chlamydia late genes.

Publications

The transcriptional repressor EUO regulates both subsets of Chlamydia late genes.

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