Chlamydia trachomatis Plasmid-Encoded Pgp4 Is a Transcriptional Regulator of Virulence-Associated Genes

Abstract

c Chlamydia trachomatis causes chronic inflammatory diseases of the eye and genital tract and has global medical importance. The chlamydial plasmid plays an important role in the pathophysiology of these diseases, as plasmid-deficient organisms are highly attenuated. The cryptic plasmid carries noncoding RNAs and eight conserved open reading frames (ORFs). To understand plasmid gene function, we generated plasmid shuttle vectors with deletions in each of the eight ORFs. The individual deletion mutants were used to transform chlamydiae and the transformants were characterized phenotypically and at the transcriptional level. We show that pgp1, -2, -6, and -8 are essential for plasmid maintenance, while the other ORFs can be deleted and the plasmid stably maintained. We further show that a pgp4 knockout mutant exhibits an in vitro phenotype similar to its isogenic plasmidless strain, in terms of abnormal inclusion morphology and lack of glycogen accumulation. Microarray and qRT-PCR analysis revealed that Pgp4 is a transcriptional regulator of plasmid-encoded pgp3 and multiple chromosomal genes, including the glycogen synthase gene glgA, that are likely important in chlamydial virulence. Our findings have major implications for understanding the plasmid's role in chlamydial pathogenesis at the molecular level.

Plasmid deficiency in urogenital isolates ofChlamydia trachomatisreduces infectivity and virulence in a mouse model

“…In the context of what has already been reported by others regarding the chlamydial plasmid and in vivo fitness, our results add to the growing body of evidence that in many hosts and at specific anatomical sites, the plasmid plays a role in modulating infectivity and virulence, but this role is not universal. Now that a means to genetically manipulate chlamydiae and selectively mutate the plasmid has been elaborated and the function of the plasmid can be dissected and studied in greater detail (Wang et al ., 2011; Song et al ., 2013). …”

Simkania negevensis, an insight into the biology and clinical importance of a novel member of the Chlamydiales order

“…An adaptive immune response was however elicited enabling protection against re-infection ( Kari et al, 2011;O'Connell et al, 2007;Porcella et al, 2015). The mechanisms by which the chlamydial plasmid modulates the pathogenesis are still unclear, but they are probably related to: (i) direct effect of proteins encoded by the plasmid (Li et al, 2008;Liu et al, 2014, p. 201) and (ii) regulation of the expression of other virulence factors encoded by the chromosome (Carlson et al, 2008;Song et al, 2013). Interestingly, the S. negevensis plasmid encodes various proteins of interest, among which is one copy of the Macrophage Infectivity Potentiator (Mip) (Collingro et al, 2011).…”

Emancipating Chlamydia: Advances in the Genetic Manipulation of a Recalcitrant Intracellular Pathogen

“…Transformation of exogenous DNA into Chlamydia has been achieved via several methods, including electroporation, dendrimer-based delivery, and a calcium chloride-based treatment (14)(15)(16)(17)(18)(19). Successful and stable transformation of C. trachomatis LGV L2 with an Escherichia coli-C. trachomatis L2 shuttle plasmid conferring ␤-lactam resistance led to the development of expression vectors for expressing Chlamydia open reading frames (ORFs), fluorescent proteins (green fluorescent protein [GFP], cyan fluorescent protein [CFP], mCherry, and mKate2), and reporter proteins (␤-galactosidase, adenylate cyclase, and glycogen synthase kinase [GSK]-tagged proteins) (19)(20)(21)(22)(23)(24)(25)(26). A conditional expression vector was also developed using a tetracycline-inducible system as well as vectors conferring chloramphenicol and blasticidin resistance (21,(26)(27)(28)(29).…”