TargeTron Inactivation of Chlamydia trachomatis
            <i>gseA</i>
            Results in a Lipopolysaccharide 3-Deoxy-
            <scp>d</scp>
            -Manno-Oct-2-Ulosonic Acid-Deficient Strain That Is Cytotoxic for Cells

DeBoer, Addison G; Lei, Lei; Yang, Chunfu; Martens, Craig; Anzick, Sarah L.; Antonioli-Schmit, Sophia; Suchland, Robert J.; McClarty, Grant; Rockey, Daniel D.

doi:10.1128/iai.00096-23

Cited by 4 publications

References 37 publications

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Zoonotic and other veterinary chlamydiae – an update, the role of the plasmid and plasmid-mediated transformation

Marti,

Shima,

Boutin

et al. 2024

Pathogens and Disease

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The obligate intracellular bacterial genus Chlamydia harbours species with zoonotic potential, particularly C. psittaci, causative agent of psittacosis, and C. abortus, which may lead to miscarriage in pregnant women. The impact of other bird chlamydiae such as C. avium, C. gallinaceae and C. buteonis, or reptilian species such as C. crocodili, amongst others, on human health is unclear. The chlamydial native plasmid, a suspected virulence factor, is present in all currently described 14 Chlamydia species except for some plasmid-free strains. The plasmid is also the primary tool to study chlamydial genetics, a still developing field that has mostly focused on C. trachomatis. Only recently, genetic transformation of C. felis, C. pecorum, C. pneumoniae, C. psittaci and C. suis has succeeded, but existing methods have yet to be refined. In this review article, we will provide an update on the recent developments concerning the zoonotic potential of chlamydiae. Furthermore, we present an overview about the current state of knowledge regarding the chlamydial plasmid in terms of prevalence and significance as a virulence factor. Finally, we give insights into the progress of developing genetic tools for chlamydial species other than C. trachomatis with a special focus on zoonotic and veterinary chlamydiae.

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Zoonotic and other veterinary chlamydiae – an update, the role of the plasmid and plasmid-mediated transformation

Marti,

Shima,

Boutin

et al. 2024

Pathogens and Disease

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Requirement of GrgA forChlamydiainfectious progeny production, optimal growth, and efficient plasmid maintenance

Lü

Wang

Wurihan

et al. 2023

Preprint

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Chlamydia, an obligate intracellular bacterial pathogen, has a unique developmental cycle involving the differentiation of invading elementary bodies (EBs) to noninfectious reticulate bodies (RBs), replication of RBs, and redifferentiation of RBs into progeny EBs. Progression of this cycle is regulated by three sigma factors, which direct the RNA polymerase to their respective target gene promoters. We hypothesized that theChlamydia-specific transcriptional regulator GrgA, previously shown to activate σ66 and σ28, plays an essential role in chlamydial development and growth. To test this hypothesis, we applied a novel genetic tool known as dependence on plasmid-mediated expression (DOPE) to createChlamydia trachomatiswith conditional GrgA-deficiency. We show that GrgA-deficientC. trachomatisRBs have a growth rate that is approximately half of the normal rate and fail to transition into progeny EBs. In addition, GrgA-deficientC. trachomatisfail to maintain its virulence plasmid. Results of RNA-seq analysis indicate that GrgA promotes RB growth by optimizing tRNA synthesis and expression of nutrient-acquisition genes, while it enables RB-to-EB conversion by facilitating the expression of a histone and outer membrane proteins required for EB morphogenesis. GrgA also regulates numerous other late genes required for host cell exit and subsequent EB invasion into host cells. Importantly, GrgA stimulates the expression of σ54, the third and last sigma factor, and its activator AtoC, and thereby indirectly upregulating the expression of σ54-dependent genes. In conclusion, our work demonstrates that GrgA is a master transcriptional regulator inChlamydiaand plays multiple essential roles in chlamydial pathogenicity.IMPORTANCEHallmarks of the developmental cycle of the obligate intracellular pathogenic bacteriumChlamydiaare the primary differentiation of the infectious elementary body (EB) into the proliferative reticulate body (RB) and the secondary differentiation of RBs back into EBs. The mechanisms regulating these transitions remain unclear. In this report, we developed an effective novel strategy termed DOPE that allows for the knockdown of essential genes inChlamydia. We demonstrate that GrgA, aChlamydia-specific transcription factor, is essential for the secondary differentiation and optimal growth of RBs. We also show that GrgA, a chromosome-encoded regulatory protein, controls the maintenance of the chlamydial virulence plasmid. Transcriptomic analysis further indicates that GrgA functions as a critical regulator of all three sigma factors that recognize different promoter sets at developmental stages. The DOPE strategy outlined here should provide a valuable tool for future studies examining chlamydial growth, development, and pathogenicity.

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Requirement of GrgA for Chlamydia infectious progeny production, optimal growth, and efficient plasmid maintenance

Lu,

Wang,

Wurihan

et al. 2024

mBio

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Chlamydia , an obligate intracellular bacterial pathogen, has a unique developmental cycle involving the differentiation of invading elementary bodies (EBs) to noninfectious reticulate bodies (RBs), replication of RBs, and redifferentiation of RBs into progeny EBs. Progression of this cycle is regulated by three sigma factors, which direct the RNA polymerase to their respective target gene promoters. We hypothesized that the Chlamydia- specific transcriptional regulator GrgA, previously shown to activate σ66 and σ28, plays an essential role in chlamydial development and growth. To test this hypothesis, we applied a novel genetic tool known as dependence on plasmid-mediated expression to create Chlamydia trachomatis with conditional GrgA deficiency. We show that GrgA-deficient C. trachomatis RBs have a growth rate that is approximately half of the normal rate and fail to transition into progeny EBs. In addition, GrgA-deficient C. trachomatis fails to maintain its virulence plasmid. Results of RNA-Seq analysis indicate that GrgA promotes RB growth by optimizing tRNA synthesis and expression of nutrient-acquisition genes, while it enables RB-to-EB conversion by facilitating the expression of a histone and outer membrane proteins required for EB morphogenesis. GrgA also regulates numerous other late genes required for host cell exit and subsequent EB invasion into host cells. Importantly, GrgA stimulates the expression of σ54, the third and last sigma factor, and its activator, AtoC, and thereby indirectly upregulating the expression of σ54-dependent genes. In conclusion, our work demonstrates that GrgA is a master transcriptional regulator in Chlamydia and plays multiple essential roles in chlamydial pathogenicity. IMPORTANCE Hallmarks of the developmental cycle of the obligate intracellular pathogenic bacterium Chlamydia are the primary differentiation of the infectious elementary body (EB) into the proliferative reticulate body (RB) and the secondary differentiation of RBs back into EBs. The mechanisms regulating these transitions remain unclear. In this report, we developed an effective novel strategy termed dependence on plasmid-mediated expression (DOPE) that allows for the knockdown of essential genes in Chlamydia . We demonstrate that GrgA, a Chlamydia -specific transcription factor, is essential for the secondary differentiation and optimal growth of RBs. We also show that GrgA, a chromosome-encoded regulatory protein, controls the maintenance of the chlamydial virulence plasmid. Transcriptomic analysis further indicates that GrgA functions as a critical regulator of all three sigma factors that recognize different promoter sets at developmental stages. The DOPE strategy outlined here should provide a valuable tool for future studies examining chlamydial growth, development, and pathogenicity.

show abstract

TargeTron Inactivation of Chlamydia trachomatis gseA Results in a Lipopolysaccharide 3-Deoxy- d -Manno-Oct-2-Ulosonic Acid-Deficient Strain That Is Cytotoxic for Cells

Cited by 4 publications

References 37 publications

Zoonotic and other veterinary chlamydiae – an update, the role of the plasmid and plasmid-mediated transformation

Zoonotic and other veterinary chlamydiae – an update, the role of the plasmid and plasmid-mediated transformation

Requirement of GrgA forChlamydiainfectious progeny production, optimal growth, and efficient plasmid maintenance

Requirement of GrgA for Chlamydia infectious progeny production, optimal growth, and efficient plasmid maintenance

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