Transcriptional and Translational Responsiveness of the Neisseria gonorrhoeae Type IV Secretion System to Conditions of Host Infections

Callaghan, Melanie M.; Klimowicz, Amy K.; Shockey, Abigail C.; Kane, John S; Pepperell, Caitlin S.; Dillard, Joseph P.

doi:10.1128/iai.00519-21

Cited by 6 publications

(14 citation statements)

References 102 publications

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“…Only copper sulfate (CuSO 4 ) had any visible effect on colony color (data not shown). Although the magnitude of activation by copper seen in the Biolog assays or on plates is only moderate, this finding aligns with other instances of copper-dependent enhancement of T4SS protein expression, described in ( Callaghan et al, 2021 ).…”

Section: Resultssupporting

confidence: 89%

“…Investigations of the expression of the gonococcal T4SS have begun to reveal a complex regulatory network, with transcriptional, translational, and post-translational mechanisms all at play ( Pachulec et al, 2014 ; Ramsey et al, 2014 , 2015 ; Callaghan et al, 2021 ). Quantitative transcript data indicate that for both the traH operon ( traH , traG , and atlA ) and the parA operon (containing parA and parB ), transcripts are readily detected in vitro.…”

Section: Resultsmentioning

confidence: 99%

“…Due to the natural transformability of N. gonorrhoeae at all stages of growth, this active DNA release can facilitate horizontal gene transfer ( Dillard and Seifert, 2001 ; Hamilton and Dillard, 2006 ; Salgado-Pabón et al, 2007 ; Shockey, 2019 ). Regulation of gonococcal T4SS expression and activity is only beginning to be understood ( Ramsey et al, 2015 ; Callaghan et al, 2021 ).…”

Section: Introductionmentioning

confidence: 99%

“…More is known about the regulation of T4SS expression in gonococci, and RNA-mediated mechanisms are recently emerging as the regulatory network is probed ( Ramsey et al, 2015 ; Callaghan et al, 2021 ). Several sRNAs have been identified within the GGI and have yet to be functionally characterized ( Remmele et al, 2014 ).…”

Section: Introductionmentioning

confidence: 99%

“…Several sRNAs have been identified within the GGI and have yet to be functionally characterized ( Remmele et al, 2014 ). Recent work has also implicated the Fur regulon in regulating some aspects of T4SS expression ( Callaghan et al, 2021 ), and this regulon is known to utilize sRNA intermediates to control iron-responsive genes ( Mellin et al, 2007 ; Yu et al, 2016 ). The GGI encodes an RNA switch in the traH 5′ untranslated region (UTR) which controls protein expression from the P traH -derived transcript ( Ramsey et al, 2015 ).…”

Section: Introductionmentioning

confidence: 99%

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Expression, Localization, and Protein Interactions of the Partitioning Proteins in the Gonococcal Type IV Secretion System

et al. 2021

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Partitioning proteins are well studied as molecular organizers of chromosome and plasmid segregation during division, however little is known about the roles partitioning proteins can play within type IV secretion systems. The single-stranded DNA (ssDNA)-secreting gonococcal T4SS has two partitioning proteins, ParA and ParB. These proteins work in collaboration with the relaxase TraI as essential facilitators of type IV secretion. Bacterial two-hybrid experiments identified interactions between each partitioning protein and the relaxase. Subcellular fractionation demonstrated that ParA is found in the cellular membrane, whereas ParB is primarily in the membrane, but some of the protein is in the soluble fraction. Since TraI is known to be membrane-associated, these data suggest that the gonococcal relaxosome is a membrane-associated complex. In addition, we found that translation of ParA and ParB is controlled by an RNA switch. Different mutations within the stem-loop sequence predicted to alter folding of this RNA structure greatly increased or decreased levels of the partitioning proteins.

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Section: Resultssupporting

confidence: 89%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Expression, Localization, and Protein Interactions of the Partitioning Proteins in the Gonococcal Type IV Secretion System

et al. 2021

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The Gonococcal Genetic Island defines distinct sub-populations of Neisseria gonorrhoeae

Youngblom

Shockey²,

Callaghan

et al. 2023

Microbial Genomics

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The incidence of gonorrhoea is increasing at an alarming pace, and therapeutic options continue to narrow as a result of worsening drug resistance. Neisseria gonorrhoeae is naturally competent, allowing the organism to adapt rapidly to selection pressures including antibiotics. A sub-population of N. gonorrhoeae carries the Gonococcal Genetic Island (GGI), which encodes a type IV secretion system (T4SS) that secretes chromosomal DNA. Previous research has shown that the GGI increases transformation efficiency in vitro, but the extent to which it contributes to horizontal gene transfer (HGT) during infection is unknown. Here we analysed genomic data from clinical isolates of N. gonorrhoeae to better characterize GGI+ and GGI− sub-populations and to delineate patterns of variation at the locus itself. We found the element segregating at an intermediate frequency (61%), and it appears to act as a mobile genetic element with examples of gain, loss, exchange and intra-locus recombination within our sample. We further found evidence suggesting that GGI+ and GGI− sub-populations preferentially inhabit distinct niches with different opportunities for HGT. Previously, GGI+ isolates were reported to be associated with more severe clinical infections, and our results suggest this could be related to metal-ion trafficking and biofilm formation. The co-segregation of GGI+ and GGI− isolates despite mobility of the element suggests that both niches inhabited by N. gonorrhoeae remain important to its overall persistence as has been demonstrated previously for cervical- and urethral-adapted sub-populations. These data emphasize the complex population structure of N. gonorrhoeae and its capacity to adapt to diverse niches.

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Bactericidal effectors of the Stenotrophomonas maltophilia type IV secretion system: functional definition of the nuclease TfdA and structural determination of TfcB

Cobe,

Dey,

Minasov

et al. 2024

mBio

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Stenotrophomonas maltophilia expresses a type IV protein secretion system (T4SS) that promotes contact-dependent killing of other bacteria and does so partly by secreting the effector TfcB. Here, we report the structure of TfcB, comprising an N-terminal domain similar to the catalytic domain of glycosyl hydrolase (GH-19) chitinases and a C-terminal domain for recognition and translocation by the T4SS. Utilizing a two-hybrid assay to measure effector interactions with the T4SS coupling protein VirD4, we documented the existence of five more T4SS substrates. One of these was protein 20845, an annotated nuclease. A S. maltophilia mutant lacking the gene for 20845 was impaired for killing Escherichia coli , Klebsiella pneumoniae , and Pseudomonas aeruginosa . Moreover, the cloned 20845 gene conferred robust toxicity, with the recombinant E. coli being rescued when 20845 was co-expressed with its cognate immunity protein. The 20845 effector was an 899 amino-acid protein, comprised of a GHH-nuclease domain in its N-terminus, a large central region of indeterminant function, and a C-terminus for secretion. Engineered variants of the 20845 gene that had mutations in the predicted catalytic site did not impede E. coli , indicating that the antibacterial effect of 20845 involves its nuclease activity. Using flow cytometry with DNA staining, we determined that 20845, but not its mutant variants, confers a loss in DNA content of target bacteria. Database searches revealed that uncharacterized homologs of 20845 occur within a range of bacteria. These data indicate that the S. maltophilia T4SS promotes interbacterial competition through the action of multiple toxic effectors, including a potent, novel DNase. IMPORTANCE Stenotrophomonas maltophilia is a multi-drug-resistant, Gram-negative bacterium that is an emerging pathogen of humans. Patients with cystic fibrosis are particularly susceptible to S. maltophilia infection. In hospital water systems and various types of infections, S. maltophilia co-exists with other bacteria, including other pathogens such as Pseudomonas aeruginosa . We previously demonstrated that S. maltophilia has a functional VirB/D4 type VI protein secretion system (T4SS) that promotes contact-dependent killing of other bacteria. Since most work on antibacterial systems involves the type VI secretion system, this observation remains noteworthy. Moreover, S. maltophilia currently stands alone as a model for a human pathogen expressing an antibacterial T4SS. Using biochemical, genetic, and cell biological approaches, we now report both the discovery of a novel antibacterial nuclease (TfdA) and the first structural determination of a bactericidal T4SS effector (TfcB).

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Transcriptional and Translational Responsiveness of the Neisseria gonorrhoeae Type IV Secretion System to Conditions of Host Infections

Cited by 6 publications

References 102 publications

Expression, Localization, and Protein Interactions of the Partitioning Proteins in the Gonococcal Type IV Secretion System

Expression, Localization, and Protein Interactions of the Partitioning Proteins in the Gonococcal Type IV Secretion System

The Gonococcal Genetic Island defines distinct sub-populations of Neisseria gonorrhoeae

Bactericidal effectors of the Stenotrophomonas maltophilia type IV secretion system: functional definition of the nuclease TfdA and structural determination of TfcB

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