Mutation of the Conserved Calcium-Binding Motif in Neisseria gonorrhoeae PilC1 Impacts Adhesion but Not Piliation

Cheng, Yuan; Johnson, Michael D. L.; Burillo-Kirch, Christine; Mocny, Jeffrey C.; Anderson, John M.; Garrett, Christopher K.; Redinbo, Matthew R.; Thomas, C.

doi:10.1128/iai.00493-13

Cited by 27 publications

(38 citation statements)

References 39 publications

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“…Also, TFP may physically obstruct cell contact with the substrate (33). Although pilY1-1611 was able to produce TFP, it is possible that pili were produced at a lower level than in the WT or that the mutation affected TFP function, as suggested previously for N. gonorrhoeae mutants with alterations in different PilY1 homologs, where it is difficult to observe differences in the number of pili and other characteristics accurately (45). Fewer TFP could result in more access for type I pili and, therefore, in increased attachment.…”

Section: Discussionmentioning

confidence: 75%

“…As with K. kingae and N. gonorrhoeae (14,53), mutation of individual X. fastidiosa PilY1 homologs does not appear to affect piliation, indicating that neither pilY1-PD0023 nor pilY1-PD1611 is required for pilus biogenesis. However, when all homologs in K. kingae, N. gonorrhoeae, and P. aeruginosa are mutated, no pili are produced (14,34,45). To verify that all three X. fastidiosa PilY1 homologs are involved in pilus biogenesis, simultaneous mutation is required.…”

Section: Discussionmentioning

confidence: 99%

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Calcium-Enhanced Twitching Motility in Xylella fastidiosa Is Linked to a Single PilY1 Homolog

Cruz

Parker

Cobine

et al. 2014

Appl Environ Microbiol

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The plant-pathogenic bacterium Xylella fastidiosa is restricted to the xylem vessel environment, where mineral nutrients are transported through the plant host; therefore, changes in the concentrations of these elements likely impact the growth and virulence of this bacterium. Twitching motility, dependent on type IV pili (TFP), is required for movement against the transpiration stream that results in basipetal colonization. We previously demonstrated that calcium (Ca) increases the motility of X. fastidiosa, although the mechanism was unknown. PilY1 is a TFP structural protein recently shown to bind Ca and to regulate twitching and adhesion in bacterial pathogens of humans. Sequence analysis identified three pilY1 homologs in X. fastidiosa (PD0023, PD0502, and PD1611), one of which (PD1611) contains a Ca-binding motif. Separate deletions of PD0023 and PD1611 resulted in mutants that still showed twitching motility and were not impaired in attachment or biofilm formation. However, the response of increased twitching at higher Ca concentrations was lost in the pilY1-1611 mutant. Ca does not modulate the expression of any of the X. fastidiosa PilY1 homologs, although it increases the expression of the retraction ATPase pilT during active movement. The evidence presented here suggests functional differences between the PilY1 homologs, which may provide X. fastidiosa with an adaptive advantage in environments with high Ca concentrations, such as xylem sap.

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Section: Discussionmentioning

confidence: 75%

Section: Discussionmentioning

confidence: 99%

Calcium-Enhanced Twitching Motility in Xylella fastidiosa Is Linked to a Single PilY1 Homolog

Cruz

Parker

Cobine

et al. 2014

Appl Environ Microbiol

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“…Strains with pilE deletions will not amplify with the primers used and will not be present in our analysis. To determine the contribution of slipped-strand phase variation, we included a phaselocked pilC mutant (54).…”

Section: Resultsmentioning

confidence: 99%

Analyzing Neisseria gonorrhoeae Pilin Antigenic Variation Using 454 Sequencing Technology

Rotman

Webber²,

Seifert

2016

J Bacteriol

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Many pathogens use homologous recombination to vary surface antigens in order to avoid immune surveillance. Neisseria gonorrhoeae, the bacterium responsible for the sexually transmitted infection gonorrhea, achieves this in part by changing the sequence of the major subunit of the type IV pilus in a process termed pilin antigenic variation (Av). The N. gonorrhoeae chromosome contains one expression locus (pilE) and many promoterless, partial-coding silent copies (pilS) that act as reservoirs for variant pilin information. Pilin Av occurs by high-frequency gene conversion reactions, which transfer pilS sequences into the pilE locus. We have developed a 454 sequencing-based assay to analyze the frequency and characteristics of pilin Av that allows a more robust analysis of pilin Av than previous assays. We used this assay to analyze mutations and conditions previously shown to affect pilin Av, confirming many but not all of the previously reported phenotypes. We show that mutations or conditions that cause growth defects can result in Av phenotypes when analyzed by phase variation-based assays. Adapting the 454 sequencing to analyze pilin Av demonstrates the utility of this technology to analyze any diversity generation system that uses recombination to develop biological diversity. IMPORTANCEMeasuring and analyzing complex recombination-based systems constitute a major barrier to understanding the mechanisms used to generate diversity. We have analyzed the contributions of many gonococcal mutations or conditions to the process of pilin antigenic variation.

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“…To reduce the background level of nonpiliated cells, the mutS alleles were placed into a strain where every third G of the pilC1 gene has been changed to a synonymous substitution to make PilC production constitutive (the phase-locked pilC1 PL allele) (47). In the phase-locked pilC strain, very few blebs emerge without IPTG, even in the MMC mutants.…”

Section: ϫ6mentioning

confidence: 99%

Neisseria gonorrhoeae MutS Affects Pilin Antigenic Variation through Mismatch Correction and Not by pilE Guanine Quartet Binding

Rotman

Seifert

2015

J Bacteriol

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Many pathogens use homologous recombination to vary surface antigens to avoid immune surveillance. Neisseria gonorrhoeae achieves this in part by changing the properties of its surface pili in a process called pilin antigenic variation (AV). Pilin AV occurs by high-frequency gene conversion reactions that transfer silent pilS sequences into the expressed pilE locus and requires the formation of an upstream guanine quartet (G4) DNA structure to initiate this process. The MutS and MutL proteins of the mismatch correction (MMC) system act to correct mismatches after replication and prevent homeologous (i.e., partially homologous) recombination, but MutS orthologs can also bind to G4 structures. A previous study showed that mutation of MutS resulted in a 3-fold increase in pilin AV, which could be due to the loss of MutS antirecombination properties or loss of G4 binding. We tested two site-directed separation-of-function MutS mutants that are both predicted to bind to G4s but are not able to perform MMC. Pilus phase variation assays and DNA sequence analysis of pilE variants produced in these mutants showed that all three mutS mutants and a mutL mutant had similar increased frequencies of pilin AV. Moreover, the mutS mutants all showed similar increased levels of pilin AV-dependent synthetic lethality. These results show that antirecombination by MMC is the reason for the effect that MutS has on pilin AV and is not due to pilE G4 binding by MutS. IMPORTANCENeisseria gonorrhoeae continually changes its outer surface proteins to avoid recognition by the immune system. N. gonorrhoeae alters the antigenicity of the pilus by directed recombination between partially homologous pilin copies in a process that requires a guanine quartet (G4) structure. The MutS protein of the mismatch correction (MMC) system prevents recombination between partially homologous sequences and can also bind to G4s. We confirmed that loss of MMC increases the frequency of pilin antigenic variation and that two MutS mutants that are predicted to separate the two different functions of MutS inhibit pilin variation similarly to a complete-loss-of-function mutant, suggesting that interaction of MutS with the G4 structure is not a major factor in this process. N eisseria gonorrhoeae is the sole causative agent of gonorrhea, the second most commonly reported sexually transmitted infection in the United States, with an estimated 800,000 new cases per year (1). N. gonorrhoeae extensively uses phase and antigenic variation to provide a reversible subpopulation of genetic variants that can be selected for during infection (2). The use of various surface antigens is one of the most effective strategies used by pathogens to evade immune surveillance. By changing outer surface components, N. gonorrhoeae can avoid recognition by the adaptive immune system, which can prolong a current infection and enable reinfection. These diversity generation mechanisms can also provide functional changes for N. gonorrhoeae.It is estimated that there are over 100 pha...

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Mutation of the Conserved Calcium-Binding Motif in Neisseria gonorrhoeae PilC1 Impacts Adhesion but Not Piliation

Cited by 27 publications

References 39 publications

Calcium-Enhanced Twitching Motility in Xylella fastidiosa Is Linked to a Single PilY1 Homolog

Calcium-Enhanced Twitching Motility in Xylella fastidiosa Is Linked to a Single PilY1 Homolog

Analyzing Neisseria gonorrhoeae Pilin Antigenic Variation Using 454 Sequencing Technology

Neisseria gonorrhoeae MutS Affects Pilin Antigenic Variation through Mismatch Correction and Not by pilE Guanine Quartet Binding

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