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Perturbing the acetylation status of the Type IV pilus retraction motor, PilT, reduces Neisseria gonorrhoeae viability.

ABSTRACT: Post-translational acetylation is a common protein modification in bacteria. It was recently reported that Neisseria gonorrhoeae acetylates the Type IV pilus retraction motor, PilT. Here, we show recombinant PilT can be acetylated in vitro and acetylation does not affect PilT ultrastructure. To investigate the function of PilT acetylation, we mutated an acetylated lysine, K117, to mimic its acetylated or unacetylated forms. These mutations were not tolerated by wild-type N. gonorrhoeae, but they were tolerated by N. gonorrhoeae carrying an inducible pilE when grown without inducer. We identified additional mutations in pilT and pilU that suppress the lethality of K117 mutations. To investigate the link between PilE and PilT acetylation, we found the lack of PilE decreases PilT acetylation levels and increases the amount of PilT associated with the inner membrane. Finally, we found no difference between wild-type and mutant cells in transformation efficiency, suggesting neither mutation inhibits Type IV pilus retraction. Mutant cells, however, form microcolonies morphologically distinct from wt cells. We conclude that interfering with the acetylation status of PilTK117 greatly reduces N. gonorrhoeae viability, and mutations in pilT, pilU and pilE can overcome this lethality. We discuss the implications of these findings in the context of Type IV pilus retraction regulation.

SUBMITTER: Hockenberry AM

PROVIDER: S-EPMC6214798 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Perturbing the acetylation status of the Type IV pilus retraction motor, PilT, reduces Neisseria gonorrhoeae viability.

Hockenberry Alyson M AM Post Deborah M B DMB Rhodes Katherine A KA Apicella Michael M So Magdalene M

Molecular microbiology 20181028 5

Post-translational acetylation is a common protein modification in bacteria. It was recently reported that Neisseria gonorrhoeae acetylates the Type IV pilus retraction motor, PilT. Here, we show recombinant PilT can be acetylated in vitro and acetylation does not affect PilT ultrastructure. To investigate the function of PilT acetylation, we mutated an acetylated lysine, K117, to mimic its acetylated or unacetylated forms. These mutations were not tolerated by wild-type N. gonorrhoeae, but they ...[more]

PMID: 29719082

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Project description:Retraction of the type IV pilus (Tfp) mediates DNA uptake, motility, and social and infection behavior in a wide variety of prokaryotes. To date, investigations into Tfp retraction-dependent activities have used a mutant deleted of PilT, the ATPase motor protein that causes the pilus fiber to retract. ΔpilT cells are nontransformable, nonmotile, and cannot aggregate into microcolonies. We tested the hypothesis that these retraction-dependent activities are sensitive to the strength of PilT enzymatic activity by using the pathogen Neisseria gonorrhoeae as a model. We constructed an N. gonorrhoeae mutant with an amino acid substitution in the PilT Walker B box (a substitution of cysteine for leucine at position 201, encoded by pilTL201C). Purified PilTL201C forms a native hexamer, but mutant hexamers hydrolyze ATP at half the maximal rate. N. gonorrhoeae pilTL201C cells produce Tfp fibers, crawl at the same speed as the wild-type (wt) parent, and are equally transformable. However, the social behavior of pilTL201C cells is intermediate between the behaviors of wt and ΔpilT cells. The infection behavior of pilTL201C is also defective, due to its failure to activate the epidermal growth factor receptor (EGFR)-heparin-binding EGF-like growth factor (HB-EGF) pathway. Our study indicates that pilus retraction, per se, is not sufficient for N. gonorrhoeae microcolony formation or infectivity; rather, these activities are sensitive to the strength of PilT enzymatic activity. We discuss the implications of these findings for Neisseria pathogenesis in the context of mechanobiology. Type IV pili are fibers expressed on the surface of many bacteria. Neisseria gonorrhoeae cells crawl, take up DNA, and communicate with each other and with human cells by retracting these fibers. Here, we show that an N. gonorrhoeae mutant expressing an enzymatically weakened type IV pilus retraction motor still crawls and takes up DNA normally. However, mutant cells exhibit abnormal social behavior, and they are less infective because they fail to activate the epidermal growth factor receptor. Our study shows that N. gonorrhoeae social and infection behaviors are sensitive to the strength of the retraction motor enzyme.

Dataset Information

Perturbing the acetylation status of the Type IV pilus retraction motor, PilT, reduces Neisseria gonorrhoeae viability.

Publications

Perturbing the acetylation status of the Type IV pilus retraction motor, PilT, reduces Neisseria gonorrhoeae viability.

Similar Datasets

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