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Proline transporters ProT and PutP are required for Staphylococcus aureus infection.

ABSTRACT: Proline acquired via specific transporters can serve as a proteinogenic substrate, carbon and nitrogen source, or osmolyte. Previous reports have documented that Staphylococcus aureus, a major community and nosocomial pathogen, encodes at least four proline transporters, PutP, OpuC, OpuD, and ProP. A combination of genetic approaches and 3H-proline transport assays reveal that a previously unrecognized transporter, ProT, in addition to PutP, are the major proline transporters in S. aureus. Complementation experiments using constitutively expressed non-cognate promoters found that proline transport via OpuD, OpuC, and ProP is minimal. Both proline biosynthesis from arginine and proline transport via ProT are critical for growth when S. aureus is grown under conditions of high salinity. Further, proline transport mediated by ProT or PutP are required for growth in media with and without glucose, indicating both transporters function to acquire proline for proteinogenic purposes in addition to acquisition of proline as a carbon/nitrogen source. Lastly, inactivation of proT and putP resulted in a significant reduction (5 log10) of bacterial burden in murine skin-and-soft tissue infection and bacteremia models, suggesting that proline transport is required to establish a S. aureus infection.

SUBMITTER: Lehman MK

PROVIDER: S-EPMC9886301 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Proline transporters ProT and PutP are required for Staphylococcus aureus infection.

Lehman McKenzie K MK Sturd Natalie A NA Razvi Fareha F Wellems Dianne L DL Carson Steven D SD Fey Paul D PD

PLoS pathogens 20230118 1

Proline acquired via specific transporters can serve as a proteinogenic substrate, carbon and nitrogen source, or osmolyte. Previous reports have documented that Staphylococcus aureus, a major community and nosocomial pathogen, encodes at least four proline transporters, PutP, OpuC, OpuD, and ProP. A combination of genetic approaches and 3H-proline transport assays reveal that a previously unrecognized transporter, ProT, in addition to PutP, are the major proline transporters in S. aureus. Compl ...[more]

PMID: 36652494

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Project description:Staphylococcus aureus causes a wide variety of invasive human infections. However, delineation of the genes which are essential for the in vivo survival of this pathogen has not been accomplished to date. Using signature tag mutagenesis techniques and large mutant pool screens, previous investigators identified several major gene classes as candidate essential gene loci for in vivo survival; these include genes for amino acid transporters, oligopeptide transporters, and lantibiotic synthesis (W. R. Schwan, S. N. Coulter, E. Y. W. Ng, M. H. Langhorne, H. D. Ritchie, L. L. Brody, S. Westbrock-Wadman, A. S. Bayer, K. R. Folger, and C. K. Stover, Infect. Immun. 66:567-572, 1998). In this study, we directly compared the virulence of four such isogenic signature tag mutants with that of the parental strain (RN6390) by using a prototypical model of invasive S. aureus infection, experimental endocarditis (IE). The oligonucleotide signature tag (OST) mutant with insertional inactivation of the gene (putP) which encodes the high-affinity transporter for proline uptake exhibited significantly reduced virulence in the IE model across three challenge inocula (10(4) to 10(6) CFU) in terms of achievable intravegetation densities (P, <0.05). The negative impact of putP inactivation on in vivo survival in the IE model was confirmed by simultaneous challenge with the original putP mutant and the parental strain as well as by challenge with a putP mutant in which this genetic inactivation was transduced into a distinct parental strain (S6C). In contrast, inactivation of loci encoding an oligopeptide transporter, a purine repressor, and lantibiotic biosynthesis had no substantial impact on the capacity of OST mutants to survive within IE vegetations. Thus, genes encoding the uptake of essential amino acids may well represent novel targets for new drug development. These data also confirm the utility of the OST technique as an important screening methodology for identifying candidate genes as requisite loci for the in vivo survival of S. aureus.

Project description:Staphylococcus aureus is a human commensal that can also cause systemic infections. This transition requires evasion of the immune response and the ability to exploit different niches within the host. However, the disease mechanisms and the dominant immune mediators against infection are poorly understood. Previously it has been shown that the infecting S. aureus population goes through a population bottleneck, from which very few bacteria escape to establish the abscesses that are characteristic of many infections. Here we examine the host factors underlying the population bottleneck and subsequent clonal expansion in S. aureus infection models, to identify underpinning principles of infection. The bottleneck is a common feature between models and is independent of S. aureus strain. Interestingly, the high doses of S. aureus required for the widely used "survival" model results in a reduced population bottleneck, suggesting that host defences have been simply overloaded. This brings into question the applicability of the survival model. Depletion of immune mediators revealed key breakpoints and the dynamics of systemic infection. Loss of macrophages, including the liver Kupffer cells, led to increased sensitivity to infection as expected but also loss of the population bottleneck and the spread to other organs still occurred. Conversely, neutrophil depletion led to greater susceptibility to disease but with a concomitant maintenance of the bottleneck and lack of systemic spread. We also used a novel microscopy approach to examine abscess architecture and distribution within organs. From these observations we developed a conceptual model for S. aureus disease from initial infection to mature abscess. This work highlights the need to understand the complexities of the infectious process to be able to assign functions for host and bacterial components, and why S. aureus disease requires a seemingly high infectious dose and how interventions such as a vaccine may be more rationally developed.

Dataset Information

Proline transporters ProT and PutP are required for Staphylococcus aureus infection.

Publications

Proline transporters ProT and PutP are required for Staphylococcus aureus infection.

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