This study demonstrates the dynamic interaction between S. aureus and the infected host, where φSa3 serves as a regulator of virulence gene expression, and increased fitness and virulence in new environments.
BackgroundSuperantigens are indispensable virulence factors for Staphylococcus aureus in disease causation. Superantigens stimulate massive immune cell activation, leading to toxic shock syndrome (TSS) and contributing to other illnesses. However, superantigens differ in their capacities to induce body-wide effects. For many, their production, at least as tested in vitro, is not high enough to reach the circulation, or the proteins are not efficient in crossing epithelial and endothelial barriers, thus remaining within tissues or localized on mucosal surfaces where they exert only local effects. In this study, we address the role of TSS toxin-1 (TSST-1) and most importantly the enterotoxin gene cluster (egc) in infective endocarditis and sepsis, gaining insights into the body-wide versus local effects of superantigens.MethodsWe examined S. aureus TSST-1 gene (tstH) and egc deletion strains in the rabbit model of infective endocarditis and sepsis. Importantly, we also assessed the ability of commercial human intravenous immunoglobulin (IVIG) plus vancomycin to alter the course of infective endocarditis and sepsis.ResultsTSST-1 contributed to infective endocarditis vegetations and lethal sepsis, while superantigens of the egc, a cluster with uncharacterized functions in S. aureus infections, promoted vegetation formation in infective endocarditis. IVIG plus vancomycin prevented lethality and stroke development in infective endocarditis and sepsis.ConclusionsOur studies support the local tissue effects of egc superantigens for establishment and progression of infective endocarditis providing evidence for their role in life-threatening illnesses. In contrast, TSST-1 contributes to both infective endocarditis and lethal sepsis. IVIG may be a useful adjunct therapy for infective endocarditis and sepsis.
The Ras-ERK (extracellular signal–regulated kinase) pathway is critical for controlling cell proliferation, and its aberrant activation drives the growth of various cancers. Because many pathogens produce toxins that inhibit Ras activity, efforts to develop effective Ras inhibitors for treating cancer could be informed by studies of Ras inhibition by pathogens. Vibrio vulnificus causes fatal infections in a manner that depends upon multifunctional-autoprocessing repeats-in-toxin (MARTX), a toxin that releases bacterial effector domains into host cells. One of the effector domains delivered by this toxin is the Ras/Rap1-specific endopeptidase (RRSP), which site-specifically cleaves the Switch I domain of the small GTPases Ras and Rap1. We solved the crystal structure of RRSP and found that its backbone shares a structural fold with the EreA/ChaN-like superfamily of enzymes. Unlike other proteases in this family, RRSP is not a metalloprotease. Through nuclear magnetic resonance (NMR) analysis and nucleotide exchange assays, we determined that RRSP processing did not release any fragments of KRAS or cause KRAS to dissociate from its bound nucleotide, but instead only locally impacted the structure. However, this structural alteration of KRAS was sufficient to disable guanine nucleotide exchange factor (GEF)-mediated nucleotide exchange and RAF binding. Thus, RRSP is a bacterial effector that represents a previously unrecognized class of protease that disconnects Ras from its signaling network while inducing limited structural disturbance to its target.
β-toxin is an important virulence factor of Staphylococcus aureus, contributing to colonization and development of disease.1–3 This cytotoxin has two distinct mechanisms of action: sphingomyelinase activity and DNA biofilm ligase activity. However, the distinct mechanism which is important for its role in infective endocarditis is unknown. We characterized the active site of β-toxin DNA biofilm ligase activity by examining deficiencies in site-directed mutants through in vitro DNA precipitation and biofilm formation assays. Possible conformational changes in mutant structure compared to wild type toxin were assessed via trypsin digestion analysis, retention of sphingomyelinase activity, and predicted structures based on the native toxin structure. We addressed the contribution of each mechanism of action in producing infective endocarditis in vivo in a rabbit model of infective endocarditis and sepsis. The H289N β-toxin S. aureus mutant, lacking sphingomyelinase activity, was decreased in lethality and infective endocarditis vegetation formation compared to wild type protein. β-toxin mutants disrupted in biofilm ligase activity did not decrease S. aureus lethality, but were deficient in infective endocarditis vegetation formation compared to wild type protein. Our study begins to characterize the DNA biofilm ligase active site of β-toxin and suggests β-toxin functions importantly in infective endocarditis through both of its mechanisms of action.
Studies have successfully elucidated the mechanism of action of several effector domains that comprise the multifunctional-autoprocessing repeats-in-toxins (MARTX) toxins of Vibrio vulnificus. However, the biochemical linkage between the cysteine proteolytic activity of Makes Caterpillars Floppy (MCF)-like effector and its cellular effects remains unknown. In this study, we identify the host cell factors that activate in vivo and in vitro MCF autoprocessing as adenosine diphosphate (ADP)-Ribosylation Factor 1 (ARF1) and ADP-Ribosylation Factor 3 (ARF3).Autoprocessing activity is enhanced when ARF1 is in its active [guanosine triphosphate (GTP)-bound] form compared to the inactive [guanosine diphosphate (GDP)bound] form. Subsequent to auto-cleavage, MCF is acetylated on its exposed N-terminal glycine residue. Acetylation apparently does not dictate subcellular localization as MCF is found localized throughout the cell. However, the cleaved form of MCF gains the ability to bind to the specialized lipid phosphatidylinositol 5-phosphate enriched in Golgi and other membranes necessary for endocytic trafficking, suggesting that a fraction of MCF may be subcellularly localized. Traditional thin-section electron microscopy, high-resolution cryoAPEX localization, and fluorescent microscopy show that MCF causes Golgi dispersal resulting in extensive vesiculation. In addition, host mitochondria are disrupted and fragmented. Mass spectrometry analysis found no reproducible modifications of ARF1 suggesting that ARF1 is not posttranslationally modified by MCF. Further, catalytically active MCF does not stably associate with ARF1. Our data indicate not only that ARF1 is a cross-kingdom activator of MCF, but also that MCF may mediate cytotoxicity by directly targeting another yet to be identified protein. This study begins to elucidate the biochemical activity of this important domain and gives insight into how it may promote disease progression.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.