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Efb is K27-ubiquitinated at K71 by host RNF114 a Immunoblots of IP products of WCL from MH-S infected with ΔEfb + Flag − Efb for indicated times (MOI = 2 5). b, c Immunoblots of WCL and IP products from WCL of HEK 293 T cells transfected with indicated plasmids. d Immunoblots of IP products of WCL from MH-S infected with ΔEfb + Flag − Efb for indicated times (MOI = 25). e Immunoblots of WCL and IP products from HEK293T cells transfected with indicated plasmids. f Immunoblots of WCL and IP products from MH-S or RNF114 knockdown (KD) MH-S infected with ΔEfb + Flag − Efb for indicated times (MOI = 25). g Immunoblots of WCL and IP products from HEK293T cells transfected with indicated plasmids. K71R, replace 71 lysine of Efb with arginine. h Immunoblots of WCL and IP products from MH-S infected with ΔEfb + Flag − Efb or ΔEfb + Flag − Efb K71R for indicated times (MOI = 25). Data are representative of three experiments with at least three independent biological replicates. Source data are provided as a Source Data file.
Source publication
Many pathogens secrete effectors to hijack intracellular signaling regulators in host immune cells to promote pathogenesis. However, the pathogenesis of Staphylococcus aureus secretory effectors within host cells is unclear. Here, we report that Staphylococcus aureus secretes extracellular fibrinogen-binding protein (Efb) into the cytoplasm of macr...
Citations
... Several studies have revealed that STING serves as a dominant regulatory molecule during S. aureus infection, exerting conflicting effects in different infection models (10,11). Our previous study found that tumor necrosis factor α (TNF-α), interleukin (IL)-1β, IL-6, and IL-12 secreted by macrophages hinder S. aureus colonization in the early stages of infection (12). This study found that STING is also involved in this process. ...
... EsxB Interacts with STING. Secretory effectors released by S. aureus can manipulate the activation of macrophages by interfering with intracellular signal transduction (12). We speculated that the STING-mediated proinflammatory responses of macrophages may counteract S. aureus infection. ...
... The STING pathway is a critical pathway that plays a vital role in regulating the inflammatory responses of macrophages during microbial infections (5). Several studies have identified STING as a dominant regulatory molecule during S. aureus infection (10,12). However, the role of STING in S. aureus infection remains contro versial. ...
Staphylococcus aureus ( S. aureus ) can evade antibiotics and host immune defenses by persisting within infected cells. Here, we demonstrate that in infected host cells, S. aureus type VII secretion system (T7SS) extracellular protein B (EsxB) interacts with the stimulator of interferon genes (STING) protein and suppresses the inflammatory defense mechanism of macrophages during early infection. The binding of EsxB with STING disrupts the K48-linked ubiquitination of EsxB at lysine 33, thereby preventing EsxB degradation. Furthermore, EsxB-STING binding appears to interrupt the interaction of 2 vital regulatory proteins with STING: aspartate-histidine-histidine-cysteine domain-containing protein 3 (DHHC3) and TNF receptor-associated factor 6. This persistent dual suppression of STING interactions deregulates intracellular proinflammatory pathways in macrophages, inhibiting STING’s palmitoylation at cysteine 91 and its K63-linked ubiquitination at lysine 83. These findings uncover an immune-evasion mechanism by S. aureus T7SS during intracellular macrophage infection, which has implications for developing effective immunomodulators to combat S. aureus infections.
... As mentioned, S. aureus Efb also interacts with Fg and belongs to SERAMs (9,23,27,34). It is reported to inhibit complement activation by engaging C3b (35)(36)(37)(38), block platelet aggregation and their interaction with leukocytes (39,40) and interact with immune cells blocking cellular-mediated immunity (23,27,41,42). Furthermore, Efb can also bind to Complement Receptor 2 on B cells, further tackling adaptive responses of the host (43). ...
Staphylococcus aureus pathology is caused by a plethora of virulence factors able to combat multiple host defence mechanisms. Fibrinogen (Fg), a critical component in the host coagulation cascade, plays an important role in the pathogenesis of this bacterium, as it is the target of numerous staphylococcal virulence proteins. Amongst its secreted virulence factors, coagulase (Coa) and Extracellular fibrinogen-binding protein (Efb) share common Fg binding motives and have been described to form a Fg shield around staphylococcal cells, thereby allowing efficient bacterial spreading, phagocytosis escape and evasion of host immune system responses. Targeting these proteins with monoclonal antibodies thus represents a new therapeutic option against S. aureus. To this end, here we report the selection and characterization of fully human, sequence-defined, monoclonal antibodies selected against the C-terminal of coagulase. Given the functional homology between Coa and Efb, we also investigated if the generated antibodies bound the two virulence factors. Thirteen unique antibodies were isolated from naïve antibodies gene libraries by antibody phage display. As anticipated, most of the selected antibodies showed cross-recognition of these two proteins and among them, four were able to block the interaction between Coa/Efb and Fg. Furthermore, our monoclonal antibodies could interact with the two main Fg binding repeats present at the C-terminal of Coa and distinguish them, suggesting the presence of two functionally different Fg-binding epitopes.
... ECP, also referred to as nonclassical secretion or protein moonlighting, has important implications for how bacterial cells interact with their external environment. Indeed, some of these excreted proteins have been reported to mediate host-pathogen interactions (24), biofilm formation (23), suppress macrophage activation (25), and are linked to survival and pathogenicity (26). Yet, the mechanisms of ECP are poorly understood (27), and it was previously unknown how mscL and arfA collectively mediate ECP in response to stress. ...
Excretion of cytoplasmic protein (ECP) is a commonly observed phenomenon in bacteria, and this partial extracellular localisation of the intracellular proteome has been implicated in a variety of stress response mechanisms. In response to hypoosmotic shock and ribosome stalling in Escherichia coli , ECP is dependent upon the presence of the large-conductance mechanosensitive channel and the alternative ribosome–rescue factor A gene products. However, it is not known if a mechanistic link exists between the corresponding genes and the respective stress response pathways. Here, we report that the corresponding mscL and arfA genes are commonly co-located on the genomes of Gammaproteobacteria and display overlap in their respective 3′ UTR and 3′ CDS. We show this unusual genomic arrangement permits an antisense RNA–mediated regulatory control between mscL and arfA , and this modulates MscL excretory activity in E. coli . These findings highlight a mechanistic link between osmotic, translational stress responses and ECP in E. coli , further elucidating the previously unknown regulatory function of arfA sRNA.
... Recently it was shown that macrophage-ingested S. aureus secretes Efb into the cytoplasm, suppresses the secretion of inflammatory cytokines, and subdues host immunity by engaging with tumor necrosis-associated factor 3 (TRAF3). Ubiquitination of Efb by host ubiquitin ligase is essential for Efb-TRAF3 interaction (Zhang et al., 2022). ...
Microbial virulence showcases an excellent model for adaptive changes that enable an organism to survive and proliferate in a hostile environment and exploit host resources to its own benefit. In Staphylococcus aureus, an opportunistic pathogen of the human host, known for the diversity of the disease conditions it inflicts and the rapid evolution of antibiotic resistance, virulence is a consequence of having a highly plastic genome that is amenable to quick reprogramming and the ability to express a diverse arsenal of virulence factors. Virulence factors that are secreted to the host milieu effectively manipulate the host conditions to favor bacterial survival and growth. They assist in colonization, nutrient acquisition, immune evasion, and systemic spread. The structural and functional characteristics of the secreted virulence proteins have been shaped to assist S. aureus in thriving and disseminating effectively within the host environment and exploiting the host resources to its best benefit. With the aim of highlighting the importance of secreted virulence proteins in bacterial virulence, the present chapter provides a comprehensive account of the role of the major secreted proteins of S. aureus in orchestrating its virulence in the human host.
