Biology subjects

Paprotka, K.

Publications and source records attributed to Paprotka, K..

5 recordsLinked to original sources

bioRxiv2024

Identification of the Staphylococcus aureus Endothelial Cell Surface Interactome by Proximity Labeling

Virulence strategies of pathogens depend on interaction with host cells. Binding and activation of receptors located on the plasma membrane is crucial for the attachment to or pathogen internalization by host cells. Identification of host cell receptors often is difficult and hence the identity of many proteins that play important roles during host-pathogen interaction remains elusive. We developed a novel proximity labeling approach, in which we decorated the opportunistic pathogen Staphylococcus aureus with ascorbate peroxidase 2. Upon addition of hydrogen peroxide the peroxidase initiates proximity biotinylation of S. aureus host receptors, thereby enabling identification of these proteins. We here demonstrate an endothelial cell surface interactome of 306 proteins of which neuronal adhesion molecule, protein tyrosine kinase PTK7, melanotransferrin, protein-tyrosine kinase Met, CD109 and others constitute novel S. aureus co-receptors. Filtering the interactome for validated surface proteins resulted in a list of 89 receptor candidates of which a 53% were described to interact with S. aureus or other pathogens.

microbiology↗

bioRxiv2024

A Novel Rapid Host Cell Entry Pathway Determines Intracellular Fate of Staphylococcus aureus

Staphylococcus aureus is an opportunistic pathogen causing severe diseases. Recently, S. aureus was recognized as intracellular pathogen, whereby the intracellular niche promotes immune evasion and antibiotic resistance. Interaction of S. aureus with versatile host cell receptors was described previously, suggesting that internalization of the pathogen can occur via several pathways. It remains elusive whether the pathway of internalization can affect the intracellular fate of the bacteria. Here, we identified a mechanism governing cellular uptake of S. aureus which relies on lysosomal Ca2+, lysosomal exocytosis and occurs concurrently to other well-known entry pathways within the same host cell population. This internalization pathway is rapid and active within only few minutes after bacterial contact with host cells. Compared to slow bacterial internalization, the rapid pathway demonstrates altered phagosomal maturation as well as translocation of the pathogen to the host cytosol and ultimately results in different rates of intracellular bacterial replication and host cell death. We show that these alternative infection outcomes are caused by the mode of bacterial uptake.

microbiology↗

bioRxiv2024

Infection of human organoids supports an intestinal niche for Chlamydia trachomatis

Several reports suggest that intestinal tissue may be a natural niche for Chlamydia trachomatis infection and a reservoir for persistent infections in the human body. Due to the human specificity of the pathogen and the lack of suitable host models, there is limited knowledge on this topic. In our study, we modelled the course of the chlamydial infection in human primary gastrointestinal (GI) epithelial cells originating from patient-derived organoids. We show that GI cells are resistant to apical infection and C. trachomatis needs access to the basolateral membrane to establish an infection. Transmission electron microscopy analysis reveals the presence of both normal as well as aberrant chlamydial developmental forms in the infected cells, suggesting a possible cell-type specific nature of the infection. Furthermore, we show that the plasmid-encoded Pgp3 is an important virulence factor for the infection of human GI cells. This is the first report of C. trachomatis infection in human primary intestinal epithelial cells supporting a possible niche for chlamydial infection in the human intestinal tissue. Author summaryChlamydial infection has a high global prevalence and is a major health concern. Untreated infections may cause complications and lead to serious health problems, especially in women. Although the infection is usually localized to the genital tract, experiments performed in a mouse infection model as well as the accumulating clinical data suggest that the human gastrointestinal (GI) tract might represent a hidden infection niche and a source of reinfections. In our study, we used the advantages of the organoid technology to model the chlamydial infection in patient-derived primary GI epithelial cells. We were able to show that these cells are resistant to the infection, however, Chlamydia could utilize a basolateral entry route for efficient infection. Chlamydia form either normal or persistent-like developmental forms in these GI epithelial cells. We also showed the importance of the plasmid-mediated virulence in the infection of human GI cells. The results obtained in the GI infection model replicated phenotypes predicted and expected for Chlamydia human intestinal infection, and therefore support a role of the human GI tract as a potential niche for chlamydial infection.

microbiology↗

bioRxiv2021

Staphylococcus aureus α-toxin induces acid sphingomyelinase release from a human endothelial cell line

Staphylococcus aureus (S. aureus) is well known to express a plethora of toxins of which the pore-forming hemolysin A (-toxin) is the best-studied cytolysin. Pore-forming toxins (PFT) permeabilize host membranes during infection thereby causing concentration-dependent effects in host cell membranes ranging from disordered ion fluxes to cytolysis. Host cells possess defense mechanisms against PFT attack, resulting in endocytosis of the breached membrane area and delivery of repair vesicles to the insulted plasma membrane as well as a concurrent release of membrane repair enzymes. Since PFTs from several pathogens have been shown to recruit membrane repair components, we here investigated whether staphylococcal -toxin is able to induce these mechanisms in endothelial cells. We show that S. aureus -toxin induced increase in cytosolic Ca2+ in endothelial cells, which was accompanied by p38 MAPK phosphorylation. Toxin challenge led to increased endocytosis of an extracellular fluid phase marker as well as increased externalization of LAMP1-positive membranes suggesting that peripheral lysosomes are recruited to the insulted plasma membrane. We further observed that thereby the lysosomal protein acid sphingomyelinase (ASM) was released into the cell culture medium. Thus, our results show that staphylococcal -toxin triggers mechanisms in endothelial cells, which have been implicated in membrane repair after damage of other cell types by different toxins.

microbiology↗

bioRxiv2020

Intracellular Staphylococcus aureus employs the cysteine protease staphopain A to induce host cell death in epithelial cells

Staphylococcus aureus is a major human pathogen, which can invade and survive in non-professional and professional phagocytes. Intracellularity is thought to contribute to pathogenicity and persistence of the bacterium. Upon internalization by epithelial cells, cytotoxic S. aureus strains can escape from the phagosome, replicate in the cytosol and induce host cell death. Here, we identified a staphylococcal cysteine protease to induce cell death by intracellular S. aureus after translocation into the host cell cytoplasm. We demonstrated that loss of staphopain A function leads to delayed onset of host cell death and prolonged intracellular replication of S. aureus in epithelial cells. Overexpression of staphopain A in a non-cytotoxic strain facilitated intracellular killing of the host cell even in the absence of detectable intracellular replication. Moreover, staphopain A contributed to efficient colonization of the lung in a mouse pneumonia model. Our study suggests that staphopain A is utilized by S. aureus to mediate escape from the host cell and thus contributes to tissue destruction and dissemination of infection. Author SummaryStaphylococcus aureus is a well-known antibiotic-resistant pathogen that emerges in hospital and community settings and can cause a variety of diseases ranging from skin abscesses to lung inflammation and blood poisoning. The bacterium asymptomatically colonizes the upper respiratory tract and skin of about one third of the human population and takes advantage of opportune conditions, like immunodeficiency or breached barriers, to cause infection. Although S. aureus is not regarded as a professional intracellular bacterium, it can be internalized by human cells and subsequently exit the host cells by induction of cell death, which is considered to cause tissue destruction and spread of infection. The bacterial virulence factors and underlying molecular mechanisms involved in the intracellular lifestyle of S. aureus remain largely unknown. We identified a bacterial cysteine protease to contribute to host cell death mediated by intracellular S. aureus. Staphopain A induced killing of the host cell after translocation of the pathogen into the cell cytosol, while bacterial proliferation was not required. Further, the protease enhanced survival of the pathogen during lung infection. These findings reveal a novel, intracellular role for the bacterial protease staphopain A.

microbiology↗