Rekrutierung von Komplementregulatoren der Faktor H Proteinfamilie als Mechanismen der Immunevasion humanpathogener Erreger

Das Komplementsystem ist ein wesentlicher Bestandteil der angeborenen Immunität des Menschen. In den Organismus eingedrungene Mikroorganismen werden vom Komplementsystem als „fremd“ erkannt und durch Opsonisierung, Phagozytose oder Lyse eliminiert. Die Aktivierung dieses Effektorsystems erfolgt über drei verschiedene Wege, den alternativen Weg, den klassischen Weg und den Lektin-Weg und führt über den terminalen Weg zur Ausbildung des Membranangriffskomplexes. Um gesunde körpereigene Zellen zu schützen, ist die Aktivierung des Komplementsystems durch Membran gebundene oder im Plasma gelöste Regulatorproteine streng reguliert. Der wichtigste Komplementregulator des alternativen Komplementwegs ist Faktor H. Das Plasmaprotein inhibiert die Komplementkaskade sowohl im Plasma als auch auf körpereigenen Oberflächen, indem es als Kofaktor für die Faktor I vermittelte Spaltung von C3b agiert und den Zerfall der C3-Konvertase des alternativen Wegs, C3bBb, beschleunigt. Pathogene nutzen die Regulationsmechanismen des Komplementsystems zum eigenen Vorteil, um sich vor dem Angriff des Komplementsystems zu schützen, indem sie Wirtsregulatoren, wie z.B. Faktor H und FHL-1 rekrutieren und an Oberflächenproteine binden. Die Komplement inhibitorischeFunktion von Faktor H verhindert die Aktivierung der Komplementkaskade auf der Erregeroberfläche und das Pathogen überlebt

The Fungus Among Us: Why the Treatment of Fungal Infections Is So Problematic

When we think of microbes that can make us sick, it is usually bacteria that cross our minds first. We tend to forget about another major microbial type that can also cause severe diseases: the fungi. Yeasts and molds make up the majority of microscopic fungi and both types can cause various infections in humans, from mild skin rashes to deadly blood infections. These fungi have found several ways to cause us harm, such as using the body’s nutrients, escaping the surveillance of the immune system, or hijacking and destroying our cells. On cellular level, we have a lot in common with fungi. These common features between human cells and fungal cells makes the development of antibiotics and vaccines to treat fungal infections very difficult. In this article, we will describe some fungal infections and explain current options for their treatment

The Staphylococcus aureus Protein Sbi Acts as a Complement Inhibitor and Forms a Tripartite Complex with Host Complement Factor H and C3b

The Gram-positive bacterium Staphylococcus aureus, similar to other pathogens, binds human complement regulators Factor H and Factor H related protein 1 (FHR-1) from human serum. Here we identify the secreted protein Sbi (Staphylococcus aureus binder of IgG) as a ligand that interacts with Factor H by a—to our knowledge—new type of interaction. Factor H binds to Sbi in combination with C3b or C3d, and forms tripartite Sbi∶C3∶Factor H complexes. Apparently, the type of C3 influences the stability of the complex; surface plasmon resonance studies revealed a higher stability of C3d complexed to Sbi, as compared to C3b or C3. As part of this tripartite complex, Factor H is functionally active and displays complement regulatory activity. Sbi, by recruiting Factor H and C3b, acts as a potent complement inhibitor, and inhibits alternative pathway-mediated lyses of rabbit erythrocytes by human serum and sera of other species. Thus, Sbi is a multifunctional bacterial protein, which binds host complement components Factor H and C3 as well as IgG and β2-glycoprotein I and interferes with innate immune recognition

Generation of a KLF15 homozygous knockout human embryonic stem cell line using paired CRISPR/Cas9n, and human cardiomyocytes derivation

Krueppel-like factor 15 (KLF15) is abundantly expressed in liver, kidney, and muscle, including myocardium. In the adult heart KLF15 is important to maintain homeostasis and to repress hypertrophic remodeling. We generated a homozygous hESC KLF15 knockout (KO) line using paired CRISPR/Cas9n. KLF15-KO cells maintained full pluripotency and differentiation potential as well as genomic integrity. We demonstrated that KLF15-KO cells can be differentiated into morphologically normal cardiomyocytes turning them into a valuable tool for studying human KLF15-mediated mechanisms resulting in human cardiac dysfunction

The FKBP-Type Domain of the Human Aryl Hydrocarbon Receptor-Interacting Protein Reveals an Unusual Hsp90 Interaction

The aryl hydrocarbon receptor-interacting protein (AIP) has been predicted to consist of an N-terminal FKBP-type peptidyl-prolyl <i>cis</i>/<i>trans</i> isomerase (PPIase) domain and a C-terminal tetratricopeptide repeat (TPR) domain, as typically found in FK506-binding immunophilins. AIP, however, exhibited no inherent FK506 binding or PPIase activity. Alignment with the prototypic FKBP12 showed a high sequence homology but indicated inconsistencies with regard to the secondary structure prediction derived from chemical shift analysis of AIP^2–166. NMR-based structure determination of AIP^2–166 now revealed a typical FKBP fold with five antiparallel β-strands forming a half β-barrel wrapped around a central α-helix, thus permitting AIP to be also named FKBP37.7 according to FKBP nomenclature. This PPIase domain, however, features two structure elements that are unusual for FKBPs: (i) an N-terminal α-helix, which additionally stabilizes the domain, and (ii) a rather long insert, which connects the last two β-strands and covers the putative active site. Diminution of the latter insert did not generate PPIase activity or FK506 binding capability, indicating that the lack of catalytic activity in AIP is the result of structural differences within the PPIase domain. Compared to active FKBPs, a diverging conformation of the loop connecting β-strand C′ and the central α-helix apparently is responsible for this inherent lack of catalytic activity in AIP. Moreover, Hsp90 was identified as potential physiological interaction partner of AIP, which revealed binding contacts not only at the TPR domain but uncommonly also at the PPIase domain

A High-Throughput Method as a Diagnostic Tool for HIV Detection in Patient-Specific Induced Pluripotent Stem Cells Generated by Different Reprogramming Methods

Induced pluripotent stem cells (iPSCs) provide a unique opportunity for generation of patient-specific cells for use in translational purposes. We aimed to compare iPSCs generated by different reprogramming methods regarding their reprogramming efficiency, pluripotency capacity, and the possibility to use high-throughput PCR-based methods for detection of human pathogenic viruses. iPSCs from skin fibroblasts (FB), peripheral blood mononuclear cells (PBMCs), or mesenchymal stem cells (MSCs) were generated by using three different reprogramming systems including chromosomal integrating and nonintegrating methods. Reprogramming efficiencies were in accordance with the literature, indicating that the parental cell type and the reprogramming method play a major role for the reprogramming efficiencies (FB: STEMCCA: 1.30 +/- 0.18, Sendai virus: 1.37 +/- 0.01, and episomal plasmids: 0.04 +/- 0.02; PBMCs: Sendai virus: 0.002 +/- 0.001, episomal plasmids: 0) but result in the same characteristics of pluripotency. We found the highest reprogramming efficiencies for MSC with 3.32 +/- 1.2 by using episomal plasmids. Since GMP standard working procedures and screening units need virus contamination-free cell lines, we studied HIV-1 contamination in the generated iPSCs. We used the high-throughput cobas (R) 6800/8800 system, which is normally used for detection of HIV-1 in plasma of patients, and found that footprint-free reprogramming methods as episomal plasmids and Sendai virus are useful for the described virus detection method. This fast, cost-effective, robust, and reliable assay demonstrates the feasibility to use high-throughput PCR-based methods for detection of human pathogenic viruses in ps-iPSC lines that were generated with nongenome integrating reprogramming methods