Chalkophore-mediated copper homeostasis in Staphylococcus lugdunensis

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Metal ions are needed as cofactors for many enzymes and are therefore essential for a functional metabolism. However, due to their redox potential, metal ions do also harbour intrinsic toxicity and cytosolic metal homeostasis needs strict control. Metal homeostasis is an important target for human innate immunity. Availability of trace metals such as iron (Fe) is actively limited within the host to reduce pathogen proliferation during infection. In contrast, a targeted increase of copper concentration within phagolysosomes kills bacteria. Bacteria produce several metallophores to improve metal acquisition. Best studied are iron-binding siderophores. However, isolated reports exist describing that in addition to iron, siderophores can bind copper. This can allow detoxification, as compound binding impacts redox characteristics. Additionally, it ensures sufficient Cu-ion supply for metabolic activity. However, the relevance of copper-binding metallophores (chalkophores) for pathogens in the context of colonisation and disease is insufficiently understood. We found that the pathogen S. lugdunensis produces the compound Ulbactin F, and found the molecule to be a chalkophore. The responsible genetic locus encodes biosynthesis systems as well as putative exporters and importers, strongly suggesting that the molecule is important for metal homeostasis. I suggest to characterize this system on the molecular level and to study its physiological relevance. I request funding for two PhD positions to study distinct topics using highly synergistic approaches. The first PhD student will investigate the relevance of Ulbactin F biosynthesis and membrane transport on metal homeostasis of S. lugdunensis. The student will perform elementary analysis of wild type and Ulbactin F deficient strains to confirm the role of the molecule on intracellular metal levels. Additionally, the student will investigate membrane transport of Ulbactin F. We identified an ABC-transporter of the energy coupling factor (ECF) type within the Ulbactin F locus. ECF-transporters are trace nutrient acquisition systems. We will study the Ulbactin F binding properties of the ECF-transporter on the molecular level. Additionally, we will use isogenic mutants and heterologous expression of the ECF-transporter to validate its role in Ulbactin F acquisition. The second PhD student will study the physiological relevance of Ulbactin F. The student will perform in vitro experiments to investigate whether Ulbactin F production provides a growth advantage under copper ion-limiting conditions or if the molecule provides resistance to copper ions. Additionally, the student will study metal dependent regulation of Ulbactin F biosynthesis and use ex vivo and in vivo infection models to investigate whether the molecule represents a virulence factor of S. lugdunensis. Altogether, I am convinced that the suggested experiments will give a complete picture of the biology of the chalkophore Ulbactin F.