EpiCross: Epigenetic Basis and Therapeutic Implications of the Cross-regulation of Arginase1

Background: Salmonella (S.) enterica serotype Typhi and Leishmania (L.) infantum are two worldwide highly prevalent intracellular pathogens cause life-threatening infections and more than 250 000 deaths per year, termed typhoid fever and visceral leishmaniasis, respectively. The control of both diseases relies on the production of the pro-inflammatory cytokines interferon-gamma (IFNg) and tumor necrosis factor (TNF) and on the generation of antimicrobial molecules such as nitric oxide (NO). NO production by inducible NO synthase (iNOS) requires the amino acid L-arginine, which is also a substrate of arginase (Arg) 1. As Arg1 is co-induced during infections, it is capable to deprive iNOS of L-arginine, and thereby promotes the survival of intracellular microorganisms. We previously found that TNF protects from infection with L. major by suppressing Arg1 transcription in myeloid cells which could be traced back to epigenetic effects of the cytokine on histone acetylation which affects the three dimensional structure of chromatin and thus the amplification of the genetic information. The molecular mechanisms that account for this epigenetic regulation are currently unknown. Hypothesis and aims: Based on these results we hypothesize that (1) Arg1 of myeloid cells, induced by microbes is a critical factor that determines the fate of infections with intracellular pathogens and that (2) TNF as well as IFNg protect against NO-sensitive microbes at least in part by unleashing the enzymatic activity of iNOS via epigenetic repression of Arg1. The central goals and methods of our application are the analysis of the functional importance and the molecular and epigenetic mechanisms of Arg1 regulation by TNF or IFNg in visceral leishmaniasis caused by L. infantum, and in typhoid fever due to systemic S. typhimurium infection. These investigations will be performed by a combined approach employing systemic infection models of mice along with a systemic analysis of transcriptional and epigenetic regulation of Arg1 along with their impact on innate immune function and host control of infection. Specific attention will be paid to the analysis of transcriptional and epigenetic regulation of Arg1 employing specific molecular biology and bioinformatics analyses. We will also screen small molecule inhibitor-libraries to identify potentially drugable regulators of the epigenetic suppression of Arg1. Candidates will be functionally validated in vivo. Expected results, novel aspects and outlook: We expect to generate key insights into the mechanisms of host susceptibility and infection control of these two important infectious diseases. The really novel aspect is the structured analysis of epigenetic regulation of innate immune genes and their contribution to the control of infection which has not been investigated thus far. Based on this knowledge we aim to identify modifiers of this epigenetic immune regulation as novel drugs for the treatment of these infections.

This project evaluated how metabolic changes in amino acid pathways affect the host responses to infection. Specifically, we analysed metabolic responses and associated anti-microbial effector pattern of macrophages, innate immune cells which can contain and neutralize invading microbes, against two pathogens with a global importance for infectious diseases, namely Salmonella typhimurium and Leishmania infantum, the causative agents of Salmonellosis/typhoid fever and Leishmaniosis/Kala azar. We focused on two metabolic enzymes which use the amino acid arginine as substrate. Inducible nitric oxide synthase (iNOS) is responsible for high output formation of nitric oxide (NO) which exerts anti-microbial activity. Arginase 1 degrades arginine thereby limiting arginine availability for iNOS and protecting microbes from NO mediated anti-microbial immune effector mechanisms. Both enzymes are induced upon infection of macrophages with either microbe, and are similarly regulated by different cytokines. Interestingly, the effects of those enzymes are different on infection control. While genetic or pharmacological blockade of Arginase1 improves the control of infection against Leishmania, this is not the case for Salmonella infection. This is due to divergent dependence on metabolites between the two pathogens, Analysis of Salmonella infected macrophages also suggest that there are pathogen specific effects on host metabolic traits. In addition, pro- and anti-inflammatory cytokines appear to have different effects on metabolic programming of macrophages depending on the nature of the pathogen thereby impacting on immune control of infection. Our data provided novel insights into the complex interrelationship between immune response, metabolic circuits, their epigenetic control and infection control point to the potential of targeted metabolic intervention to ameliorate the course of infections.