Scientific Report 2005

Immunology

Molecular Mechanisms of Host-Pathogen Interactions

R.J. Ulevitch, V.V. Kravchenko, C. Fearns, T.-H. Chuang, J.C. Mathison, Q. Pan, J. da Silva Correia, K. Iwata, K.D. Janda, G. Kaufmann, M. Meijler

Infection by microbial pathogens often sets in motion chains of events that cause severe injury to the host, and nowhere is this phenomenon illustrated more dramatically than in the response by humans to infection by gram-negative bacteria. In his book Lives of a Cell, Lewis Thomas characterized the host response to the endotoxin, or lipopolysaccharide, of gram-negative bacteria as being “read by our tissues as the very worst of bad news. . . . There is nothing intrinsically poisonous about endotoxin, but it must look awful, or feel awful, when sensed by cells. Cells believe that it signifies the presence of gram-negative bacteria, and they will stop at nothing to avoid this threat.” In other words, the innate immune response to infection has caused a serious disease in humans.

Clearly, much human suffering could be eased if such overzealous host responses could be tempered. However, such responses, when not overzealous, are a normal part of the host’s homeostatic mechanisms, designed to respond to the threat of infection by gram-negative bacteria. Accordingly, we are attempting to (1) define the mechanisms of innate immunity and (2) learn how to control these responses without compromising host defenses against pathogens. Most recently, we contributed to the understanding of innate immunity through our studies of Toll-like receptors (TLRs) and of effector mechanisms that mediate host responses to infection.

It is now well appreciated that the innate immune system is positioned at the intersection of multiple host pathways, including those for microbial and viral recognition, enhancement of adaptive immune responses, and, possibly, cancer immunosurveillance. Each pathway depends on ligand recognition by specific cellular receptors that are either membrane bound (plasma membrane as well as endosomal compartments) or cytosolic. The most important class of membrane-bound receptors are the TLRs. Among cytosolic receptors, an important family known as the Nod/Caterpillar family has been identified. Within this family, 2 proteins, Nod1 and Nod2, are involved in recognition of bacterial ligands distinct from the ligands for TLRs. Activation of TLR and Nod signaling pathways leads to production of multiple cytokines with proinflammatory and anti-inflammatory activities. Such responses are central to host responses to infection. However, when a breakdown occurs in the normal regulatory mechanisms that control these pathways, disease may result.

Perhaps the most well-understood link between innate immunity and human disease is in the host response to infection. When dysregulation of innate immune responses occurs, clinical abnormalities such as septic shock and acute respiratory distress syndrome occur. Dysregulation of innate immune responses may also play a role in human diseases in which chronic inflammation is responsible for disease progression, including autoimmune and autoinflammatory diseases. Genetic studies in humans have revealed strong associations among various members of the Nod family of proteins and human diseases.

During the past year, we made considerable progress in several different areas. First we established mice that lack both copies of the gene for Triad3A, an E3 ubiquitin ligase that controls the expression of some TLRs. We are breeding these mice to begin studies of the cellular phenotype that occurs when the innate immune system is activated. At the same time, we are continuing our studies of the interactions between Triad3A and potential endogenous substrates. Although it is too early to know the exact consequences of the deletion of Triad3A, our initial observations suggest that homozygous Triad 3A^–/– mice have lower body weight and size during the first 8 weeks of life than do their wild-type littermates.We continue to study the biological functions of Nod1 and Nod2. We discovered a signaling pathway in MCF-7 cells in which Nod1 negatively regulates the response of this human breast cancer cell line to estrogen and also influences the level of expression of the estrogen receptor. We found that MCF-7 cells containing Nod1 had greater proliferative responses in tissue culture than did cells lacking Nod1. Most importantly, in a xenograft model of tumor growth in mice with severe combined immunodeficiency, growth of MCF-7 cells containing Nod1 was greater than growth of cells lacking Nod1. We are working out the molecular details of the pathway leading from Nod1 to the estrogen receptor.

We are also continuing our studies on the biological function of Nod2, with a specific emphasis on its role in the assembly of a macromolecular protein complex that induces release of the cytokine IL-1 from cells after activation with the Nod2 ligand. Using cells genetically deficient in specific components associated with IL-1 release, we showed that the Nod2 pathway requires the kinase receptor interacting protein-2 and an adaptor protein known as ASC. Currently, we are using cells deficient in other proteins that may also be implicated in the release of IL-1. Our long-term goal is to identify the nature of this activating complex so that strategies can be devised to block IL-1 release in chronic inflammatory diseases.

Finally, in collaboration with K.D. Janda, Department of Chemistry, we are studying a class of bacterial products known as quorum-sensing factors. These factors play a crucial role in the adaptation of bacteria to the host and serve as a means for bacteria to communicate with one another. Quorum-sensing factors are involved in the induction of virulence factors and the establishment of biofilms. Thus, this class of bacterial molecules clearly plays a role in pathogenesis of infectious processes.

Now we are investigating the role in quorum sensing of members of the homoserine lactone family, which induce inflammation and cell death via their effects on host cells. Inflammation and cell death induced by homoserine lactones may be important processes in diseases such as cystic fibrosis in which the local concentrations of the quorum-sensing factor can be quite high in the lung and in which inflammation and cell remodeling and/or death are unchecked. By understanding how the quorum-sensing molecules activate host cells, we may be able to devise new therapeutic strategies for treatment of diseases in which these molecules are involved.

Publications

Fort, M.M., Mozaffarian, A., Stover, A.G., da Silva Correia, J., Johnson, D.A., Crane, R.T., Ulevitch, R.J., Persing, D.H., Bielefeldt-Ohmann, H., Probst, P., Jeffery, E., Fling, S.P., Hershberg, R.M. A synthetic TLR4 antagonist has anti-inflammatory effects in two murine models of inflammatory bowel disease. J. Immunol. 174:6416, 2005.

Ulevitch, R.J. Therapeutics targeting the innate immune system. Nat. Rev. Immunol. 4:512, 2004.

Ulevitch, R.J., Mathison, J.C., da Silva Correia, J. Innate immune responses during infection. Vaccine 22(Suppl. 1):S25, 2004.