Structure and regulation of the cytoplasmic membrane complexes formed during signal transduction by the Toll-like receptors

Humans have evolved complex and effective ways of fighting infections caused by microbes such as bacteria and viruses, the immune system. Sometimes the immune system goes wrong and this can cause serious diseases such as rheumatoid arthritis and diabetes. Our research aims to understand at a molecular level how the cells of the immune system are able to recognise microbes and the ways in which these cells respond to cause the familiar symptoms of an infection such as fever and tiredness, and to generate specific antibodies that fight the invading microbes. In this project we will study the way in which immune system cells respond to a powerful toxins, such as endotoxin or LPS, released by certain types of bacteria during an infection. Although normal responses to endotoxin are an important part of the fight against infection, it can cause a highly dangerous condition called endotoxic shock in patients with severe septicaemia and this pathological response frequently causes organ failure and death. In fact endotoxin shock is responsible for about 135000 deaths every year in Europe alone. The proposed studies should shed light on why endotoxin can produce a beneficial reaction in some conditions and a damaging response in others.

The Toll-like receptors respond to microbial products such as lipopolysaccharide or endotoxin from Gram negative bacteria and activate a downstream signalling pathway that leads to the production cytokines and co-stimulatory molecules. Our previous work using bioinformatic approaches provides strong evidence that stimulus induced dimerization of the TLR cytoplasmic domains causes the formation of new molecular scaffold for the recruitment of signalling adaptor molecules. We plan to study TLR mediated signalling events by investigating how the adaptor molecules engage specifically with activated receptor TIR dimers at the molecular level. We will utilise NMR spectroscopy and X-ray crystallography to determine the molecular structure of the receptor and adaptor TIR domains, independently and in complex. This will provide detailed information critical to understanding the precise interactions involved in receptor-adaptor binding. In the second part of the project we will study the next step in the signalling pathway. After binding of adaptor proteins into an active receptor complex, the non-receptor protein kinase IRAK is recruited into the signalling complex. This involves interactions between the death domains of the adaptor protein MyD88 and the IRAK family protein kinases. These kinases undergo auto- and cross-phosphorylation and we propose to study the role of these modifications in the assembly and dissociation of the death domain complexes formed during the signalling process. In a related project we will determine the structure of trimeric complexes of the adaptors tube and dMyD88 with the kinase pelle in the Drosophila innate immunity and dorsoventral patterning pathways.