Signalling and activation of TLR4 by Gram-negative bacteria in epithelial cells

Abstract: Lipopolysaccharide (LPS) is a well-characterized pathogen-associated molecule found primarily as a constituent of the outer membrane of Gram-negative bacteria. LPS is known to act on different cell types in a pleiotropic manner. Central to LPS induced transduction is the activation of a homologue of the Drosophila Toll receptor, termed Toll-like receptor 4 (TLR4). In addition to various signalling transduction proteins, extracellular modulators such as CD14, LBP and MD-2 are also required for LPS signalling. Stimulation of the TLR4 signalling cascade predominantly terminates in the activation of the transcription factor NFkappaB. One key function of the mucosal epithelial lining is to detect and determine the presence of bacteria. In this thesis, the significance of a TLR4 signalling process has been investigated in epithelial cells following exposure to LPS or Gram-negative bacteria. The Grain-negative bacterium Yersinia pseudotuberculosis can circumvent NF-kappaB activation by translocation of the YopJ protein. This finding has now been extended by demonstrating that YopJ is also acting on the MAPK signalling cascade involving the transcription factors CREB and AP-1. The varying magnitude of host responses to LPS in different cell types, intestinal and bladder epithelia, provide evidence of a tissue specific design to handle invasion of bacteria. In the alimentary tract, which is rich in micro-organisms, TLR4 expression is barely detectable in epithelial cells. Hence, the threshold to mount an inflammatory response is elevated. The reverse is true in a germ-free compartment, Re the urine bladder, where higher levels of TLR4 receptors increase the sensing system to swiftly respond upon invasion of bacteria. In the last part of this thesis, additional modes of LPS/TLR4 induced activation of NF-kappaB in epithelial cells are explored. A potentially novel signalling pathway involving the activation of Akt kinase through p38 MAPK, and thus independent of PI3-kinase, is described. In conclusion, the results presented in this thesis underline the important regulatory role of the sophisticated epithelial lining in mammals. Further understanding of how epithelial cells set, direct and determine host responses and how these are interpreted into a global immune response is highly warranted.