The immune system is composed of both innate and adaptive components. The latter, involving antibodies and T cell-mediated cellular immunity is exquisitely sensitive, but requires time for expansion of the rare antigen-specific cells upon initial infection. In contrast, the innate immune system utilizes evolved recognition mechanisms and provides a first line of defense until the adaptive response can reach a significant magnitude. Among the innate immune responses are the responses of macrophages to bacterial cell wall components. The best studied of these is bacterial lipopolysaccharide (LPS), which is the major component of the outer leaflet of the outer membrane of gram-negative bacteria. Recent progress has identified CD14 as an important recognition component for responses to LPS and other bacterial cell wall components. While the responses of macrophages to LPS are ordinarily beneficial, in that they are pro-inflammatory and attract immune effector cells to the site of infection, when activated systemically, as in cases of bacterial sepsis, the result is life-threatening septic shock. This proposal is concerned with understanding the intracellular events that occur in macrophages upon contact with physiologically relevant concentration of LPS, with the view that such understanding may lead to improved methods of treating septic shock. Experiments of AIM 1 will determine the roles of the three MAP kinase-like pathways (classical MAP kinase, p38/HOG-1, and JNK) in mediating the responses to LPS. Experiments of AIM 2 will study the roles of the intracellular tyrosine kinases Syk, Hck, Lyn, and Fgr in initiating LPS signaling events by examining macrophages from mice made genetically defective in one or more of these tyrosine kinases.
In Aim 3, the mechanism by which the signaling adapter protein Shc becomes tyrosine phosphorylated in response to LPS will be determined, as this may allow us to identify the putative signaling chain of the LPS receptor complex. Moreover, these experiments will assess the role of Shc in activation of Ras in response to LPS and in mediating other signaling events. Finally, in this aim, the role of the signaling component Vav in LPS responses will be assessed by studying LPS responses in macrophages from Vav-deficient mice.
