Bacterial infections are still a major cause of morbidity and mortality, especially in immunocompromised individuals. Peptidoglycan and endotoxin can reproduce most of the major signs and symptoms of infections, including production of cytokines and induction of septic shock. The prerequisite for release of cytokines is interaction of peptidoglycan and endotoxin with their specific receptors on host cells, which results in triggering of signal transduction pathways that culminate in induction of transcription of cytokine genes and secretion of cytokines. The overall goal of this project is to identify peptidoglycan receptor and peptidoglycan-activated signal transduction pathways. This will enable rational development of new therapeutic agents that can block the consequences of peptidoglycan-induced cell activation (sepsis syndrome, arthritis, etc.). The applicant proposes to test three hypotheses to determine if CD14, a macrophage endotoxin receptor, also serves as a receptor for peptidoglycan. Hypothesis I: Peptidoglycan binds to membrane CD14 and soluble CD14, which function as cell-activating receptors for PGN. The applicant proposes to test this hypothesis by i) demonstrating binding of peptidoglycan to membrane CD14 and soluble CD14; ii) determining if CD14-negative cells are activated by soluble CD14:peptidoglycan complexes; and iii) defining structural requirements of membrane and soluble CD14 for peptidoglycan binding and cell activation. Hypothesis II: Interaction of peptidoglycan with CD14 induces a cascade of phosphorylation events that starts with activation of Lyn and results in activation of MAP kinases. The applicant proposes to test this hypothesis by: i) defining the pathway through which Lyn transmits the activation signal into the cell; ii) identifying the pathways of activation of MAP kinases by peptidoglycan; and iii) determining if peptidoglycan-induced activation of MAP kinases is Lyn-dependent. Hypothesis III: Interaction of peptidoglycan with CD14 induces signal transduction pathways that culminate in activation of transcription factors that induce transcription of cytokine genes and secretion of cytokines. The applicant will test this hypothesis by: i) determining if peptidoglycan induces transcription of transcription factor and cytokine genes; ii) identifying transcription factors activated by peptidoglycan; iii) determining if peptidoglycan-induced transcription factors are required and sufficient for activation of cytokine genes; and iv) determining if Lyn and MAP kinases induce activation of transcription factors and cytokine genes.
Showing the most recent 10 out of 29 publications
