Polymorphonuclear cells (PMN) play integral roles in the immune response to invading microorganisms. Activated PMN, however, can damage host tissues due to the production of highly toxic proinflammatory mediators. To learn more about the mechanisms controlling PMN activation, we are studying how adaptor molecules coordinate the signaling pathways initiated by Fc receptors for immunoglobulin (Fc-gammaR) or beta2 integrins, receptors that regulate critical PMN functions. We have new information that the hematopoietic adaptor SLP-76 (SH2 domain-containing Leukocyte-specific Phosphoprotein of 76 kD) is inducibly tyrosine phosphorylated following Fc-gammaR and integrin ligation. Consistent with a possible role for this adaptor in the signaling pathways used by these receptors, SLP-76-/-PMN demonstrate reduced Fc-gammaR and absent integrin-mediated activation in vitro, including defective production of reactive oxygen species (ROS). From these and related observations, we hypothesize that SLP-76, and the SLP-76-associated adaptors PRAM-1 (PML-RARalpha target gene encoding an Adaptor Molecule-1) and ADAP (Adhesion and Degranulation-promoting Adaptor Protein) regulate PMN function by integrating Fc-gammaR and integrin-mediated signals. In this proposal, we will use complementary approaches to investigate this hypothesis and to rigorously dissect the spatial and biochemical requirements for these molecules during PMN activation. Structure-function analyses in Aim 1 will define the domains within SLP-76 that are required to mediate its cellular localization, interaction with associated proteins and PMN-specific functions.
In Aim 2, we will use similar approaches to further investigate the roles of PRAM-1 and ADAP in PMN.
In Aim 3, we will generate and characterize myeloid lineage-restricted gene-targeted mice to determine whether the loss of SLP-76, PRAM-1 and ADAP expression in vivo influences PMN activation in animal models of infection and autoimmunity. These investigations will increase out understanding of PMN function and may provide insights into the development of new treatments for human disorders associated with PMN activation, including immunodeficiency, infection and autoimmunity.
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