The undue susceptibility of human neonates to infections with intracellular pathogene may result in part from an intrinsic and selective immaturity in production of IFN-gamma by neo-natal T cells and in part from the failure of neonatal M0 to respond to IFN-gamma with increased TNF production. This proposal addresses the central hypothesis that priming of T cells, which occurs by previous antigen or mitogen- induced activation, and of M0, which occurs by exposure to IFN-gamma, acts in part by modifying cellular signal transduction mechanisms.
Aim 1 seeks to determine if the basis for diminished IFN-gamma production, in spite of normal interleukin 2 (IL2) production, by neonatal t cells: 1. reflects in large part the absence of previously primed T cells with """"""""memory"""""""" phenotype; 2. greater production of IFN-gamma by """"""""memory"""""""" T cells reflects more efficient mechanisms transducing the signal to transcribe IFN-gamma mRNA; or 3. alternatively reflects differences in nucleoprotein/chromatin configuration necessary for IFN-gamma transcription. T cells of """"""""memory"""""""" and """"""""non-memory"""""""" phenotype will be identified and purified using monoclonal antibodies; we will also determine if """"""""memory"""""""" T cells can be derived from """"""""non-memory"""""""" T cells by stimulation and propagation in vitro. Using these cells, T cell activation induced protein phosphorylation will be studied using 2D gels and phosphotyrosine antibodies to determine if phosphorylation occurs on tyrosine or serine/threonine; results will be correlated with IFN gamma production and differences in lymphocyte-specific tyrosine kinase (lck) and protein kinase C (PKC) subtype expression and activity using specific cDNA probes, antibodies and functional assay. Alternatively, if correlative differences in signal mechanisms are not confirmed, IFN-gamma compared to IL-2 gene nucleoprotein/chromatin structure will be assessed by DNase mapping and gel retardation assays.
Aim 2 seeks to explore, in a manner analogous to the T cell studies, the role of enhanced/altered signal transduction by PKC and the myeloid specific tyrosine kinases hck and fgr in IFN-gamma enhancement of M0 TNF production. Activation induced protein phosphorylation and expression and activity of these kinases will be correlated with IFN-gamma enhancement of TNF production. If an association between increased expression/activity of a specific kinase and IFN-gamma enhancement of TNF production is found, we will attempt to mimic these effects by introducing into M0 cell lines expression vectors containing that kinase(s). The proposed studies will dissect the lines expression vectors containing that kinase(s). The proposed studies will dissect the mechanisms by which IFN-gamma production by T cells and TNF production by M0 are regulated and enhance out understanding of the selective immaturity of these neonatal defenses.

Project Start
1990-04-01
Project End
1999-03-31
Budget Start
1995-04-01
Budget End
1996-03-31
Support Year
16
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Washington
Department
Pediatrics
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Schoenborn, Jamie R; Wilson, Christopher B (2007) Regulation of interferon-gamma during innate and adaptive immune responses. Adv Immunol 96:41-101
Bilic, Ivan; Koesters, Christina; Unger, Bernd et al. (2006) Negative regulation of CD8 expression via Cd8 enhancer-mediated recruitment of the zinc finger protein MAZR. Nat Immunol 7:392-400
Cowley, Shaun M; Iritani, Brian M; Mendrysa, Susan M et al. (2005) The mSin3A chromatin-modifying complex is essential for embryogenesis and T-cell development. Mol Cell Biol 25:6990-7004
Way, Sing Sing; Wilson, Christopher B (2005) The Mycobacterium tuberculosis ESAT-6 homologue in Listeria monocytogenes is dispensable for growth in vitro and in vivo. Infect Immun 73:6151-3
Way, Sing Sing; Thompson, Lucas J; Lopes, Jared E et al. (2004) Characterization of flagellin expression and its role in Listeria monocytogenes infection and immunity. Cell Microbiol 6:235-42
Way, Sing Sing; Wilson, Christopher B (2004) Cutting edge: immunity and IFN-gamma production during Listeria monocytogenes infection in the absence of T-bet. J Immunol 173:5918-22
Makar, Karen W; Wilson, Christopher B (2004) DNA methylation is a nonredundant repressor of the Th2 effector program. J Immunol 173:4402-6
Kollmann, Tobias R; Way, Sing Sing; Harowicz, Heidi L et al. (2004) Deficient MHC class I cross-presentation of soluble antigen by murine neonatal dendritic cells. Blood 103:4240-2
Shnyreva, Maria; Weaver, William M; Blanchette, Mathieu et al. (2004) Evolutionarily conserved sequence elements that positively regulate IFN-gamma expression in T cells. Proc Natl Acad Sci U S A 101:12622-7
Alaniz, Robert C; Sandall, Sharsti; Thomas, Elaine K et al. (2004) Increased dendritic cell numbers impair protective immunity to intracellular bacteria despite augmenting antigen-specific CD8+ T lymphocyte responses. J Immunol 172:3725-35

Showing the most recent 10 out of 51 publications