Systemic lupus erythematosus (SLE) is characterized by disordered T lymphocyte signal transduction. T cells exhibit impaired protein kinase A (PKA)-catalyzed protein phosphorylation due to a profound deficiency of type I PKA (PKA-I) isozyme phosphotransferase activity. Recently, we identified a concomitant deficiency of PKA-II isozyme activity in SLE T cells. Deficient PKA-II activity is associated with (a) autophosphorylation and aberrant translocation of the beta isoform of the type II regulatory subunit (RIIbeta) from the cytosol to the nucleus; (b) accumulation and retention of nuclear RIIbeta; and, (c) reduced or undetectable c-Fos cytosolic protein. Therefore, we hypothesize that aberrant nuclear translocation of autophosphorylated RIIbeta results in (a) deficient PKA-II activity, (b) under-phosphorylation of nuclear cAMP response element binding protein (CREB) transcription factor, (c) impaired c-fos transcriptional activation, (d) reduced levels of c-fos transcript and c-Fos protein, (e) decreased formation of AP-1 transcription factor, and (f) impaired IL-2 transcriptional activation.
The specific aims of this proposal are: (l) To demonstrate that autophosphorylated RIIbeta-subunit is a transcription factor that forms a RIIb-CREB heteromeric complex and acts as a transcriptional repressor of CREB-mediated c-fos transcription in normal primary T cells; (2) To identify the mechanism(s) leading to aberrant translocation of the RIIbeta-subunit from the cytosol to the nucleus in SLE T cells; (3) To determine if PKA-II-catalyzed phosphorylation of CREB is impaired and hinders its binding to CREB binding protein (CBP) and transcriptional activation of the c-Fos promoter in SLE T cells; and, (4) To determine if there is diminished AP-1 binding to consensus AP-1 sites of the IL-2 promoter/enhancer that results in reduced IL-2 production in SLE T cells. The principal goal of the proposed experiments is to establish the mechanism(s) by which deficient PKA-II isozyme activity contributes to altered c-Fos and IL-2 transcriptional activation, loss of AP-1, and diminished IL-2 production by SLE T cells. Demonstrating a connection between reduced T cell IL-2 production and deficient PKA-II isozyme activity in SLE T cells will address a principal gap in our understanding of the molecular and cellular pathophysiology of T cell immunodysfunctions in SLE.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI046526-04
Application #
6706914
Study Section
General Medicine A Subcommittee 2 (GMA)
Program Officer
Johnson, David R
Project Start
2001-03-01
Project End
2006-02-28
Budget Start
2004-03-01
Budget End
2005-02-28
Support Year
4
Fiscal Year
2004
Total Cost
$252,000
Indirect Cost
Name
Wake Forest University Health Sciences
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
937727907
City
Winston-Salem
State
NC
Country
United States
Zip Code
27157
Khan, Islam U; Kammer, Gary M (2004) Protein kinase A and signal transduction in T lymphocytes: biochemical and molecular methods. Methods Mol Med 102:73-85
Elliott, Michael R; Shanks, Ryan A; Khan, Islam U et al. (2004) Down-regulation of IL-2 production in T lymphocytes by phosphorylated protein kinase A-RIIbeta. J Immunol 172:7804-12
Khan, Islam U; Tsokos, George C; Kammer, Gary M (2003) Abnormal B cell signal transduction in systemic lupus erythematosus. Curr Dir Autoimmun 6:89-104
Elliott, Michael R; Tolnay, Mate; Tsokos, George C et al. (2003) Protein kinase A regulatory subunit type II beta directly interacts with and suppresses CREB transcriptional activity in activated T cells. J Immunol 171:3636-44
Kammer, Gary M; Perl, Andras; Richardson, Bruce C et al. (2002) Abnormal T cell signal transduction in systemic lupus erythematosus. Arthritis Rheum 46:1139-54
Kammer, Gary M (2002) Deficient protein kinase a in systemic lupus erythematosus: a disorder of T lymphocyte signal transduction. Ann N Y Acad Sci 968:96-105
Laxminarayana, Dama; Khan, Islam U; Kammer, Gary (2002) Transcript mutations of the alpha regulatory subunit of protein kinase A and up-regulation of the RNA-editing gene transcript in lupus T lymphocytes. Lancet 360:842-9