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"PROFILING OF INTERFERON REGULATORY FACTOR 5 EXPRESSION AND ACTIVATION IN BLOOD CELLS OF PATIENTS WITH SYSTEMIC LUPUS ERYTHEMATOSUS"

by
Rivka C. Stone
Department of Biochemistry and Molecular Biology
B.S. 2005, New Jersey Institute of Technology



Thesis Advisor: Betsy J. Barnes, Ph.D.
Assistant Professor
Department of Biochemistry & Molecular Biology

Thursday, April 14, 2011
Cancer Center, G Level Seminar Room, 10:00 A.M.


Abstract

Systemic lupus erythematosus (SLE) is a chronic autoimmune disorder affecting multiple organs and is characterized by a type I interferon (IFN) gene signature as well as the production of autoantibodies. Although the underlying etiology of SLE remains obscure, several lines of evidence document a complex interaction between environmental and genetic factors. The transcription factor interferon regulatory factor 5 (IRF5) is one SLE susceptibility gene recently identified. Four IRF5 risk variants, consisting of a CGGGG insertion/deletion as well as SNPs rs2004640, rs10954213 and rs10488631, define distinct haplotypes H1-H9; the H2 risk haplotype shows the highest probability of causality in SLE. IRF5 regulates the expression of proinflammatory cytokines and type I IFNs involved in SLE pathogenesis. In this study, we observed a significant upregulation of IRF5 expression in SLE patients versus healthy donors. Using genotyped patient samples, we found that upregulation of IRF5 transcript and protein expression was associated with the H2 risk haplotype. Profiling of IRF5 transcripts by molecular cloning and next-generation sequencing revealed distinct differential expression of IRF5 in healthy donors and SLE patients; fourteen new full-length IRF5 transcripts and many novel splice junctions were identified. For further insight into the SLE-relevant activities of various IRF5 protein isoforms, we observed differential regulation of proinflammatory IRF5 targets IL12, TNFA, IFNA, and IFNB. To examine IRF5 activation in vivo, we studied its nuclear localization in blood cells of SLE patients and found that IRF5 is already activated in monocytes of these individuals. SLE serum was identified as the trigger for IRF5 nuclear accumulation. Neither IFN nor immune complexes in SLE serum could induce nuclear localization; instead, autoantigens comprised of apoptotic and necrotic cellular material triggered IRF5 nuclear accumulation. Production of IRF5-regulated cytokines IFN, TNF and IL6 in monocytes stimulated with SLE serum or autoantigens correlated with the kinetics of IRF5 nuclear localization. This study provides the first formal proof that genetic variants of IRF5 showing association with SLE regulate its splicing and expression profile, while activation and cellular function of IRF5 are dependent on the SLE blood environment. Results point to IRF5 as a potential therapeutic target for intervention in SLE.


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