Session Title: Cytokines, Mediators, and Gene Regulation I
Session Type: Abstract Submissions (ACR)
Systemic lupus erythematosus (SLE) is a complex autoimmune disease which afflicts mainly women in the reproductive years, and causes painful arthritis, skin disease and complications in the kidneys and brain. Abnormal T lymphocytes in SLE not only regulate autoantibody-producing B cells, but are also responsible for target organ infiltration. Currently, no specific therapies target T cell defects. Moreover, there are no known molecular markers to predict disease activity. Aberrant molecular mechanisms responsible for SLE T cell defects are thus promising targets for therapy, as well as potential biomarkers for disease. A key defect of T cells from lupus patients is that they produce insufficient amounts of the vital cytokine interleukin (IL)-2. Reduced IL-2 production is linked to reduced cytotoxicity, defective regulatory T cell function, and impaired activation-induced cell death leading to persistence of autoreactive T cells. We previously showed that T cells from SLE patients express decreased levels of the T cell receptor (TCR) – associated CD3 zeta (ζ) chain, a feature directly linked to their poor IL-2 production. We recently showed that the serine arginine (SR) protein splicing factor 2/ alternative splicing factor (SF2/ASF) enhances the expression of CD3ζ chain by limiting the production of an unstable mRNA splice variant. In this study we asked whether the expression of SF2/ASF is aberrant in T cells from SLE patients and if SF2/ASF regulates T cell function, specifically IL-2 production.
T cells were isolated by negative selection from peripheral blood of SLE patients and healthy controls. SF2/ASF mRNA and protein expression was assessed using real time pcr and immunoblotting respectively. T cells were transiently transfected with siRNA or plasmids using electroporation. T cells were activated with anti-CD3 and anti-CD28 antibodies. Transcriptional activity was studied using the dual luciferase reporter assay system. IL-2 production was measured in supernatants by enzyme linked immunosorbent assay (ELISA). Chromatin immunoprecipitation assays were used to assess transcription factor binding to the IL-2 promoter.
We show that SF2/ASF expression levels are decreased in T cells from SLE patients, and correlate inversely with patients’ SLE disease activity index (SLEDAI). Importantly, overexpression of SF2/ASF in SLE T cells enhances their IL-2 production. In parallel, silencing SF2/ASF expression in normal T cells reduces their IL-2 secretion. Using reporter assays, we show that SF2/ASF increases transcriptional activity of the IL-2 promoter. Further, SF2/ASF induces the expression of the nuclear factor of activated T cells (NFAT) and c-fos transcription factors, and induces increased binding of NFAT and c-fos to the IL-2 promoter. Finally, we show recruitment of SF2/ASF to the IL2 promoter indicating its direct role in IL-2 transcription.
Conclusion: Our results identify SF2/ASF as a novel regulator of IL-2 expression in human T cells and a potential molecular mechanism underlying the altered T cell defect in SLE.
V. R. Moulton,
A. P. Grammatikos,
G. C. Tsokos,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-serine-arginine-protein-sf2asf-is-a-novel-regulator-of-il-2-transcription-and-restores-il-2-production-in-t-lymphocytes-from-sle-patients/