Background/Purpose:

Systemic lupus erythematosus (SLE) is a chronic debilitating autoimmune disease that primarily afflicts women in the childbearing years. Female hormones especially estrogen are implicated in the pathogenesis of disease, however the precise molecular mechanisms regulated by estrogen in immune cells are not well understood. We previously identified novel roles for the multifunctional protein serine/arginine-rich splicing factor 1 (SRSF1) in T cells. We showed that SRSF1 promotes expression of the CD3zeta signaling gene and activates IL-2 production in normal T cells. T cell specific Srsf1-conditional knockout mice exhibit defects in T cell function and develop autoimmune disease. SRSF1 expression levels are decreased in T cells from SLE patients, and associate with severe disease activity. Importantly, overexpression of SRSF1 into SLE T cells rescues IL-2 production. These results imply that the SRSF1 is an important molecule in T cells and immune-mediated disease. However, not much is known about the regulation of SRSF1, and if hormones may contribute to its expression.

Methods:

Peripheral blood was collected from healthy women in the follicular phase of the menstrual cycle. T cells were isolated by negative selection from peripheral blood. T cells were cultured (in RPMI without phenol red supplemented with charcoal-stripped serum), with increasing concentrations (0, 1nM, 10nM, 100nM) of beta-estradiol for 18 – 24 hours. To assess mRNA stability, transcription was blocked with actinomycin D, cells collected at 0, 0.5, 1, 1.5 and 2 hours, and Srsf1expression assessed by RT-qPCR. To assess posttranslational protein degradation mechanisms, the proteasome inhibitor MG132 or the lysosome inhibitor Bafilomycin A1 were added to cultures followed by western blots for SRSF1. The Srsf1 promoter was cloned into a pGL3-luciferase plasmid. T cells were transfected by electroporation and luciferase activity measured by a luminometer.

Results:

Exposure to estrogen led to a dose dependent increase in Srsf1 mRNA expression, but decreased protein levels in T cells. This discrepancy between mRNA and protein levels suggests that estrogen controls SRSF1 at transcriptional and post-transcriptional levels and may upregulate transcriptional activity of its promoter but downregulate protein levels via mRNA decay, translation or protein degradation.Accordingly, estrogen increased luciferase activity of the Srsf1 promoter-luciferase construct indicating that estrogen activates Srsf1 transcription. Estrogen did not affect mRNA stability, however protein analyses showed that the proteasome is involved in the degradation of SRSF1. In silico analysis revealed microRNA (miR) target sites within the Srsf13`UTR, of which miR-21-5p, miR-27, and miR-200b-3p are known to be regulated by estrogen and to be dysregulated in SLE.

Conclusion:

Our results suggest that estrogen can modulate the expression of SRSF1 via transcriptional and posttranscriptional mechanisms in human T lymphocytes, thus revealing a potential molecular link between hormones, immune cells and autoimmune disease.

« Back to 2018 ACR/ARHP Annual Meeting

ACR Meeting Abstracts - https://acrabstracts.org/abstract/estrogen-controls-the-expression-of-serine-arginine-rich-splicing-factor-1-srsf1-in-human-t-lymphocytes-via-transcriptional-and-post-transcriptional-mechanisms/