Minocycline Suppresses Activation of Nuclear Factor of Activated T Cells 1 (NFAT1) in Human CD4+ T Cells

David R.M Graham,Janice E Clements,Gregory L Szeto,Joel L Pomerantz

doi:10.1074/jbc.m110.210518

Abstract

Minocycline is a tetracycline family antibiotic that has anti-inflammatory and immunomodulatory properties. These properties have shown promise in the treatment of conditions such as rheumatoid arthritis, Huntington disease, and multiple sclerosis. As lymphocyte activation is involved in the pathogenesis of many of these diseases, T cells are postulated to be a primary target in minocycline therapy. Previous studies have demonstrated attenuation of CD4(+) T cell activation by minocycline, but a specific mechanism has not been elucidated. In this study, we investigated the effect of minocycline on the activity of three key transcription factors regulating CD4(+) T cell activation: NF-κB, AP-1 (activator protein 1), and NFAT (nuclear factor of activated T) cells. Our data demonstrate that minocycline selectively impairs NFAT-mediated transcriptional activation, a result of increased phosphorylation and reduced nuclear translocation of the isoform NFAT1. Minocycline increased the activity of the NFAT kinase GSK3 and decreased intracellular Ca(2+) flux, both of which facilitate NFAT1 phosphorylation. These findings provide a novel mechanism for minocycline induced suppression of CD4(+) T cell activation and may better inform the application of minocycline as an immunomodulatory agent.

Highlights

T cell activation contributes to disease pathogenesis by creating an inflammatory environment that leads to cellular damage and death as well as decreased immune function
To determine which pathways contributed to the suppressive effect of minocycline on activation, primary human CD4ϩ T cells from healthy donors were co-transfected with a reporter plasmid expressing firefly luciferase responsive to NF-␬B, NFAT, or AP-1, along with a plasmid expressing Renilla luciferase under herpes simplex virus thymidine kinase promoter to control for transfection efficiency
Minocycline pretreatment did not significantly impact transcriptional activation mediated by NF-␬B or AP-1 when cells were activated by phorbol 12-myristate 13-acetate (PMA) alone or PMA with ionomycin (Fig. 1, A and B)

Summary

Introduction

T cell activation contributes to disease pathogenesis by creating an inflammatory environment that leads to cellular damage and death as well as decreased immune function. To determine which pathways contributed to the suppressive effect of minocycline on activation, primary human CD4ϩ T cells from healthy donors were co-transfected with a reporter plasmid expressing firefly luciferase responsive to NF-␬B, NFAT, or AP-1, along with a plasmid expressing Renilla luciferase under herpes simplex virus thymidine kinase promoter to control for transfection efficiency. Minocycline pretreatment mediated a dose-dependent decrease in nuclear translocation of NFAT1 relative to untreated, activated controls at both 2 and 4 h post-activation (Fig. 2A).

Results

Conclusion