Endogenous antisense regulates CD39 in Crohn’s disease upon interaction with nucleolin and heterogeneous-nuclear-ribonucleoprotein-A1

Abstract

The ectoenzyme CD39 hydrolyzes ATP to generate immunosuppressive adenosine. We have shown that CD39 levels and activity are reduced in Treg and Th17-cells from Crohn’s patients. This impairment is linked to heightened levels of endogenous antisense (AS) present at the 3′ end of the human CD39 gene and regulating CD39 at mRNA and protein level. Here we note that CD39-specific AS is predominantly located in the nucleus of peripheral blood-derived Treg and Th17-cells of healthy subjects (HS) and Crohn’s disease patients. RNA pulldown assay followed by mass spectrometry shows that AS regulates CD39 upon interaction with nucleolin (NUCL) and heterogenous-nuclear-ribonucleoprotein-A1 (HNRNPA1). Silencing of NUCL and/or HNRNPA1 results in higher CD39 levels in both Treg and Th17-cells. Blockade of antisense in vitro using FANA oligonucleotides decreases AS while boosting CD39 in HS and Crohn’s Treg and Th17-cells. AS silencing in vivo ameliorates experimental colitis induced by trinitrobenzene-sulfonic-acid in NOD/scid/gamma mice reconstituted with antisense+ human CD4-cells and concomitantly administered FANA oligonucleotides. Compared to untreated controls, FANA-treated mice show decreased disease activity index, higher colon length, lower histology score and, phenotypically, increased proportions of circulating CD39+CD4+-cells and decreased frequencies of peripheral blood and splenic CD4+IL-17+-cells. In conclusion, CD39-specific AS dampens CD39 levels in Crohn’s Treg and Th17-cells by interacting with NUCL and HNRNPA1. Inhibition of AS controls disease activity in experimental colitis suggesting that AS blockade might serve as a novel therapeutic tool to restore CD39 and curb inflammation in Crohn’s disease.