Human in vitro induced T regulatory cells and memory T cells share common demethylation of specific FOXP3 promoter region.
BACKGROUND
The FOXP3 gene is the master regulator for T regulatory cells and is under tight DNA methylation control at the Treg specific demethylated region (TSDR) in its first intron. This said, methylation of its promoter region, the significance of which is unknown, has also been associated with various immune-related disease states such as asthma, food allergy, auto-immunity and cancer. Here, we used induced T regulatory cells (iTreg) as a target cell population to identify candidate hypomethylated CpG sites in the FOXP3 gene promoter to design a DNA methylation quantitative assay for this region.


RESULTS
Three CpG sites at the promoter region showed clear demethylation pattern associated with high FOXP3 expression after activation in presence of TGFβ and were selected as primary targets to design methylation-dependent RT-PCR primers and probes. We then examined the methylation of this 'inducible-promoter-demethylated-region' (IPDR) in various FOXP3+ T cell subsets. Both naïve and memory thymic-derived Treg cells were found to be fully demethylated at both the IPDR and TSDR. Interestingly, in addition to iTregs, both CD25- and CD25(lo) conventional memory CD4+CD45RA- T cells displayed a high fraction of IPDR demethylated cells in absence of TSDR demethylation.


CONCLUSIONS
This implies that the fraction of memory T cells should be taken in account when interpreting FOXP3 promoter methylation results from clinical studies. This approach, which is available for testing in clinical samples could have diagnostic and prognostic value in patients with immune or auto-inflammatory diseases.
Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Stanford University, Stanford, CA USA.|Division of Allergy, Department of Pediatrics, CHU Sainte-Justine, Montreal, Canada.
Philippe
P
Bégin
Epiontis, Berlin, Germany.
Janika
J
Schulze
Epiontis, Berlin, Germany.
Udo
U
Baron
Epiontis, Berlin, Germany.
Sven
S
Olek
Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Stanford University, Stanford, CA USA.
Rebecca N
RN
Bauer
San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy.
Laura
L
Passerini
Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Stanford University, Stanford, CA USA.|San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), Division of Regenerative Medicine, Stem Cells and Gene Therapy, San Raffaele Scientific Institute, Milan, Italy.
Rosa
R
Baccheta
Division of Allergy, Immunology and Rheumatology, Department of Pediatrics, Stanford University, Stanford, CA USA.
Kari C
KC
Nadeau
Assay; Epigenetic; FOXP3; IPDR; Induced; Methylation; Promoter; TSDR; Treg