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. 2012 Oct 9;109(41):16606-11.
doi: 10.1073/pnas.1210129109. Epub 2012 Sep 26.

cAMP response element modulator α controls IL2 and IL17A expression during CD4 lineage commitment and subset distribution in lupus

Christian M Hedrich  1 , Jose C Crispin, Thomas Rauen, Christina Ioannidis, Sokratis A Apostolidis, Mindy S Lo, Vasileios C Kyttaris, George C Tsokos
Affiliations

cAMP response element modulator α controls IL2 and IL17A expression during CD4 lineage commitment and subset distribution in lupus

Christian M Hedrich et al. Proc Natl Acad Sci U S A. .

Abstract

Appropriate expression of IL-2 plays a central role during the priming and differentiation of T cells. A tight balance between IL-2 and the effector cytokine IL-17A is essential for immune homeostasis. Epigenetic mechanisms have been documented as a key component of cytokine regulation during lineage commitment. The molecular mechanisms that induce chromatin remodeling are less well understood. We investigated epigenetic regulators that mediate the diametric expression of IL-2 and IL-17A in naive, central memory, and effector memory CD4(+) T cells. We demonstrate that cAMP response modulator (CREM)α contributes to epigenetic remodeling of IL2 in effector memory T cells through the recruitment of DNMT3a. CREMα also reduces CpG-DNA methylation of the IL17A promoter. CREMα expression is regulated at the epigenetic level by CpG-DNA methylation, which allows increased CREMα expression in effector memory CD4(+) T cells. T cells from patients with systemic lupus erythematosus (SLE) express increased levels of CREMα and exhibit a phenotype that is similar to effector memory CD4(+) T cells with epigenetically predetermined expression patterns of IL-2 and IL-17A. We conclude that CREMα mediates epigenetic remodeling of the IL2 and IL17A gene during T-cell differentiation in favor of effector memory T cells in health and disease.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
IL-2 and IL-17A mRNA expression and promoter methylation in CD4+ T cells. (A) IL-2 mRNA expression of naive, central memory (CM) and effector memory (EM) CD4+ T cells in response to stimulation with anti-CD3 and anti-CD28 antibodies (mean ± SD). (B) CpG-DNA methylation of the IL2 promoter in CD4+ T cells. Methylation index (MI) as assessed relative to methylated (100%) and unmethylated (0%) control DNA are shown (mean ± SD). (C) IL-17A mRNA expression of naive, CM and EM CD4+ T cells in response to stimulation with anti-CD3 and anti-CD28. (D) CpG-DNA methylation of the IL17A promoter in CD4+ T cells (mean ± SD).
Fig. 2.
Fig. 2.
CREMα controls IL-2 and IL-17A expression on the epigenetic level. (A) IL-2 mRNA expression in primary human T cells 24 h after transfection with empty pcDNA3.1, CREMα, or DNMT3a expression plasmids. CREMα or DNMT3a mediate reduced IL-2 expression (mean ± SD). (B) CpG-DNA methylation (MI) of the IL2 promoter of primary human T cells 24 h after transfection. T cells that were transfected with pcDNA3.1 empty vector exhibited lower IL2 promoter methylation (MI: 5.8) compared with CREMα (MI: 30.1; P = 0.001) or DNMT3a (MI: 27.1; P < 0.001). (C) Jurkat T cells were cotransfected with pcDNA3.1 or CREMα and control siRNA or DNMT3a siRNA. Twenty-four hours after transfection, CREMα overexpression resulted in a significant reduction of IL-2 mRNA expression (lane 4; P = 0.001). Knock-down of DNMT3a reversed these effects (lane 6; P = 0.003). (D) Jurkat T cells were cotransfected with pcDNA3.1 or CREMα and control siRNA or DNMT3a siRNA and harvested after 24 h. CREMα resulted in increased IL2 promoter methylation (MI: 16.4 vs. 78.3; P = 0.007). DNMT3a knock-down reversed these effects (MI: 33.8%). (E) IL-17A expression in primary human T cells 24 h after transfection with pcDNA3.1, CREMα, or DNMT3a. CREMα resulted in increased IL-17A mRNA expression (P = 0.002), whereas DNMT3a did not show an effect. (F) CpG-DNA MI of the IL17A promoter was assessed 24 h after transfection with pcDNA3.1, CREMα, or DNMT3a. T cells that were transfected with pcDNA3.1 exhibited higher MIs (MI: 79.4) compared with cells transfected with CREMα (MI: 51.7; P = 0.03). DNMT3a resulted in complete methylation of the IL17A promoter (MI: 95.4; P = 0.07). (G) Jurkat T cells were cotransfected with pcDNA3.1 or CREMα and control siRNA or DNMT3a siRNA and harvested after 24 h. CREMα resulted in an increase of IL-17A mRNA expression (lane 4; P = 0.004). Knock-down of DNMT3a in combination with CREMα overexpression did not bring on additional effects (lane 6). (H) Jurkat T cells were cotransfected with pcDNA3.1 or CREMα and control siRNA or DNMT3a siRNA and harvested after 24 h. CREMα resulted in reduced IL17A promoter methylation (MI: 59.8 vs. 38.2; P = 0.006). DNMT3a knock-down did not have additional effects.
Fig. 3.
Fig. 3.
CREMα mRNA expression and promoter methylation in CD4+ T cells. (A) pGL3-basic and CREM P1 reporter plasmids were methylated. Promoter activities of the unmethylated and the methylated reporters were assessed in primary human T cells (mean ± SD). (B) T cells were transfected with pcDNA3.1 or DNMT3a. DNMT3a resulted in reduced CREMα mRNA expression (P < 0.001). (C) CREMα mRNA expression in naive, central memory (CM) and effector memory (EM) CD4+ T cells in response to stimulation with anti-CD3 and anti-CD28 antibodies (mean ± SD). (D) CpG-DNA methylation (MI) of the CREM promoter P1 in CD4+ T cells (mean ± SD).
Fig. 4.
Fig. 4.
CREM, IL2, and IL17A mRNA expression and promoter methylation in T cells from SLE patients. T cells from SLE patients and healthy controls were assessed for the expression of (A) CREMα, (C) IL-2, and (E) IL-17A mRNA. Controls were compared with SLE patients with low disease activity (SLEDAI 0–4) and active disease (SLEDAI > 4). T cells from SLE patients and controls were screened for CpG-DNA methylation of the proximal promoter of (B) CREM, (D) IL2, and (F) IL17A.
Fig. 5.
Fig. 5.
SLE patients exhibit a higher percentage of effector CD4+ T cells. (A) A representative pair of age, sex, and ethnicity matched SLE patient and the corresponding healthy control. T cells were stained for CD3, CD4, CD45RA, and CCR7 to define naive, central memory (CM), and effector memory (EM) CD4+ T cells. (B) Frequency of EM CD4+ T cells in total CD4+ T cells from SLE patients (SLEDAI: 4–8) and controls.
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