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. 2015 Nov;45(11):3200-3.
doi: 10.1002/eji.201545646. Epub 2015 Oct 1.

Epigenetic analysis of regulatory T cells using multiplex bisulfite sequencing

Daniel B Rainbow  1 , Xin Yang  1 , Oliver Burren  1 , Marcin L Pekalski  1 , Deborah J Smyth  1 , Marcus D R Klarqvist  1 , Christopher J Penkett  2 , Kim Brugger  3 , Howard Martin  3 , John A Todd  1 , Chris Wallace  1   4 , Linda S Wicker  1
Affiliations

Epigenetic analysis of regulatory T cells using multiplex bisulfite sequencing

Daniel B Rainbow et al. Eur J Immunol. 2015 Nov.
No abstract available

Keywords: Bisulfite sequencing; FOXP3; Methylation; Next-generation sequencing; Regulatory T cell.

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Figures

Figure 1
Figure 1
FOXP3 TSDR sequencing of purified human CD4+ T‐cell subsets. (A) Naïve (CD4+, CD127+, CD45RA+, CD62L+) and memory (CD4+, CD127+, CD45RA−, CD62L+) effector cells and Tregs (CD4+, CD25+, CD127−) were sorted from a male (B) and a female (C) donor by flow cytometry and analyzed for FOXP3 TSDR methylation (two representative donors out of a total of four, for more information see Supporting Information Fig. 2 and Supporting Information Table 1). See Supporting Information Fig. 7 for gating strategy defining CD4+ TCRαβ+ T cells. The x axis shows the nine methylation sites analyzed and each row indicates the methylation status of one copy of the sequenced TSDR. Light green represents a C (methylated) and dark green represents a T (demethylated). The y axis is the percentage of sequencing reads for each methylation/demethylation pattern. Six replicates of 3000 cells per replicate (5 ng DNA) were analyzed from a single donor and the median (with range) reads with eight or nine sites demethylated at FOXP3 for the replicates is shown below each graph. (D) A schematic of the FOXP3 gene and the TSDR sequence are shown.
Figure 2
Figure 2
CTLA4 demethylation in human FACS‐purified cells. Cells were purified and analyzed as detailed in Fig. 1 for CTLA4 methylation status. The median (with range) reads with all seven sites demethylated at CTLA4 for the replicates is shown below each graph. Data are representative of four donors (for more information see, Supporting Information Fig. 2 and Supporting Information Table 2).
See this image and copyright information in PMC

References

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