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Methods for analyzing the role of DNA methylation and chromatin structure in regulating T lymphocyte gene expression

  • Qianjin Lu
  • Bruce Richardson
Open Access
Article

Abstract

Chromatin structure, determined in part by DNA methylation, is established during differentiation and prevents expression of genes unnecessary for the function of a given cell type. We reported that DNA methylation and chromatin structure contributes to lymphoidspecific ITGAL (CD11a) and PRF1 (perforin) expression. We used bisulfite sequencing to compare methylation patterns in the ITGAL promoter and 5′ flanking region of T cells and fibroblasts, and in the PRF1 promoter and upstream enhancer of CD4+ and CD8+ T cells with fibroblasts. The effects of methylation on promoter function were tested using regional methylation of reporter constructs, and confirmed by DNA methyltransferase inhibition. The relationship between DNA methylation and chromatin structure was analyzed by DNaseI hypersensitivity. Herein we described the methods and results in greater detail.

Indexing terms

DNA methylation T-lymphocytes Gene expression regulation 

Abbreviations

PHA

phytohemagglutinin

5-azaC

5-azacytidine

dC

deoxycytosine

SAM

Sadenosylmethionine

MeCP2

methyl-CpG-binding protein 2

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Copyright information

© Springer 2004

Authors and Affiliations

  • Qianjin Lu
    • 1
    • 2
  • Bruce Richardson
    • 1
    • 2
  1. 1.Department of MedicineUniversity of Michigan, the Ann Arbor Veterans Affairs Medical CenterAnn ArborUSA
  2. 2.the Second Xiangya HospitalCentral South UniversityChangshaChina

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