Age-associated demethylation of KIR2DL3 core promoter precedes transcription. DNA methylation was quantified by qPCR in CD158b/j⁻ CD4 and CD8 T cells and in CD158b/j⁺ T cells. (A) KIR2DL3 transcripts were low in CD158b/j⁻ (18 young and 14 elderly persons) compared with CD158b/j⁺ T cells (13 young and 12 elderly persons) irrespective of age, although a slight age-associated increase for CD8 T cells was seen. (B) Results for methylation-specific PCR of CpG sites −170/−160, −120/−116, −50, and −27/−23 in CD4 and CD8 T cells negative for CD158b/j cell-surface expression were compared with CD158b/j⁺ T cells; results are shown as box plots of 15 young (□) and 15 elderly (▩) persons. (C) CD8⁺CD45RO⁺CD158b/j⁻ and CD8⁺CCR7⁻CD45RA⁺ from 10 young and 10 elderly persons were compared for demethylation of the KIR2DL3 promoter. (D) The methylation of the KIR2DL3 promoter in CD158b/j⁻ CD8 T cells was examined by bisulfite sequencing. Closed symbols represent methylated, open symbols unmethylated, CpG. A representative example of an elderly and a young person is shown.