Promoter methylation of PIG7/LITAF. (A) Methylation-specific PCR (MSP), quantitative RT-PC, and Western blot analysis of PIG7/LITAF in RL and SCI-1 cell lines before and after treatment with 4 μM 5-Aza-2′-deoxycitine. The treatment with demethylating Aza restored PIG7/LITAF expression at the RNA and protein levels. In the QRT-PCR data analysis, gene-expression levels were normalized with GAPDH as an internal control; bars indicate standard deviations. C+ indicates methylated control. (B) Methylation-specific PCR (MSP) in 4 primary biopsies from patients with Burkitt lymphoma. M indicates methylated; U, unmethylated; C+, methylated control. (C) Sequence analysis of the PIG7/LITAF promoter region after bisulfite modification. The black box indicates the methylated cytosine, and the white box indicates the unmethylated cytosine in the dinucleotide CpG. Samples analyzed were a positive methylated control (C+); cells from a nontumoral tonsil (unmethylated); Namalwa (unmethylated); SCI-1 (unmethylated); and a lymph-node biopsy from a patient with Burkitt lymphoma (P10931) showing methylation. The numbers correspond to (1) genomic sequence of PIG7/LITAF promoter region, (2) methylated sequence of the PIG7/LITAF promoter region after bisulfite modification, and (3) unmethylated sequence of the PIG7/LITAF promoter region after bisulfite modification.