The cyclin D1 promoter region showed differential cytosine methylation in cyclin D1⁺ and cyclin D1^– cell lines. (A) Diagram of the cyclin D1 promoter region analyzed. The bent arrow represents the transcription start site, the vertical lines mark the locations of CpG dinucleotides, and filled rectangles indicate the positions of PCR primers used for sodium bisulfite genomic sequence analysis. Msp indicates the positions of restriction sites. (B). Bisulfite analysis of the cyclin D1 promoter region methylation in cyclin D1-overexpressing cell lines (Granta, NCEB1, and U266), a cyclin D1 low-expressing cell line (K562), and cyclin D1–nonexpressing cell lines (HL60 and Manca). (C) Bisulfite analysis of the upstream region of the cyclin D1 promoter in Granta and HL60 cell lines. CpG sites in the region (–2385 to –845) were not analyzed. Diagrams in A and C (upper panels) show locations of CpG dinucleotides relative to the cyclin D1 gene transcription start.