RESTRICTION LANDMARK GENOMIC SCANNING (RLGS-M)-BASED GENOME-WIDE SCANNING OF MOUSE-LIVER TUMORS FOR ALTERATIONS IN DNA METHYLATION STATUS

Restriction landmark genomic scanning for methylation (RLGS-PVI) was u sed to detect, and subsequently clone, genomic regions with alteration s in DNA methylation associated with tumorigenesis. Use of a methylati on-sensitive enzyme for the landmark cleavage allows analysis of chang es in methylation patterns, In this study, we used RLGS-M to analyze S V4O T antigen-induced mouse liver tumors derived from interspecific F- 1 hybrids between Mus spretus (S) and C57BL/6 (B6), Because 575 S- and B6-specific RLGS loci/spots have been mapped, tumor-related alteratio ns in the RLGS profile could be immediately localized to specific chro mosomal regions, We previously found that the loss of contiguous loci/ spots could be attributed primarily to DNA loss, whereas loss of solit ary loci/spots could be attributed primarily to DNA methylation, In th is study, we examined 30 mouse liver tumor samples for loss of the 507 mapped loci/spots, Fourteen solitary loci/spots found to be absent or reduced in more than 75% of tumor samples were cloned and subjected t o DNA sequence analyses, Two loci were identified as alpha 4 integrin and p16/CDKN2, genes reported to be involved in tumorigenesis. Thus, R LGS-M can detect alterations in the methylation status of known tumor suppressor genes and provide a method for detecting and subsequently c loning novel genomic regions that undergo alterations in methylation d uring tumorigenesis.