POSITION EFFECT TAKES PRECEDENCE OVER TARGET SEQUENCE IN DETERMINATION OF ADENINE METHYLATION PATTERNS IN THE NUCLEAR GENOME OF A EUKARYOTE, TETRAHYMENA-THERMOPHILA

Approximately 0.8% of the adenine residues in the macronuclear DNA of the ciliated protozoan Tetrahymena thermophila are modified to N-6-met hyladenine, DNA methylation is site specific and the pattern of methyl ation is constant between clonal cell lines. In vivo, modification of adenine residues appears to occur exclusively in the sequence 5'-NAT-3 ', but no consensus sequence for modified sites has been found. In thi s study, DNA fragments containing a site that is uniformly methylated on the 50 copies of the macronuclear chromosome were cloned into the e xtrachromosomal rDNA. In the novel location on the rDNA minichromosome , the site was unmethylated. The result was the same whether the seque nces were introduced in a methylated or unmethylated state and regardl ess of the orientation of the sequence with respect to the origin of D NA replication. The data show that sequence is insufficient to account for site-specific methylation in Tetrahymena and argue that other fac tors determine the pattern of DNA methylation.