Methylation of ribosomal RNA genes in the macronucleus ofTetrahymena thermophila

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Abstract
We have investigated the occurrence of methylated adenine residues in the macronuclear ribosomal RNA qenes of Tetrahymena thermophila. It has been shown previously that macronuclear DNA, including the palindromic ribosomal RNA genes (rDNA), of Tetrahymena thermophila contains the modified base N-6-methyladenine, but no 5-methylcytosine. Purified rDNA was digested with restriction enzymes Sau 3AI, Mbol and Dpnl to map the positions and levels of N-6-methyladenine in the seguence 5' GATC 3'. A specific pattern of doubly methylated GATC seguences was found; hemimethylated sites were not detected. The patterns and levels of methylation of these sites did not change significantly in different physiological states. A molecular form of the rDNA found in the newly developing macronucleus and for several generations following the sexual process, conjugation, contained no detectably methylated GATC sites. However, both the bulk macronuclear DNA and palindromic rDNA from the same macronuclei were methylated. Possible roles for N-6-methyladenine in macronuclear DNA are discussed in light of these findings.