The Macronuclear Ribosomal DNA of Tetrahymena pyriformis is a Palindrome
Format of Original
Journal of Molecular Biology
The macronuclear ribosomal DNA of Tetrahymena pyriformis (strain BVII, syngen 1) is extraohromosomal. The majority of the molecules are linear with measured mean molecular weights of 13.0 × 106 to 13.8 × 106 in various experiments. Several lines of evidence indicated that the double-stranded rDNA molecule is a palindrome in the terminology of Wilson & Thomas (1974). That is, taking into account the polarity of the polynucleotide strands, the sequence is the same read from either end of the molecule. The rDNA molecules had a partial denaturation map that was symmetrical about the center of the molecule. There were two sites susceptible to cleavage by the restriction enzyme EcoRI. Partial denaturation of the fragments indicated that these two sites were equidistant from the center of the rDNA molecule. There were two different sites susceptible to cleavage by the restriction enzyme Bam, which were also equidistant from the center of the rDNA molecule.
The two polynucleotide strands of a palindromic DNA molecule are similar or identical and each is self-complementary. Thus each strand will rapidly renature with first-order kinetics following denaturation. Thermal denaturation of the rDNA of Tetrahymena revealed a complex melting curve with at least six transitions. When the temperature was reduced under conditions such that intermolecular reassociation was not expected to occur, the rDNA abruptly lost 35% of its hyperchromicity. When the products of this snapback renaturation were examined in the electron microscope, many of the molecules observed were double stranded, and had a contour length half the length of the native rDNA. Thermal denaturation of the snapback molecules yielded the same transitions as the original melt, but in different proportions, as predicted from the partial denaturation map. There was little or no mismatching in the self-complementary strands that could be detected by comparison of melting temperature (tm) between the melt and the remelt. Cruciform structures were generated by incubating the rDNA in vitro.
Although the rDNA of the amicronucleate Tetrahymena strain GL is slightly different in size and buoyant density, and in its partial denaturation map from that of strain BVII, it too has a palindromic structure.