Short inverted repeats at a free end signal large palindromic DNA formation in Tetrahymena

Large palindromic DNAs are formed in many cell types, but their molecular mechanism is unknown. During nuclear differentiation in Tetrahymena, the ribosomal RNA genes (rDNA) are converted from a single integrated copy to an extrachromosomal head-to-head palindrome. Using in vitro mutagenesis and Tetrahymena transformation, we show that two properties of the rDNA are necessary and sufficient for palindrome formation. The first is a pair of 42 bp inverted repeats found at the rDNA's 5' end. Its inverted symmetry, but not specific sequence, is important. The second is a free end next to the repeats. It is normally created by chromosome breakage in vivo, but can also be provided by restriction endonuclease cutting before transformation. We also demonstrate that the ability to form palindromes is not restricted to developing nuclei, but is present in vegetative cells as well. This process may represent a general mechanism for palindrome formation in eukaryotes.