A Distant 10-bp Sequence Specifies the Boundaries of a Programmed DNA Deletion in Tetrahymena

Programmed DNA deletion occurs at thousands of specific sites in most ciliates studied. To understand the mechanism of this prominent DNA rearranging process, we analyzed one of the deletion elements (the M-element) in Tetrahymena by making specific mutations in cloned DNAs and testing their effects on rearrangement in vivo. We found that a 10-bp polypurine sequence (5'-AAAAAGGGGG) plays a crucial role. This sequence is located at a short distance (approximately 45 bp) outside of the element on either side. Removal of it abolishes the deletion process. Moving it short distances away causes the deletion boundary to move with it. Insertion of this sequence into a site within the element induces new boundaries to form near the insertion site. Sequence analysis reveals that all new boundaries created are 41-54 bp away from the sequence. Thus, this sequence is necessary and sufficient to determine the boundaries of DNA deletion, and it does so from a short distance outside of the element. These results offer a possible explanation for the control of DNA deletion in ciliates and suggest that a new type of mechanism for site-specific DNA rearrangements is involved.