TRANSIENT DNA BREAKS ASSOCIATED WITH PROGRAMMED GENOMIC DELETION EVENTS IN CONJUGATING CELLS OF TETRAHYMENA-THERMOPHILA

Thousands of programmed genomic deletion events occur during macronucl ear development in Tetrahymena thermophila. Two of the deleted segment s, called M and R, have been particularly well characterized. Using li gation-mediated PCR, we have detected DNA strand breaks that correlate temporally and structurally with the deletion events in the M and R r egions. The ends appear at positions that correspond precisely to boun daries of deleted sequences, as defined by observed chromosomal juncti ons found after deletion is complete. They occur exclusively during th e known DNA rearrangement period in macronuclear development. The brea ks are staggered by 4 bp in the complementary strands. Several alterna tive breaks were found at the end of one deleted region, consistent wi th multiple alternative chromosomal junctions detected previously. The free 5' ends generated at the breaks are phosphorylated. A purine res idue always occurs at the free 3' ends, with an adenosine appearing in 11 of 12 cases. Patterns found in the detected break sites suggest ru les that define the ends of the deleted segments within a transposon-l ike deletion mechanism.