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The germline chromosomes in Paramecium and other ciliated protozoa contain regions ()f DNA that are excised and eliminated during the development of a new macronuclear genome. Paramecium tetraurelia internal eliminated sequences (IESs) are invariably flanked by a 5[prime]-TA-3[prime] dinucleotide sequence that is part of a larger 8-bp terminal inverted-repeat consensus sequence. Both features, the absolutely conserved 5[prime]-TA-3[prime] and the remaining 6-bp terminal inverted repeat, are shared with the mariner/Tc1 class of transposons. In this article we describe a mutant cell line (AIM-2) detective in excision of a single IES from the coding region of the A51 surface antigen gene. Excision of the 370-bp IES6649 is prevented by a single A to G transition in the invariably conserved 5[prime]-TA-3[prime] dinucleotide. Failure to excise IES6649 also revealed a 29-bp lES located inside IES6649. Additional experiments with the previously isolated AIM-1 mutant, which also contains an internal lES, shows that alternate excision using the wild-type end of IES2591 with an end from the internal lES is extremely rare or nonexistent. These results indicate that lESs are discrete elements whose excision depends upon nucleotides located within the consensus sequence, but also suggest that additional information is required to match one end of an lES with its excision partner.

The excision of internal eliminated sequences (IESs) from the germline micronuclear DNA occurs during' the differentiation of a new macronuclear genome in ciliated protozoa. In Paramecium, lESs are generally short (28-882 bp), AT rich DNA elements that show few conserved sequence features with the exception of an inverted-terminal-repeat consensus sequence that has similarity to the ends of mariner/Tc1 transposons (KLOBUTCHER and HERRICK 1995). We have isolated and analyzed a mutant cell line that cannot excise a 370-bp IESs (IES2591) from the coding region of the 51A variable surface protein gene. A single micronuclear C to T transition within the consensus sequence prevents excision. The inability to excise IES259l has revealed a 28-bp IES inside the larger IES, suggesting that reiterative integration of these elements can occur. Together, the consensus sequence mutation and the evidence for reiterative integration support the theory that Paramecium IESs evolved from transposable elements. Unlike a previously studied Paramecium IES, the presence of this IES in the macronucleus does not completely inhibit excision of its wild-type micronuclear copy through multiple sexual generations.