REITERATIVE DG ADDITION BY EUPLOTES-CRASSUS TELOMERASE DURING EXTENSION OF NONTELOMERIC DNA

Telomerase from the ciliate Euplotes crassus incorporates G(4)T(4) tel omeric repeats onto both telomeric and non-telomeric single-stranded D NA 3'-ends via reverse transcription of a templating domain in its RNA subunit. Here we describe an unusual mode of template copying that is characteristic of DNA synthesis onto nontelomeric 3'-ends in vitro. W hen dTTP was eliminated from telomerase reactions, telomeric primers o r DNA products generated from the telomerase endonuclease were extende d by precise copying of the RNA template. In contrast, telomerase cata lyzed the addition of up to 13 dG residues onto primers with non-telom eric 3'-ends under the same reaction conditions. Introducing mismatche s in the 3'-terminus of telomeric primers that reduced primer compleme ntarity to the RNA template induced reiterative dG incorporation, indi cating that the reaction is influenced by Watson-Crick base pair forma tion between the primer and the RNA template. Unexpectedly, the reiter ative dG addition mode was confined to telomerase derived from develop ing cells that undergo new telomere formation. This reaction was not o bserved in vegetatively growing cells. We postulate that indiscriminat e dG addition by telomerase occurs by reiterative copying of C residue s in the telomerase RNA templating domain and reflects lateral instabi lity of the primer-template interaction during de novo telomere format ion.