The effects of nucleoside analogs on telomerase and telomeres inTetrahymena

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Abstract
The ribonucleoproteln enzyme telomerase is a specialized type of cellular reverse transcrlptase which synthesizes one strand of telomerlc DNA, using as the template a sequence in the RNA moiety of telomerase. We analyzed the effects of various nucleoside analogs, known to be chain-terminating inhibitors of retroviral reverse transcriptases, on Tetrahymena thermophila telomerase activity in vitro.Yle also analyzed the effects of such analogs on telomere length and maintenance in vivo, and on vegetative growth and mating of Tetrahymena cells. Arabinofuranyl-guanosine triphos-phate (Ara-GTP) and ddGTP both efficiently inhibited telomerase activity in vitro, while azidothymldine triphosphate (AZT-TP), dldeoxylnosine triphosphate (ddlTP) or ddTTP were less efficient inhibitors. All of these nucleoside triphosphate analogs, however, produced analog-specific alterations of the normal banding patterns seen upon gel electrophoresis of the synthesis products of telomerase, suggesting that their chain terminating and/or competitive actions differ at different positions along the RNA template. The analogs AZT, 3′-deoxy-2′,3′-dldehydrothymidine (d4T) and Ara-G in nucleoside form caused consistent and rapid telomere shortening In vegetatively growing Tetrahymena. In contrast, ddG or ddl had no effect on telomere length or cell growth rates. AZT caused growth rates and viability to decrease in a fraction of cells, while Ara-G had no such effects even after several weeks in culture. Neither AZT, Ara-G, acyclc-guanosine (Acyclo-G), ddG nor ddl had any detectable effect on cell mating, as assayed by quantitation of the efficiency of formation of progeny from mated cells. However, AZT decreased the efficiency of programmed de novo telomere addition during macronuclear development In mating cells.