UTILIZATION OF RIBONUCLEOTIDES AND RNA PRIMERS BY TETRAHYMENA TELOMERASE

Telomerase is a ribonucleoprotein (RNP) DNA polymerase involved in telomere synthesis, A short sequence within the telomerase RNA component provides a template for de novo addition of the G-rich strand of a telomeric simple sequence repeat onto chromosome termini, In vitro, telomerase can elongate single-stranded DNA primers processively: one primer can be extended by multiple rounds of template copying before product dissociation, Telomerase will incorporate dNTPs or ddNTPs and will elongate any G-rich, single-stranded primer DNA, In this report, we show that Tetrahymena telomerase was able to incorporate a ribonucleotide, rGTP, into product polynucleotide, Synthesis of the product [d(TT)r(GGGG)](n) was processive, suggesting that the chimeric product remained associated with the enzyme both at the active site and at a second, previously characterized, template-independent product binding site, As predicted by this finding, RNA-containing oligonucleotides served as primers for elongation, More than 3 nt of RNA at a primer 3' end decreased the quantity of product synthesis but increased the affinity of the primer for telomerase. Thus, RNA-containing primers were effective as competitive inhibitors of DNA primer elongation by telomerase, These results support the possible evolutionary origin of telomerase as an RNA-dependent RNA polymerase.