Ec. Greene et al., FLEXIBLE POSITIONING OF THE TELOMERASE-ASSOCIATED NUCLEASE LEADS TO PREFERENTIAL ELIMINATION OF NONTELOMERIC DNA, Molecular and cellular biology, 18(3), 1998, pp. 1544-1552
In addition to a reverse transcriptase activity, telomerase is associa
ted with a DNA endonuclease that removes nucleotides from a primer 3'
terminus prior to telomere repeat addition, Here we examine the DNA sp
ecificity of the primer cleavage-elongation reaction carried out by th
e Euplotes crassus telomerase. We show that the primer cleavage activi
ty copurified with the E. crassus telomerase polymerase, indicating th
at it either is an intrinsic property of telomerase or is catalyzed by
a tightly associated factor, Using chimeric primers containing stretc
hes of telomeric DNA that could be precisely positioned an the RNA tem
plate, we found that the cleavage site is more flexible than originall
y proposed, Primers harboring mismatches in dT tracts that aligned opp
osite nucleotides 37 to 40 in the RNA template were cleaved to elimina
te the mismatched residues along with the adjacent 3' sequence, The cl
eaved product was then elongated to generate perfect telomeric repeats
, Mismatches in dG tracts were not removed, implying that the nuclease
does not track coordinately with the polymerase active site. Our data
indicate that the telomerase-associated nuclease could provide a rudi
mentary proofreading function in telomere synthesis by eliminating mis
matches between the DNA primer and the 5' region of the telomerase RNA
template.