Functional interaction between telomere protein TPP1 and telomerase.
Human chromosome end-capping and telomerase regulation require POT1 (Protection of Telomeres 1) and TPP1 proteins, which bind to the 3' ssDNA extension of human telomeres. POT1-TPP1 binding to telomeric DNA activates telomerase repeat addition processivity. We now provide evidence that this POT1-TPP1 activation requires specific interactions with telomerase, rather than it being a DNA substrate-specific effect. First, telomerase from the fish medaka, which extends the same telomeric DNA primer as human telomerase, was not activated by human POT1-TPP1. Second, mutation of a conserved glycine, Gly100 in the TEN (telomerase essential N-terminal) domain of TERT, abolished the enhancement of telomerase processivity by POT1-TPP1, in contrast to other single amino acid mutations. Chimeric human-fish telomerases that contained the human TEN domain were active but not stimulated by POT1-TPP1, showing that additional determinants of processivity lie outside the TEN domain. Finally, primers bound to mouse POT1A and human TPP1 were activated for extension by human telomerase, whereas mPOT1A-mTPP1 was most active with mouse telomerase, indicating that these mammalian telomerases have specificity for their respective TPP1 proteins. We suggest that a sequence-specific interaction between TPP1 in the TPP1-POT1-telomeric DNA complex and the G100 region of the TEN domain of TERT is necessary for high-processivity telomerase action.
Department of Chemistry and Biochemistry, Howard Hughes Medical Institute, University of Colorado, Boulder, 80309, USA.
Arthur J
AJ
Zaug
Elaine R
ER
Podell
Jayakrishnan
J
Nandakumar
Thomas R
TR
Cech
D000818:Animals; D002460:Cell Line; D004789:Enzyme Activation / Q000235:genetics; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D009990:Oryzias; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000089804:Shelterin Complex; D019098:Telomerase / Q000378:metabolism*; D034501:Telomere-Binding Proteins / Q000235:genetics / Q000378:metabolism*
C495876:ACD protein, human; C544060:Acd protein, mouse; C440849:POT1 protein, human; D011993:Recombinant Fusion Proteins; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins; C509186:TERT protein, human; D019098:Telomerase; C509187:Tert protein, mouse