Telomere DNA G-quadruplex folding within actively extending human telomerase

Telomerase maintains telomere length by reverse transcribing short G-rich DNA repeat sequences from its internal RNA template. G-rich telomere DNA repeats readily fold into G-quadruplex (GQ) structures in vitro, and the presence of GQ-prone sequences throughout the genome introduces challenges to re...

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Bibliographic Details
Published inbioRxiv
Main Authors Jansson, Linnea I, Parks, Joseph W, Hentschel, Jendrik, Chang, Terren R, Baral, Rishika, Bagshaw, Clive R, Stone, Michael D
Format Paper
LanguageEnglish
Published Cold Spring Harbor Cold Spring Harbor Laboratory Press 26.10.2018
Subjects
Catalysis
Chromosomes
Deoxyribonucleic acid
DNA
DNA structure
Fluorescence resonance energy transfer
Genomes
Nucleotide sequence
Primers
Ribonucleic acid
RNA
Salts
Telomerase
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Summary:Telomerase maintains telomere length by reverse transcribing short G-rich DNA repeat sequences from its internal RNA template. G-rich telomere DNA repeats readily fold into G-quadruplex (GQ) structures in vitro, and the presence of GQ-prone sequences throughout the genome introduces challenges to replication in vivo. Using a combination of ensemble and single-molecule telomerase assays we discovered that GQ folding of the nascent DNA product during processive addition of multiple telomere repeats modulates the kinetics of telomerase catalysis and dissociation. Telomerase reactions performed with telomere DNA primers of varying sequence or using K+ versus Li+ salts yield changes in DNA product profiles consistent with formation of GQ structure within the telomerase-DNA complex. Single-molecule FRET experiments reveal complex DNA structural dynamics during real-time catalysis, supporting the notion of nascent product folding within the active telomerase complex. To explain the observed distributions of telomere products, we fit telomerase time series data to a global kinetic model that converges to a unique set of rate constants describing each successive telomere repeat addition cycle. Our results highlight the potential influence of the intrinsic folding properties of telomere DNA during telomerase catalysis and provide a detailed characterization of GQ modulation of polymerase function. Footnotes * To correct an author name mis-spelling.
DOI:10.1101/435545