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Mechanisms of telomerase inhibition by oxidized and therapeutic dNTPs

Sanford, Samantha (2021) Mechanisms of telomerase inhibition by oxidized and therapeutic dNTPs. Doctoral Dissertation, University of Pittsburgh. (Unpublished)

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Telomeres cap chromosome ends and are essential for genome stability and human health, but they shorten in most human somatic cells with cell division due to the end replication problem. Telomerase a specialized reverse transcriptase that lengthens telomeres by adding GGTTAG repeats to chromosome ends and is upregulated in most human cancers to enable limitless proliferation. Here, we uncover two distinct mechanisms by which naturally occurring oxidized and therapeutic dNTP DNA precursors inhibit telomerase-mediated telomere elongation. We conducted a series of direct telomerase extension assays in the presence of modified dNTPs on various telomeric substrates. We provide direct evidence that telomerase can add the metabolized form of NRTIs, dideoxyadenosine 5’ triphosphate (ddITP) and 3’-azido-3’deoxythymidine triphosphate (AZT-TP), to the telomeric end, causing chain termination. In contrast, telomerase continues elongation after inserting oxidized 2-OH-dATP or the therapeutic 6-thioguanine metabolite, 6-thio-dGTP, but insertion disrupts translocation and inhibits further repeat addition. Kinetics reveal that telomerase poorly selects against 6-thio-dGTP, inserting with similar catalytic efficiency as dGTP. Furthermore, telomerase processivity factor POT1-TPP1 fails to restore processive elongation in the presence of inhibitory dNTPs. These findings reveal mechanisms for targeting telomerase with modified dNTPs in cancer therapy.


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Item Type: University of Pittsburgh ETD
Status: Unpublished
CreatorsEmailPitt UsernameORCID
Sanford, Samanthasas354@pitt.edusas354
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Committee ChairOpresko, Patriciaplo4@pitt.eduplo4
Committee CoChairAyyavoo, Velpandivelpandi@pitt.eduvelpandi
Committee MemberVan Houten, Bennettvanhoutenb@upmc.eduvanhoutenb
Committee MemberBerman, Andreaajb190@pitt.eduajb190
Committee MemberMartinson, Jeremyjmartins@pitt.edujmartins
Date: 19 January 2021
Date Type: Publication
Defense Date: 12 August 2020
Approval Date: 19 January 2021
Submission Date: 12 November 2020
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 190
Institution: University of Pittsburgh
Schools and Programs: School of Public Health > Infectious Diseases and Microbiology
Degree: PhD - Doctor of Philosophy
Thesis Type: Doctoral Dissertation
Refereed: Yes
Uncontrolled Keywords: telomeres, telomerase
Date Deposited: 19 Jan 2021 20:50
Last Modified: 19 Jan 2021 20:50

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