# Determining the effects of ribonucleotides on telomere integrity

> **NIH NIH F31** · UNIVERSITY OF KANSAS MEDICAL CENTER · 2022 · $35,263

## Abstract

Project Summary/Abstract
Ribonucleotides (ribos) are misinserted by DNA polymerases and have recently been recognized as the most
common lesion across most of the genome. Compared to deoxyribonucleotides, ribos have an additional 2'-OH
that makes them much more susceptible to spontaneous hydrolysis. Additionally, unrepaired ribos promote
genomic instability and are associated with various human diseases. In comparison to genomic ribos, the impact
of ribos in telomeres is unknown. Telomeres are protective noncoding DNA structures at the ends of
chromosomes that are essential for maintaining genomic integrity, as highlighted by the role of telomere
dysfunction in a myriad of human diseases. Due to the lack of a tool to selectively perturb ribo insertion rates
into telomeres, the prevalence, and effects of telomeric ribos remains unexplored. To this end, we have
developed solutions for investigating the role of telomeric ribos by discovering a variant telomerase that lacks
any ribo discrimination activity and by developing an assay to determine the levels of telomeric ribos.
Furthermore, I have identified two telomerase disease associated variants (A716V and Y717N) which appear to
perturb telomerase ribo discrimination. The overarching goal of this proposal is to determine the levels and
effects of telomeric ribos, as well as establish a causal role for increased telomeric ribo insertion and human
disease. I hypothesize that ribonucleotides are present in telomeres and that increased ribo insertion promotes
telomere uncapping and genomic instability. To test this, I propose the following aims: 1) Characterize the kinetic
and structural effects of telomerase disease-associated variants and 2) Determine how each variant effects
telomere integrity and cell viability. To accomplish these aims I will characterize the pre-steady state kinetics of
A716V and Y717N using the telomerase homolog Tribolium castaneum telomerase reverse transcriptase
(tcTERT) (Aim 1A), as well as use human telomerase to determine differences in telomerase processivity (Aim
1B). I will complement the kinetic analysis by characterizing the structural rearrangements of A716V and Y717N
in tcTERT using X-ray crystallography (Aim 1C). Following the comprehensive biochemical characterization of
each variant, I will determine the levels of telomeric ribos in cell lines expressing each telomerase variant (Aim
2A). Finally, the effects of increased telomeric ribos will be determined using immunofluorescence microscopy
to identify telomere dysfunction induced foci (Aim 2B), which are a commonly used marker of a telomere DNA
damage response. This innovative project will be completed at the University of Kansas Medical Center under
the supervision of an expert sponsorship team. In addition to technical skills, the training plan also prioritizes
training in scientific communication and mentorship. The proposed experiments will provide me with the
foundation for a successful post-doctoral fello...

## Key facts

- **NIH application ID:** 10464674
- **Project number:** 1F31GM146446-01
- **Recipient organization:** UNIVERSITY OF KANSAS MEDICAL CENTER
- **Principal Investigator:** Griffin Alan Welfer
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $35,263
- **Award type:** 1
- **Project period:** 2022-05-01 → 2024-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10464674

## Citation

> US National Institutes of Health, RePORTER application 10464674, Determining the effects of ribonucleotides on telomere integrity (1F31GM146446-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10464674. Licensed CC0.

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