# Mechanisms of Genome Instability Mediated by Simple DNA Repeats

> **NIH NIH R35** · TUFTS UNIVERSITY MEDFORD · 2024 · $573,959

## Abstract

Project Summary
The focus of my lab is to understand the mechanisms of genome instability caused by structure-prone
DNA repeats. We are particularly interested in the mechanisms of repeat expansions that are responsible
for over fifty hereditary diseases in humans. Recent advances in long-read sequencing revealed a new
paradigm: massive genome-wide expansions of structure-prone DNA repeats in human cancers. Thus,
understanding the mechanisms responsible for large-scale repeat expansions is fundamentally important
and has broad biomedical implications.
My lab was the first to show that expandable DNA repeats stall replication fork progression in every
experimental system studied, including bacteria, yeast, and human cells. This led us to propose that
repeats can be added while replication fork escapes from a “repetitive trap”. We first confirmed this idea in
a yeast experimental system. Depending on the mode of replication fork progression through a repeat,
expansions occur by incorporation of unprocessed flaps during Okazaki fragments maturation, replicative
or post-replicative template-switching, or break-induced replication. Recently, we developed a new
experimental system to study large-scale expansions in human cells, which implicates DNA replication as
well. Repeat expansions also occur in terminally differentiated somatic cells that do not undergo DNA
replication. We have, therefore, adjusted our experimental system to study repeat expansions in non-
dividing yeast cells. Our results point to DNA nick repair as a possible mechanism.
We plan to move our research in several directions. First, we will elucidate the role of DNA nick repair in
expansions of Friedreich’s ataxia (FRDA) (GAA)n repeats in dividing and non-dividing yeast cells by
introducing targeted nicks with Cas9 nickases. We will establish its genetic controls and study the role of
the human FAN1 nuclease expressed in yeast. Second, we will unravel the mechanisms of large-scale
repeat expansions in human cells by conducting candidate gene analysis in a plasmid system utilizing
SV40 replication machinery. We will also study the effects of compounds that disrupt or stabilize DNA triplexes
formed by these repeats on their replication and expansion. We will further extend our studies into a different
system based on the EBNA1-dependent replication which involves regular cellular replication fork. Third, we will
carry out structure-functional analysis of other expandable homopurine-homopyrimidine repeats, including the
(AAGGG)n repeat, which is responsible for cerebellar ataxia, neuropathy, vestibular areflexia syndrome
(CANVAS), the (CCCTCT)n repeat that modifies the expressivity of X-linked dystonia parkinsonism (XDP), and
the (GAAA)n repeat, which recurrently expands in many human cancers. At present, there are no data on DNA
structures formed by those repeats or on the mechanisms of their expansions. We will address these matters by
using the broad arsenal of methods and experimen...

## Key facts

- **NIH application ID:** 10764034
- **Project number:** 2R35GM130322-06
- **Recipient organization:** TUFTS UNIVERSITY MEDFORD
- **Principal Investigator:** SERGEI MIRKIN
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $573,959
- **Award type:** 2
- **Project period:** 2019-03-14 → 2029-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10764034

## Citation

> US National Institutes of Health, RePORTER application 10764034, Mechanisms of Genome Instability Mediated by Simple DNA Repeats (2R35GM130322-06). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10764034. Licensed CC0.

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