# Cell cycle timing and molecular mechanisms of structural variant formation following incomplete replication

> **NIH NIH R01** · UNIVERSITY OF MICHIGAN AT ANN ARBOR · 2024 · $494,161

## Abstract

Project Summary / Abstract
Mutagenesis resulting from replication failure is a direct cause of tissue dysfunction and cancer when it occurs
in somatic tissues and of de novo and inherited genetic diseases when it occurs in gametogenic stem cells or
meiosis. A primary mutagenic outcome of replication failure is structural variant (SV) formation, especially copy
number variants (CNVs), which create large changes in genomic content in single mutational steps. Fundamental
gaps in knowledge exist regarding the DNA repair mechanisms that lead to SV formation. While multiple
mechanisms may be involved, models that derive from template switching, break-induced replication (BIR), and
other forms of double-strand break (DSB) repair have been forwarded to account for a large proportion of human
CNVs but lack direct experimental evidence. Our prior work has shown that incomplete replication leads to a
high frequency of CNVs in human cells, with hotspots in large, transcribed genes corresponding to common
fragile sites (CFSs) that provide a model system for characterizing SV formation mechanisms.
 Recent literature has revealed much about the damage response pathways that promote proper completion
of replication. One finding was Mitotic DNA Synthesis (MiDAS), where failed replication in S is rescued by a
conservative form of replication activated as late as mitosis. As a BIR-like pathway, MiDAS accuracy is thought
to be low such that the temporal association between MiDAS and CFS expression suggests a potential
mechanistic link to SV formation. Our major goals are to explore the relationships between replication rescue,
end-joining, BIR, other forms of DSB repair, and SV formation, how CFS expression relates to CNV formation,
and how extensible observations at CFS/CNV hotspot loci are to SV formation genome wide. Our central
hypothesis is that hotspot CNV formation occurs during replication rescue, either via MiDAS or an alternative
pathway to MiDAS, notably, theta-mediated end joining (TMEJ). A driving rationale is that we must monitor SV
formation in real time as a primary experimental outcome, something we have been uniquely dedicated to doing.
Our approach will therefore apply our recent technology advances for directly detecting rare SV junctions in
experimental samples to provide answers to longstanding questions about the origins of human SVs.
 We will address our goals through three specific aims to (1) Identify the precise cell cycle stage(s) when
structural variants form following replication stress; (2) Establish the replication rescue and DNA repair pathways
that create structural variant junctions; and (3) Extend mitotic SV formation mechanisms from CFSs to the whole
genome and BRCA2 deficiency. The combination is significant as it will provide direct experimental tests of the
mechanisms that execute SV formation in at-risk genomic loci and extend those findings to multiple genomic
regions and cell lineages relevant to both somatic and heritable ...

## Key facts

- **NIH application ID:** 10827971
- **Project number:** 5R01GM147026-02
- **Recipient organization:** UNIVERSITY OF MICHIGAN AT ANN ARBOR
- **Principal Investigator:** THOMAS W GLOVER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $494,161
- **Award type:** 5
- **Project period:** 2023-04-12 → 2027-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10827971

## Citation

> US National Institutes of Health, RePORTER application 10827971, Cell cycle timing and molecular mechanisms of structural variant formation following incomplete replication (5R01GM147026-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10827971. Licensed CC0.

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