# Mitochondrial G-quadruplex structures in health and disease

> **NIH NIH R01** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2020 · $381,327

## Abstract

PROJECT SUMMARY
G-quadruplex structures (G4) arise in guanine-rich sequences and have a high potential for formation in the
mitochondrial DNA (mtDNA) due to its strand specific biases in nucleotide content. Preliminary studies and
prior published work suggest that G4s impact mitochondrial function, but the evidence remains largely indirect,
obscuring the role of these fascinating structures in normal and pathological mitochondrial biology. This
proposal will address this gap in knowledge by defining the specific regions of mtDNA that form G4 in the cell
and the conditions that promote G4 emergence and stability. The proposal is significant because a fuller
understanding of the regulation of mtDNA maintenance and expression may be important in future approaches
to diverse pathologies including heritable mitochondrial diseases, metabolic syndromes and sporadic cancers.
The proposal is innovative in its development of novel reagents to detect mitochondrial G4s and in its novel
approach to the therapy of mitochondrial disorders. The overarching hypothesis is that physiological G4
formation within mtDNA is widespread and regulates mitochondrial transcription and replication. The first
specific aim will employ an innovative tool, a mitochondrial-targeted intrabody that binds to G4 sequences, to
probe for G4-interacting sequences in the mitochondrial matrix by chromatin immunoprecipitation (ChIP). The
sequences of mtG4s will be identified and their relative abundance will be evaluated under a range of
conditions. The conditions include basal, elevated and inhibited mitochondrial function, and under G4-activated
or G4-inhibited conditions. The role of DNA unwinding enzymes will also be evaluated. The second specific
aim will expand upon a recent observation that induced G4 formation, using G4 binding agents, selects against
specific pathogenic mtDNA variants that enhance G4 formation. Such variants typically exist in a state known
as heteroplasmy, where healthy mtDNA is also present and the ratio between pathogenic and wild type
sequence determines penetrance and severity. We will use patient cell lines and patient-derived cybrid cells to
evaluate the range of pathogenic variants that may be susceptible to this approach. We will also expand the
identification of novel G4 binding compounds that discriminate between pathogenic and wild type alleles.
Overall, these studies will contribute mechanistic evidence for specific G4 structure formation in different
conditions, connect their formation to the regulation of mtDNA replication and transcription, as well as develop
new tools and reagent to positively impact mtDNA content in certain heteroplasmic conditions.

## Key facts

- **NIH application ID:** 9864288
- **Project number:** 1R01GM135443-01
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** Brett A Kaufman
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $381,327
- **Award type:** 1
- **Project period:** 2020-08-01 → 2024-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9864288

## Citation

> US National Institutes of Health, RePORTER application 9864288, Mitochondrial G-quadruplex structures in health and disease (1R01GM135443-01). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9864288. Licensed CC0.

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