# DEVELOP GENOMEWIDE TECHNOLOGY TO PROFILE DNA ADP-RIBOSYLATION

> **NIH NIH R21** · JOHNS HOPKINS UNIVERSITY · 2024 · $204,688

## Abstract

PROJECT SUMMARY
Our research program seeks to develop innovative technologies to investigate the role of ADP-ribosylation, an
under-studied DNA modification implicated in critical biological processes, including DNA repair, genome
replication, and pathogen defense. Though prevalent across all kingdoms of life, from bacteria to humans, the
precise function and extent of DNA ADP-ribosylation remain elusive. Current knowledge largely stems from
studies on the bacterial DarT-DarG toxin-antitoxin system, and more recently, the discoveries of DNA
modification by the anticancer drug target PARP1 in human cells. One significant challenge in the field is the
absence of technology capable of identifying DNA ADP-ribosylation sites across the whole genome. Past
efforts have involved meticulous mutagenesis using radioactive labeling on predefined oligo sequences or
breaking down the genome to individual bases for modification assessment. However, these methods are
limited in their ability to provide the broader genomic context of DNA ADP-ribosylation.
Our goal is to bridge this gap through the development of two complementary techniques, using the well-
characterized DarT-mediated ADP-ribosylation as our model. The first repurposes tools we have developed to
study protein ADP-ribosylation and applies them to DNA, using high-throughput Illumina-based sequencing to
pinpoint the modification site. The second approach involves nanopore sequencing, using a modified
transmembrane protein as both a channel for DNA passage and a biosensor for base identification. By
monitoring changes in electric current, we can distinguish specific bases and detect DNA modifications. To
optimize this method, we will collaborate with leading nanopore epigenetics expert Dr. Winston Timp at
Hopkins and adapt established machine learning approaches for electric current data analyses. We will test
and refine our methods using DarT-modified oligos with defined sequences and genomic DNA standards.
In summary, this program aims to unravel the complexities of DNA ADP-ribosylation using cutting-edge
sequencing methods. These strategies will enable the investigation of the sequence context of ADP-
ribosylation sites and, in due course (beyond the current scope), the study of motifs from diverse pathogen
toxins and other modifying enzymes such as human PARPs. By developing tools to reveal the mechanisms of
DNA repair, gene regulation, and pathogen defense, we anticipate that our work may catalyze significant
advancements in healthcare, potentially leading to the development of new treatment and detection tools for a
range of diseases, from infections to cancers.

## Key facts

- **NIH application ID:** 10870413
- **Project number:** 1R21GM154218-01
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Anthony K L Leung
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $204,688
- **Award type:** 1
- **Project period:** 2024-09-01 → 2026-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10870413

## Citation

> US National Institutes of Health, RePORTER application 10870413, DEVELOP GENOMEWIDE TECHNOLOGY TO PROFILE DNA ADP-RIBOSYLATION (1R21GM154218-01). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10870413. Licensed CC0.

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