# Bacterial Rogue Methyltransferases Inducing Human Epimutations

> **NIH NIH F32** · CLEVELAND CLINIC LERNER COM-CWRU · 2022 · $21,430

## Abstract

PROJECT SUMMARY: DNA methylation is dysregulated in every form of cancer. While there is substantial
knowledge regarding the location and function of abnormal DNA methylation across tumor types, we are severely
lacking in understanding how such abnormal DNA methylation is established during cancer formation. Colorectal
cancer (CRC), the third most common cancer type worldwide, has been heavily associated with alterations in
DNA methylation, however, mutations in DNA methylation/demethylation pathway genes are rare and cannot
account for the exaggerated DNA hypermethylation. Roughly 1.5 kg of bacteria reside in our gut and have been
shown to play a substantial role in the development of CRC. Recent evidence suggests that these bacteria can
alter the colon cell epigenome by inducing aberrant DNA hypermethylation, although the mechanism promoting
this is still not completely understood. One such microorganism, Fusobacterium nucleatum, is able to invade into
colon epithelial cells and has been heavily associated with CRC development, as well as hypermethylation of
tumor suppressor genes. Roughly 90% of bacteria, including Fusobacterium nucleatum, contain DNA
methyltransferase enzymes as a component of their restriction-modification system; a defense mechanism
aimed at protecting their genome against invading viruses by methylating specific motifs within their DNA as to
distinguish between their own and invading viral DNA. To date, >3500 different R-M motifs have been identified,
thus our gut microbiome is filled with microbial genome-modifying enzymes, however their potential to modify
human DNA has not been evaluated. The long-term objective of this proposal is to evaluate whether F. nucleatum
DNA methyltransferase enzymes are able to access and aberrantly methylate colon cell DNA, potentially giving
rise to the epimutations observed in CRC development. This will be tested with three specific aims. The first aim
will experimentally evaluate F. nucleatum methyltransferase activity in vitro, by ectopically expressing F.
nucleatum DNA methyltransferase enzymes in CRC cell lines and evaluating their ability to enter the nucleus,
methylate DNA, as well as determine the locations of DNA methylation. The second aim will take advantage of
the microbiome core facilities at Lerner Research Institute to preform bacterial co-culture experiments using F.
nucleatum that have been genetically modified to contain tagged-DNA methyltransferase enzyme, to test
whether F. nucleatum DNA methyltransferase can access and methylate colon cell DNA. The third aim is to
detect F. nucleatum activity in vivo, by searching for F. nucleatum R-M motif enrichment at CRC hypermethylated
loci, using our labs published MBD-seq data and the TCGA CRC cohort. Results could reveal a novel paradigm
in host-microbiome interactions, as direct epigenetic crosstalk between microbial and host cells has not been
fully explored. However, if true, this new paradigm will transform studies of bacter...

## Key facts

- **NIH application ID:** 10439471
- **Project number:** 5F32CA260774-02
- **Recipient organization:** CLEVELAND CLINIC LERNER COM-CWRU
- **Principal Investigator:** Emily E Fink
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $21,430
- **Award type:** 5
- **Project period:** 2021-06-01 → 2022-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10439471

## Citation

> US National Institutes of Health, RePORTER application 10439471, Bacterial Rogue Methyltransferases Inducing Human Epimutations (5F32CA260774-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10439471. Licensed CC0.

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