# Regulation and recognition of H3K79me

> **NIH NIH F31** · JOHNS HOPKINS UNIVERSITY · 2021 · $46,036

## Abstract

Project Summary/Abstract
Patterns of dynamic post-translational modifications on histone proteins of nucleosomes regulate local and
global eukaryotic gene expression by recruiting specialized effector proteins. Mono-, di-, and tri- methylation of
histone H3 lysine 79 (H3K79me1/2/3) is a hallmark of actively transcribed genes and is central to many
fundamental biological processes. Importantly, aberrant hypermethylation of H3K79 is found in the Mixed
Lineage Leukemia 1-rearranged (MLL-r) leukemias which are resistant to treatment. New potent treatments for
these leukemias are desperately needed, but the underlying mechanisms of H3K79me regulation and
recognition are poorly understood. Dot1L is the sole methyltransferase responsible for establishing H3K79me.
It is stimulated by mono-ubiquitylation of histone H2B (H2Bub). In cells, Dot1L is found in complex with partner
proteins AF9 and AF10, which are known to upregulate H3K79me levels. However, very little is known about
their effects on Dot1L binding affinity and activity on H2Bub-modified nucleosomes. In Aim 1, I will compare
the binding affinity and activity of Dot1L + AF9 and Dot1L + AF10 on nucleosomes containing H2Bub and
H3K79me1/2/3 modifications using biochemical techniques. To guide future structure-based leukemia
therapies, I will also determine the cryo-EM structure of Dot1L + AF9 + AF10 on H2Bub nucleosome.
H3K79me2/3 is known to regulate fundamental biological processes, yet no H3K79me effector proteins have
been identified. H3K79 is located within the globular core of the nucleosome and may require the use of
nucleosomal substrates as bait in pulldown experiments to identify H3K79me binding proteins. However, no
pulldown studies have used H3K79me-modified nucleosomes to identify binding proteins. In Aim 2, I will
identify H3K79me binding proteins using H3K79me2/3-modified nucleosomes as bait in protein pulldowns from
mammalian nuclear extract. I will identify potential hits using mass-spectrometry based proteomics and
validate them for specific H3K79me binding using a biochemical technique. Finally, I will study the mechanism
of H3K79me recognition by determining a cryo-EM structure of a high-affinity H3K79me binding protein on
nucleosome. The outcomes from these proposed experiments in Aims 1 and 2 will form a basis for guiding the
design of drug-based leukemia therapies targeting Dot1L partner proteins and will reveal how H3K79 methyl
marks are recognized by effector proteins to modulate gene expression.

## Key facts

- **NIH application ID:** 10235041
- **Project number:** 1F31CA261154-01
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Chad Hicks
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $46,036
- **Award type:** 1
- **Project period:** 2021-04-01 → 2024-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10235041

## Citation

> US National Institutes of Health, RePORTER application 10235041, Regulation and recognition of H3K79me (1F31CA261154-01). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10235041. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*