# The TET-DNMT-ASXL1-OGT axis: relevance to clonal hematopoiesis, heterochromatin integrity and cancer

> **NIH NIH R35** · LA JOLLA INSTITUTE FOR IMMUNOLOGY · 2024 · $1,098,000

## Abstract

Abstract
During the previous funding period, we made important strides in answering the fundamental mechanistic
question of how TET2, DNMT3A and ASXL1 mutations give rise to clonal hematopoiesis (CH), cancer and
inflammation. We performed biochemical and genomic analyses that linked TET deficiency to an unexpected
loss of DNA methylation in heterochromatin. Given that DNMT3A and TET proteins have opposite biochemical
functions, our finding of decreased DNA methylation in heterochromatin of both Dnmt3a KO and Tet2 KO cells
is the only common feature that explains the unexpectedly greater disease severity of Dnmt3a KO, Tet2 KO
(DKO) mice compared to mice with individual Dnmt3a or Tet2 gene disruptions. We also showed that CH- and
cancer-associated mutations in ASXL1 result in reduced histone 3 lysine methylation (H3K9me2/me3) in hetero-
chromatin, thus linking all three of the top proteins mutated in clonal hematopoiesis – TET2, DNMT3A and ASXL1
– to the fundamental process of impaired heterochromatin integrity due to reduced DNA or H3K9 methylation in
heterochromatin. Interference with DNA or H3K9 methylation in heterochromatin has been known for decades
to result in increased expression of transposable elements, genome instability and inflammation, all common
features of cancer, inflammation and aging. Finally, we showed that OGT normally restrains TET activity in
mESC, and that consequently, OGT deficiency increases TET activity genome-wide and results in genome-wide
DNA demethylation. In this renewal application, we will ask how the TET-OGT interaction regulates DNA
methylation, focusing on the protein-protein interactions and regulatory mechanisms that operate within TET-
OGT complexes in mESC and hematopoietic-lineage cells. We will define how DNMTs and TETs cooperate to
reduce DNA methylation in heterochromatin by examining DNMT3A redistribution and DNMT1/UHRF1 stability.
We will explore the mechanism of how CH-associated mutations in ASXL1 lead to reduced H3K9 methylation in
heterochromatin, again focusing on protein-protein interactions and regulatory mechanisms that operate within
two distinct ASXL1-associated protein complexes that we have recently defined. We will identify individual
members of TE families that are uniquely upregulated in TET2, DNMT3A and ASXL1-mutant cells, so as to
determine how increased TE expression can have potentially indirect and stochastic effects on the expression
of nearby genes by acting as promoters, enhancers or both. Lastly, we will perform CRISPR/Cas9 screens to
elucidate how TET deficiency and ASXL1 mutations are linked to inflammation. Our studies will illuminate the
mechanistically elusive connections between DNA and H3K9 methylation, clonal hematopoiesis, cancer,
inflammation and aging, and improve our understanding of heterochromatin, a compartment of the genome that
is very poorly understood.

## Key facts

- **NIH application ID:** 10983243
- **Project number:** 2R35CA210043-08A1
- **Recipient organization:** LA JOLLA INSTITUTE FOR IMMUNOLOGY
- **Principal Investigator:** Anjana Rao
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $1,098,000
- **Award type:** 2
- **Project period:** 2016-09-01 → 2031-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10983243

## Citation

> US National Institutes of Health, RePORTER application 10983243, The TET-DNMT-ASXL1-OGT axis: relevance to clonal hematopoiesis, heterochromatin integrity and cancer (2R35CA210043-08A1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10983243. Licensed CC0.

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