# ADNP mechanisms in R-loop regulation during differentiation

> **NIH NIH R01** · WISTAR INSTITUTE · 2022 · $431,640

## Abstract

Project Summary
Aberrations in chromatin structure can be a causal mechanism of numerous human disease and developmental
syndromes. R-loops are RNA containing chromatin structures that are deregulated in numerous developmental
disorders and cancers. In most cases the underlying mechanisms and the functional significance of R-loop
deregulation and whether they are causal to specific disease is not clear. Activity dependent neuroprotective
protein (ADNP) is a homeodomain containing transcriptional repressor that is critical for neuronal differentiation
and neurodevelopment. ADNP mutations cause ADNP syndrome, a condition characterized by intellectual
disability and features of autism spectrum disorder. ADNP is upregulated during neurodevelopment. ADNP
contains zinc finger motifs and a homeodomain, both of which can bind nucleic acids. ADNP syndrome mutations
result in protein products that lack the homeodomain, underscoring the importance of this region to ADNP
function. However, the precise contributions of the zinc fingers and homeodomain to ADNP localization and
function in gene expression during neuronal differentiation is not well understood. Our preliminary data have
uncovered a novel role for ADNP in the resolution of R-loops. Biochemical assays revealed that ADNP resolves
R-loops – in a homeodomain dependent manner. ADNP also suppresses R-loops in vivo, as deletion of ADNP
in mouse embryonic stem cells (mESCs) resulted in R-loop accumulation specifically at ADNP target sites, but
not at sites not bound by ADNP. Last, ADNP-deficient mESCs and mESCs exclusively expressing an ADNP
mutant lacking the homeodomain fail to differentiate into neural progenitor cells (NPCs), indicating an essential
role for ADNP and its homeodomain in neuronal differentiation. These results suggest a potentially novel
mechanism – R-loop resolution – for ADNP, and possibly other homeodomain-containing proteins, in gene
regulation, which can link R-loop dysfunction to numerous disorders and diseases caused by mutations in
homeodomain proteins. Based on our preliminary data, we hypothesize that ADNP drives neuronal
differentiation by suppressing R-loops; as a corollary, we propose that resolving R-loops that accumulate upon
ADNP loss may rescue gene expression programs to enable neuronal differentiation. We propose two aims to
test our central hypothesis. In Aim 1, we will elucidate the molecular basis of R-loop resolution by ADNP by
identifying roles of the ADNP protein domains and their interactions with nucleic acids. In Aim 2, we will elucidate
how ADNP suppression of R-loops influences neuronal differentiation.

## Key facts

- **NIH application ID:** 10444141
- **Project number:** 1R01GM143229-01A1
- **Recipient organization:** WISTAR INSTITUTE
- **Principal Investigator:** Kavitha Sarma
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $431,640
- **Award type:** 1
- **Project period:** 2022-09-06 → 2026-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10444141

## Citation

> US National Institutes of Health, RePORTER application 10444141, ADNP mechanisms in R-loop regulation during differentiation (1R01GM143229-01A1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10444141. Licensed CC0.

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