# Cell biological and proteomic investigation of pathogenic DDX3X missense mutations during neurogenesis

> **NIH NIH R21** · DUKE UNIVERSITY · 2022 · $241,500

## Abstract

Abstract
De novo mutations in DDX3X, an X-linked RNA helicase, account for 1-3% of intellectual disability (ID) in females
and are associated with a broad range of phenotypes, including developmental delay, epilepsy, autism and brain
malformations. Approximately half of the mutations are nonsense, whereas the other 50% are missense
mutations. Initial reports suggested that DDX3X mutations primarily cause loss-of-function phenotypes.
However, our collaborators identified recurrent missense mutations in DDX3X individuals that were consistently
associated with more severe clinical outcomes and specific brain malformations not observed in other DDX3X
individuals. These data strongly suggest some DDX3X missense mutations exert dominant negative phenotypes.
We have further discovered that DDX3X is required for proper brain development; specifically, loss of Ddx3x in
the embryonic mouse brain impairs the ability of neural progenitors to make neurons. We also demonstrated that
DDX3X missense mutations associated with severe impairment have nearly complete loss of helicase activity,
impaired translation of key targets, and form ectopic RNA-protein granules. These findings indicate that ectopic
RNA-protein granules and aberrant DDX3X protein interactions may contribute to disease severity. However,
these mechanisms have not been examined. In this proposal, we seek to elucidate the molecular pathogenesis
of DDX3X missense mutations linked to severe clinical deficits in DDX3X syndrome. We will test the hypothesis
that clinically severe DDX3X missense mutations perturb its cellular dynamics and protein interactome
during neurogenesis. First, we will characterize DDX3X granules in neural progenitors and neurons expressing
mild and severe DDX3X missense mutations. We will also determine whether granules are linked to impaired
neurogenesis. Second, we will use an unbiased proteomic screen in neural progenitors and neurons to determine
how different DDX3X missense mutations impair the WT protein interactome. These studies are critical for
understanding the molecular underpinnings of dominant negative phenotypes associated with DDX3X
syndrome. Further, our studies will broadly establish a paradigm for understanding other neurodevelopmental
disorders in which RNA regulation plays a central role.

## Key facts

- **NIH application ID:** 10474429
- **Project number:** 5R21HD104514-02
- **Recipient organization:** DUKE UNIVERSITY
- **Principal Investigator:** Debra L. Silver
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $241,500
- **Award type:** 5
- **Project period:** 2021-09-01 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10474429

## Citation

> US National Institutes of Health, RePORTER application 10474429, Cell biological and proteomic investigation of pathogenic DDX3X missense mutations during neurogenesis (5R21HD104514-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10474429. Licensed CC0.

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