DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis

Abstract

The DEAD-box RNA helicase DDX3X is frequently mutated in pediatric medulloblastoma. We dissect how these mutants affect DDX3X function with structural, biochemical, and genetic experiments. We identify an N-terminal extension (“ATP-binding loop”, ABL) that is critical for the stimulation of ATP hydrolysis by RNA. We present crystal structures suggesting that the ABL interacts dynamically with ATP and confirming that the interaction occurs in solution by NMR chemical shift perturbation and isothermal titration calorimetry. DEAD-box helicases require interaction between two conserved RecA-like helicase domains, D1 and D2 for function. We use NMR chemical shift perturbation to show that DDX3X interacts specifically with double-stranded RNA through its D1 domain, with contact mediated by residues G302 and G325. Mutants of these residues, G302V and G325E, are associated with pediatric medulloblastoma. These mutants are defective in RNA-stimulated ATP hydrolysis. We show that DDX3X complements the growth defect in a ded1 temperature-sensitive strain of Schizosaccharomyces pombe, but the cancer-associated mutants G302V and G325E do not complement and exhibit protein expression defects. In conclusion, taken together, our results suggest that impaired translation of important mRNA targets by mutant DDX3X represents a key step in the development of medulloblastoma.

Authors:
; ; ; ;
Publication Date:
Research Org.:
Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES)
OSTI Identifier:
1242551
Alternate Identifier(s):
OSTI ID: 1178837
Grant/Contract Number:  
W-31-109-Eng-38; CCSG 2 P30 CA21765
Resource Type:
Published Article
Journal Name:
Journal of Molecular Biology
Additional Journal Information:
Journal Name: Journal of Molecular Biology Journal Volume: 427 Journal Issue: 9; Journal ID: ISSN 0022-2836
Publisher:
Elsevier
Country of Publication:
United Kingdom
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; crystallography; NMR; biochemistry; genetics

Citation Formats

Epling, Leslie B., Grace, Christy R., Lowe, Brandon R., Partridge, Janet F., and Enemark, Eric J. Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis. United Kingdom: N. p., 2015. Web. doi:10.1016/j.jmb.2015.02.015.
Epling, Leslie B., Grace, Christy R., Lowe, Brandon R., Partridge, Janet F., & Enemark, Eric J. Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis. United Kingdom. https://doi.org/10.1016/j.jmb.2015.02.015
Epling, Leslie B., Grace, Christy R., Lowe, Brandon R., Partridge, Janet F., and Enemark, Eric J. Fri . "Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis". United Kingdom. https://doi.org/10.1016/j.jmb.2015.02.015.
@article{osti_1242551,
title = {Cancer-Associated Mutants of RNA Helicase DDX3X Are Defective in RNA-Stimulated ATP Hydrolysis},
author = {Epling, Leslie B. and Grace, Christy R. and Lowe, Brandon R. and Partridge, Janet F. and Enemark, Eric J.},
abstractNote = {The DEAD-box RNA helicase DDX3X is frequently mutated in pediatric medulloblastoma. We dissect how these mutants affect DDX3X function with structural, biochemical, and genetic experiments. We identify an N-terminal extension (“ATP-binding loop”, ABL) that is critical for the stimulation of ATP hydrolysis by RNA. We present crystal structures suggesting that the ABL interacts dynamically with ATP and confirming that the interaction occurs in solution by NMR chemical shift perturbation and isothermal titration calorimetry. DEAD-box helicases require interaction between two conserved RecA-like helicase domains, D1 and D2 for function. We use NMR chemical shift perturbation to show that DDX3X interacts specifically with double-stranded RNA through its D1 domain, with contact mediated by residues G302 and G325. Mutants of these residues, G302V and G325E, are associated with pediatric medulloblastoma. These mutants are defective in RNA-stimulated ATP hydrolysis. We show that DDX3X complements the growth defect in a ded1 temperature-sensitive strain of Schizosaccharomyces pombe, but the cancer-associated mutants G302V and G325E do not complement and exhibit protein expression defects. In conclusion, taken together, our results suggest that impaired translation of important mRNA targets by mutant DDX3X represents a key step in the development of medulloblastoma.},
doi = {10.1016/j.jmb.2015.02.015},
journal = {Journal of Molecular Biology},
number = 9,
volume = 427,
place = {United Kingdom},
year = {Fri May 01 00:00:00 EDT 2015},
month = {Fri May 01 00:00:00 EDT 2015}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
https://doi.org/10.1016/j.jmb.2015.02.015

Citation Metrics:
Cited by: 43 works
Citation information provided by
Web of Science

Save / Share: