Division of Labor in an Oligomer of the DEAD-Box RNA Helicase Ded1p

Putnam, Andrea; Gao, Zhaofeng; Liu, Fei; Jia, Huijue; Yang, Quansheng; Jankowsky, Eckhard

doi:10.1016/j.molcel.2015.06.030

Cited by 62 publications

(121 citation statements)

References 42 publications

(104 reference statements)

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“…Oligomerization depends on the C-terminal tail of Ded1, and truncation of the C-terminal 69 residues blocks multimerization and hinders duplex unwinding (32). Our data show that removal of this region strongly suppresses duplex unwinding in vitro, but only if the conserved RDYR motif is deleted (Figs.…”

Section: Discussionmentioning

confidence: 58%

“…It is possible that removal of this positively charged region in the CTE negatively impacts RNA binding, as suggested by weaker binding to heparin resin (data not shown) and by the RNA binding defect caused by deletion of the entire C-terminal tail of Ded1p up to the helicase core (29). Alternatively, or in addition, this region might be critical for oligomerization (32).…”

Section: Resultsmentioning

confidence: 98%

“…2A). Both full-length and truncated protein show sigmoidal functional binding isotherms, suggesting that DDX3 functions as an oligomer, as yeast DED1 (32).…”

Section: Resultsmentioning

confidence: 99%

“…Interestingly, the Ded1/DDX3 subfamily of DEAD-box proteins multimerizes both in vitro and in vivo (32). Oligomerization depends on the C-terminal tail of Ded1, and truncation of the C-terminal 69 residues blocks multimerization and hinders duplex unwinding (32).…”

Section: Discussionmentioning

confidence: 99%

“…The tails of Ded1/DDX3 subfamily members are thought to be largely unstructured and contain diverse motifs with different functions. For example, the N-terminal tail of DDX3 contains a Crm1-dependent nuclear export sequence (31) and an eIF4E-binding motif (10, 11), whereas the C-terminal tail contains conserved sequences of unknown function that are essential for oligomerization (5,32). The tails of DED1 additionally contain assembly domains that modulate translation and alleviate lethality associated with protein overexpression when deleted (10).…”

mentioning

confidence: 99%

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Autoinhibitory Interdomain Interactions and Subfamily-specific Extensions Redefine the Catalytic Core of the Human DEAD-box Protein DDX3

Floor

Condon

Sharma

et al. 2016

Journal of Biological Chemistry

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112

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DEAD-box proteins utilize ATP to bind and remodel RNA and RNA-protein complexes. All DEAD-box proteins share a conserved core that consists of two RecA-like domains. The core is flanked by subfamily-specific extensions of idiosyncratic function. The Ded1/DDX3 subfamily of DEAD-box proteins is of particular interest as members function during protein translation, are essential for viability, and are frequently altered in human malignancies. Here, we define the function of the subfamily-specific extensions of the human DEAD-box protein DDX3. We describe the crystal structure of the subfamily-specific core of wild-type DDX3 at 2.2 Å resolution, alone and in the presence of AMP or nonhydrolyzable ATP. These structures illustrate a unique interdomain interaction between the two ATPase domains in which the C-terminal domain clashes with the RNA-binding surface. Destabilizing this interaction accelerates RNA duplex unwinding, suggesting that it is present in solution and inhibitory for catalysis. We use this core fragment of DDX3 to test the function of two recurrent medulloblastoma variants of DDX3 and find that both inactivate the protein in vitro and in vivo. Taken together, these results redefine the structural and functional core of the DDX3 subfamily of DEADbox proteins.DEAD-box proteins are ATP-dependent RNA-binding proteins that remodel RNA structures and RNA-protein complexes, stably clamp RNA, and promote fluidity within RNA granules (1-3). The human DEAD-box protein DDX3 (encoded by DDX3X) and its yeast ortholog Ded1p have been implicated in numerous functions including translation initiation (4 -12). Messenger RNA molecules containing especially long or structured 5Ј leader sequences are particularly sensitive to DDX3 activity (6,7,12,13). DDX3X is frequently mutated in numerous cancer types (5), such as chronic lymphocytic leukemia (14 -16), natural killer/T-cell lymphoma (17), head and neck squamous cell carcinoma (18,19), and lung cancer (20). DDX3X is also one of the most frequently mutated genes in the highly malignant brain tumor medulloblastoma (21-24). In medulloblastoma, many mutations are predicted to inactivate DDX3, and some have been demonstrated to diminish activities in vitro (17,25).DEAD-box proteins are defined by 12 different motifs that function in ATP binding or hydrolysis and RNA binding, or couple ATP and RNA binding (1). Outside of these conserved motifs, each DEAD-box protein subfamily has unique tails that lie N-or C-terminal to the helicase core and contain elements that define the unique properties of that subfamily. For example, DDX21 has a GUCT domain in its C-terminal extension (26, 27), DDX5 has tandem P68HR domains in its C-terminal extension, and DDX43 has a KH1 domain in its N-terminal extension. However, as the tails of each DEAD-box protein subfamily are idiosyncratic, whereas the cores are very similar (28,29), it is essential to study individual subfamilies of DEAD-box proteins in detail to understand the role of subfamily-specific tails.DDX3 is a member of the Ded1/DDX...

show abstract

Section: Discussionmentioning

confidence: 58%

Section: Resultsmentioning

confidence: 98%

“…2A). Both full-length and truncated protein show sigmoidal functional binding isotherms, suggesting that DDX3 functions as an oligomer, as yeast DED1 (32).…”

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Autoinhibitory Interdomain Interactions and Subfamily-specific Extensions Redefine the Catalytic Core of the Human DEAD-box Protein DDX3

Floor

Condon

Sharma

et al. 2016

Journal of Biological Chemistry

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112

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Structural Basis of Human Helicase DDX21 in RNA Binding, Unwinding, and Antiviral Signal Activation

Chen

et al. 2020

Advanced Science

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RNA helicase DDX21 plays vital roles in ribosomal RNA biogenesis, transcription, and the regulation of host innate immunity during virus infection. How DDX21 recognizes and unwinds RNA and how DDX21 interacts with virus remain poorly understood. Here, crystal structures of human DDX21 determined in three distinct states are reported, including the apo‐state, the AMPPNP plus single‐stranded RNA (ssRNA) bound pre‐hydrolysis state, and the ADP‐bound post‐hydrolysis state, revealing an open to closed conformational change upon RNA binding and unwinding. The core of the RNA unwinding machinery of DDX21 includes one wedge helix, one sensor motif V and the DEVD box, which links the binding pockets of ATP and ssRNA. The mutant D339H/E340G dramatically increases RNA binding activity. Moreover, Hill coefficient analysis reveals that DDX21 unwinds double‐stranded RNA (dsRNA) in a cooperative manner. Besides, the nonstructural (NS1) protein of influenza A inhibits the ATPase and unwinding activity of DDX21 via small RNAs, which cooperatively assemble with DDX21 and NS1. The structures illustrate the dynamic process of ATP hydrolysis and RNA unwinding for RNA helicases, and the RNA modulated interaction between NS1 and DDX21 generates a fresh perspective toward the virus–host interface. It would benefit in developing therapeutics to combat the influenza virus infection.

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Regulatory effects of cotranscriptional RNA structure formation and transitions

Liu

Zhang

2016

WIREs RNA

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RNAs, which play significant roles in many fundamental biological processes of life, fold into sophisticated and precise structures. RNA folding is a dynamic and intricate process, which conformation transition of coding and noncoding RNAs form the primary elements of genetic regulation. The cellular environment contains various intrinsic and extrinsic factors that potentially affect RNA folding in vivo, and experimental and theoretical evidence increasingly indicates that the highly flexible features of the RNA structure are affected by these factors, which include the flanking sequence context, physiochemical conditions, cis RNA-RNA interactions, and RNA interactions with other molecules. Furthermore, distinct RNA structures have been identified that govern almost all steps of biological processes in cells, including transcriptional activation and termination, transcriptional mutagenesis, 5'-capping, splicing, 3'-polyadenylation, mRNA export and localization, and translation. Here, we briefly summarize the dynamic and complex features of RNA folding along with a wide variety of intrinsic and extrinsic factors that affect RNA folding. We then provide several examples to elaborate RNA structure-mediated regulation at the transcriptional and posttranscriptional levels. Finally, we illustrate the regulatory roles of RNA structure and discuss advances pertaining to RNA structure in plants. WIREs RNA 2016, 7:562-574. doi: 10.1002/wrna.1350 For further resources related to this article, please visit the WIREs website.

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Division of Labor in an Oligomer of the DEAD-Box RNA Helicase Ded1p

Cited by 62 publications

References 42 publications

Autoinhibitory Interdomain Interactions and Subfamily-specific Extensions Redefine the Catalytic Core of the Human DEAD-box Protein DDX3

Autoinhibitory Interdomain Interactions and Subfamily-specific Extensions Redefine the Catalytic Core of the Human DEAD-box Protein DDX3

Structural Basis of Human Helicase DDX21 in RNA Binding, Unwinding, and Antiviral Signal Activation

Regulatory effects of cotranscriptional RNA structure formation and transitions

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