The DEAD Box Protein eIF4A. 2. A Cycle of Nucleotide and RNA-Dependent Conformational Changes

Lorsch, Jon R.; Herschlag, Daniel

doi:10.1021/bi9724319

Cited by 146 publications

(107 citation statements)

References 44 publications

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“…S5). As proposed for the dsRNA melting mechanism of Vasa and other biochemical studies on eIF4A, we expect that eIF4A cooperatively binds to ATP and dsRNA in concert with the closure of the two domains and subsequently induces the unwinding of the bound dsRNA (16,17) (Fig. 4A).…”

Section: Implication Of the Two Different Binding Modes In The Functimentioning

confidence: 53%

Crystal structure of the eIF4A–PDCD4 complex

Chang

Cho

Sohn

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

104

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Tumor suppressor programmed cell death protein 4 (PDCD4) inhibits the translation initiation factor eIF4A, an RNA helicase that catalyzes the unwinding of secondary structure at the 5 -untranslated region of mRNAs and controls the initiation of translation. Here, we determined the crystal structure of the human eIF4A and PDCD4 complex. The structure reveals that one molecule of PDCD4 binds to the two eIF4A molecules through the two different binding modes. While the two MA3 domains of PDCD4 bind to one eIF4A molecule, the C-terminal MA3 domain alone of the same PDCD4 also interacts with another eIF4A molecule. The eIF4A-PDCD4 complex structure suggests that the MA3 domain(s) of PDCD4 binds perpendicular to the interface of the two domains of eIF4A, preventing the domain closure of eIF4A and blocking the binding of RNA to eIF4A, both of which are required events in the function of eIF4A helicase. The structure, together with biochemical analyses, reveals insights into the inhibition mechanism of eIF4A by PDCD4 and provides a framework for designing chemicals that target eIF4A.translation inhibition ͉ tumor suppressor ͉ RNA helicase ͉ domain closure ͉ MA3 domain P rogrammed cell death protein 4 (PDCD4) is a translation inhibitor that suppresses neoplastic transformation in cultured cells and transgenic mice (1-3). Loss or reduced expression of PDCD4 has been implicated in the development and progression of a variety of aggressive human cancers (4-6). PDCD4 is regulated by the S6K1 kinase and the SCF ␤TRCP ubiquitin ligase in response to the activation of the mTOR pathway by mitogens, and the controlled degradation of PDCD4 is essential for efficient protein synthesis and consequently for cell growth and proliferation (7).PDCD4 is believed to perform its tumor suppressor function primarily through interaction with eIF4A and eIF4G, which are components of mRNA-binding complex eIF4F (8). eIF4A is a DEAD-box RNA helicase, with two domains, that unwinds the secondary structures in 5Ј-untranslated region (UTR) and cap of mRNA and thereby facilitates ribosome scanning (8). eIF4G is an adaptor protein that coordinates assembly of translation factors and the small ribosomal subunit (8). PDCD4 is believed to inhibit cap-dependent translation by directly inhibiting the helicase activity of eIF4A or by competing with eIF4G for binding to eIF4A and preventing assembly into a eukaryotic initiation complex, eIF4F, or both (1, 9, 10).PDCD4 is formed with the two MA3 domains at its middle (mMA3) and C-terminal regions (cMA3) (9, 11-13). Mutational and NMR binding analysis have shown that PDCD4 uses both MA3 domains to interact with eIF4A and prevents translation (9, 10). However, other studies have demonstrated that the cMA3 domain alone is sufficient for the inhibition of RNA helicase and translation (12). The interactions between eIF4A and PDCD4 have been analyzed in several mutational and NMR mapping studies (1, 9, 10). Nevertheless, it is unclear from these studies how PDCD4 inhibits eIF4A at the molecular level. To elucidate ...

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Section: Implication Of the Two Different Binding Modes In The Functimentioning

confidence: 53%

Crystal structure of the eIF4A–PDCD4 complex

Chang

Cho

Sohn

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

104

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“…In addition, the lack of contributions of the g-phosphate to nucleotide affinity is an indication for ATP-driven conformational changes. Limited proteolysis experiments provide further evidence for ATP-induced conformational changes (Lorsch and Herschlag, 1998b;Henn et al, 2002;Cheng et al, 2005;Low et al, 2007).…”

Section: Nucleotide Bindingmentioning

confidence: 88%

“…Indirect evidence for such a conformational reorganization is available from different proteolysis patterns in the presence or absence of substrates (Lorsch and Herschlag, 1998b;Henn et al, 2002;Cheng et al, 2005;Low et al, 2007). Nucleotide binding kinetics support nucleotide-driven conformational rearrangements: ATP and ADP binding follows a single exponential in the absence of RNA, but a second slow phase appears when RNA is present.…”

Section: Cooperativity Of Rna and Nucleotide Bindingmentioning

confidence: 99%

The mechanism of ATP-dependent RNA unwinding by DEAD box proteins

Hilbert

Karow²,

Klostermeier³

2009

BCHM

143

171

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DEAD box proteins catalyze the ATP-dependent unwinding of double-stranded RNA (dsRNA). In addition, they facilitate protein displacement and remodeling of RNA or RNA/protein complexes. Their hallmark feature is local destabilization of RNA duplexes. Here, we summarize current data on the DEAD box protein mechanism and present a model for RNA unwinding that integrates recent data on the effect of ATP analogs and mutations on DEAD box protein activity. DEAD box proteins share a conserved helicase core with two flexibly linked RecAlike domains that contain all helicase signature motifs. Variable flanking regions contribute to substrate binding and modulate activity. In the presence of ATP and RNA, the helicase core adopts a compact, closed conformation with extensive interdomain contacts and high affinity for RNA. In the closed conformation, the RecA-like domains form a catalytic site for ATP hydrolysis and a continuous RNA binding site. A kink in the backbone of the bound RNA locally destabilizes the duplex. Rearrangement of this initial complex generates a hydrolysisand unwinding-competent state. From this complex, the first RNA strand can dissociate. After ATP hydrolysis and phosphate release, the DEAD box protein returns to a low-affinity state for RNA. Dissociation of the second RNA strand and reopening of the cleft in the helicase core allow for further catalytic cycles.

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“…The helicase activity of eIF4F (via eIF4A c ) is thought to unwind local secondary structure in the 5Ј UTR of mRNAs to facilitate cap-dependent ribosome recruitment (6-8). In the crystal form, eIF4A f has a distended ''dumbbell'' structure consisting of two domains (11-13), which undergo conformational changes in response to RNA and ATP binding (14).There are three eIF4A family members: eIF4AI, eIF4AII, and eIF4AIII. eIF4AI and eIF4AII show 90-95% similarity at the amino acid level, are involved in translation, and appear to have the same biological activity in vitro (15,16).…”

mentioning

confidence: 99%

Stimulation of mammalian translation initiation factor eIF4A activity by a small molecule inhibitor of eukaryotic translation

Bordeleau

Matthews

Wojnar

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

213

245

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RNA helicases are the largest group of enzymes in eukaryotic RNA metabolism. The DEXD͞H-box putative RNA helicases form the helicase superfamily II, whose members are defined by seven highly conserved amino acid motifs, making specific targeting of selected members a challenging pharmacological problem. The translation initiation factor eIF4A is the prototypical DEAD-box RNA helicase that works in conjunction with eIF4B and eIF4H and as a subunit of eIF4F to prepare the mRNA template for ribosome binding, possibly by unwinding the secondary structure proximal to the 5 m 7 GpppN cap structure. We report the identification and characterization of a small molecule inhibitor of eukaryotic translation initiation that acts in an unusual manner by stimulating eIF4A-associated activities. Our results suggest that proper control of eIF4A helicase activity is necessary for efficient ribosome binding and demonstrate the feasibility of selectively targeting DEADbox RNA helicases with small molecules.chemical biology ͉ DEAD-box helicase ͉ pateamine T he ribosome recruitment step of translation initiation is ratelimiting and an important regulatory point whereby cellular environmental cues (e.g., amino acid starvation, mitogenic stimulation, and hypoxia) are linked to the process of translation (1). Two distinct pathways exist for recruitment of the ribosome to the mRNA template. One mechanism is cap-dependent and is facilitated by the presence of the 5Ј cap structure (m 7 GpppN, where N is any nucleotide) on the mRNA. It is catalyzed by the eIF4 class of translation initiation factors and involves the recruitment of ribosomes near the 5Ј end of the mRNA template (1). The second mode involves ribosome recruitment in a cap-independent fashion to an internal ribosome entry site (IRES). Initiation factor requirement for internal ribosome binding varies among IRESes, with some not requiring any factors (2).Preparation of the mRNA template for cap-dependent ribosome recruitment is achieved by eIF4F, eIF4A, eIF4B, eIF4H, and ATP hydrolysis (1). The eIF4F complex is comprised of three subunits: (i) eIF4E, which binds the mRNA cap structure in an ATP-independent fashion; (ii) eIF4A, an RNA helicase that exhibits RNA-dependent ATPase activity and ATPstimulated RNA binding activity (3); and (iii) eIF4G, a modular scaffold that mediates mRNA binding of the 43S preinitiation complex through interactions with eIF3. eIF4B, and eIF4H cooperate with eIF4A to make its helicase activity more processive (4, 5). eIF4A exists as a free form (referred to herein as eIF4A f ) and as a subunit of eIF4F (eIF4A c ) and is thought to cycle through the eIF4F complex during initiation (6-8). When localized in the eIF4F complex, eIF4A c is Ϸ20-fold more efficient as an RNA helicase than when found alone (4, 9), leading to the proposal that eIF4A c is the functional helicase for translation initiation (10). The helicase activity of eIF4F (via eIF4A c ) is thought to unwind local secondary structure in the 5Ј UTR of mRNAs to facilitate cap-dependent ribosome...

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The DEAD Box Protein eIF4A. 2. A Cycle of Nucleotide and RNA-Dependent Conformational Changes

Cited by 146 publications

References 44 publications

Crystal structure of the eIF4A–PDCD4 complex

Crystal structure of the eIF4A–PDCD4 complex

The mechanism of ATP-dependent RNA unwinding by DEAD box proteins

Stimulation of mammalian translation initiation factor eIF4A activity by a small molecule inhibitor of eukaryotic translation

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