Dbp5, a DEAD-box protein required for mRNA export, is recruited to the cytoplasmic fibrils of nuclear pore complex via a conserved interaction with CAN/Nup159p

Schmitt, Christel; Kobbe, Cayetano von; Bachi, Ángela; Panté, Nelly; Rodrigues, João; Boscheron, Cécile; Rigaut, Guillaume; Wilm, Matthias; Séraphin, Bertrand; Carmo‐Fonseca, Maria; Izaurralde, Elisa

doi:10.1093/emboj/18.15.4332

Cited by 257 publications

(290 citation statements)

References 69 publications

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“…With this single-base pair addition, the GRTH cDNA contains 357 additional base pair and an ATG codon (ϩ1) 5Ј in-frame to the original cDNA ATG codon (earlier defined as ϩ1 bp, now revised as ϩ343 bp). This sequence of 1630 bp contains an open reading frame encoding 483 amino acids, with the additional 114 amino acids resulting from 5Ј sequences of our earlier reported clone ( N-terminal sequences are conserved between GRTH of rat, mouse, and human and DBP5 of mouse and human (8,9). Furthermore, our recent studies demonstrated that the TATA-less GRTH gene contains multiple transcriptional start sites located within 80 bp 5Ј to the first ATG codon (ϩ1).…”

Section: Resultsmentioning

confidence: 95%

Cell-specific and Hormone-regulated Expression of Gonadotropin-regulated Testicular RNA Helicase Gene (GRTH/Ddx25) Resulting from Alternative Utilization of Translation Initiation Codons in the Rat Testis

Tsai‐Morris

Dufau

2003

Journal of Biological Chemistry

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Gonadotropin-regulated testicular RNA helicase (GRTH) is a novel DEAD-box protein withRNA helicases are ribonucleic acid-binding proteins that regulate RNA structure, possess ATPase activity, and unwind double-stranded RNA in an ATP-dependent manner (1, 2). A large number of RNA helicases have been identified, and these were classified into superfamilies based on specific amino acid sequences in the conserved motifs common to all family members. Among these proteins, DEAD-box proteins were grouped in a family of RNA helicases; that is, the DEAD-box family (Asp-Glu-Ala-Asp), whose members exert regulatory roles in various aspects of RNA metabolism including translation, nuclear transcription, pre-mRNA splicing, mRNA export, and ribosome biogenesis (1, 2). In addition, some members of the DEAD-box family participate in regulatory events during organ maturation and cellular differentiation (1, 2).The gonadotropin-regulated testicular RNA helicase (GRTH/ Ddx25), 1 cloned from the rat Leydig cell, mouse and human testis cDNA libraries, is a novel member of the DEAD-box protein family of RNA helicases and is the first member found to be regulated by a hormone (3). Purified recombinant GST-GRTH displayed ATPase and ATP-dependent bi-directional RNA helicase activities. It also increased in vitro translation of luciferase RNA templates (3). Northern analysis indicated that this helicase is highly expressed in rat, mouse, and human testes and is weakly expressed in the pituitary and hypothalamus. In vivo/in vitro studies demonstrated that GRTH is transcriptionally up-regulated by human chorionic gonadotropin at doses that cause down-regulation of luteinizing hormone/human chorionic gonadotropin (hCG) receptors, steroidogenic enzymes, and androgen formation (4). Furthermore, in vitro studies revealed that induction of GRTH mRNA by hCG is mediated via second messenger and androgen in Leydig cells. Inhibition of androgen production by inhibitors of steroidogenic enzymes or blockade of androgen action by a receptor antagonist abrogated the stimulation of GRTH mRNA by gonadotropin or cAMP. In situ hybridization analysis demonstrated that GRTH is predominantly expressed in the testis in both somatic Leydig cells and meiotic spermatocytes and haploid germinal cells of the seminiferous epithelium and is developmentally regulated. In the present study, we identified GRTH protein species, evaluated their protein levels, and studied their cellular distribution within the testicular compartments and their hormonal regulation in the adult rats testis. The regulation of this enzyme by gonadotropin and androgen and its stage-specific localization in germ cells indicate that GRTH could participate in the regulation of androgen-dependent steroidogenesis and spermatogenesis.

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Section: Resultsmentioning

confidence: 95%

Cell-specific and Hormone-regulated Expression of Gonadotropin-regulated Testicular RNA Helicase Gene (GRTH/Ddx25) Resulting from Alternative Utilization of Translation Initiation Codons in the Rat Testis

Tsai‐Morris

Dufau

2003

Journal of Biological Chemistry

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“…Also, 60 S export was rescued by Nup214 versions that cannot bind Dbp5. Dbp5 interaction with Nup214-Nup159 is required for mRNA export (63,64), suggesting that the observed 60 S export defects are not primarily caused by general inhibition of mRNA export.…”

Section: Discussionmentioning

confidence: 98%

Nup214-Nup88 Nucleoporin Subcomplex Is Required for CRM1-mediated 60 S Preribosomal Nuclear Export

Bernad¹,

Engelsma

Sanderson³

et al. 2006

Journal of Biological Chemistry

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The nuclear pore complex (NPC) conducts macromolecular transport to and from the nucleus and provides a kinetic/hydrophobic barrier composed of phenylalanine-glycine (FG) repeats. Nuclear transport is achieved through permeation of this barrier by transport receptors. The transport receptor CRM1 facilitates export of a large variety of cargoes. Export of the preribosomal 60 S subunit follows this pathway through the adaptor protein NMD3. Using RNA interference, we depleted two FG-containing cytoplasmically oriented NPC complexes, Nup214-Nup88 and Nup358, and investigated CRM1-mediated export. A dramatic defect in NMD3-mediated export of preribosomes was found in Nup214-Nup88-depleted cells, whereas only minor export defects were evident in other CRM1 cargoes or upon depletion of Nup358. We show that the large C-terminal FG domain of Nup214 is not accessible to freely diffusing molecules from the nucleus, indicating that it does not conduct 60 S preribosomes through the NPC. Consistently, derivatives of Nup214 lacking the FG-repeat domain rescued the 60 S export defect. We show that the coiled-coil region of Nup214 is sufficient for 60 S nuclear export, coinciding with recruitment of Nup88 to the NPC. Our data indicate that Nup214 plays independent roles in NPC function by participating in the kinetic/ hydrophobic barrier through its FG-rich domain and by enabling NPC gating through association with Nup88.

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“…Dbp5 predominantly localizes to the cytoplasm and is particularly concentrated at the rim of nuclear pores, where it associates with cytoplasmic fibrils of the nuclear pore complex 116 . This localization requires the nucleo porin protein Nup159 (NUP214 in mammal s) 117,118 .…”

Section: Nuclear Specklesmentioning

confidence: 99%

From unwinding to clamping — the DEAD box RNA helicase family

Linder

Jankowsky

2011

Nat Rev Mol Cell Biol

983

1,069

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Nearly all aspects of RNA metabolism, from transcription and translation to mRNA decay, involve RNA helicases, which are enzymes that use ATP to bind or remodel RNA and RNA-protein complexes (ribonucleoprotein (RNP) complexes) 1 . RNA helicases are found in all three domains of life, and many viruses also encode one or more of these proteins 2,3 . Together with the structurally related DNA helicases that function in replication, recombination and repair, the RNA helicases are classified into superfamilies and families, based on sequence and structural features 3,4 . DEAD box proteins form the largest helicase family, with 37 members in humans and 26 in Saccharomyces cerevisiae 3 , and are characterized by the presence of an Asp-Glu-Ala-Asp (DEAD) motif.DEAD box helicases have central and, in many cases, essential physiological roles in cellular RNA metabolism 5 (FIG. 1). The proteins generally function as part of larger multicomponent assemblies, such as the spliceosom e or the eukaryotic translation initiation machinery 6 . Mutations and deregulation of several DEAD box proteins have been linked to disease states, including cancer 7 . Although all DEAD box proteins contain a structurally highly conserved core with conserved ATP-binding and RNA-binding sites, different proteins have been associated with diverse and seemingly unrelated functions, including the disassembly of RNPs, chaperoning during RNA folding and even stabil ization of protein complexes on RNA 5,6 . How these highly conserved proteins fulfil such an array of different functions has been a long-standing question.Research has now started to illuminate the molecular basis for this functional diversity. In this Review, we summarize how structural data, together with biochemical and biophysical studies, have revealed unexpected modes by which these proteins function. We also discuss how novel molecular and cell biological approaches have better defined the cellular roles of DEAD box helicases. We outline our current view on common structural and mechan istic themes that have emerged, and on physical models of how these 'unusual' RNA helicases work.Structures of DEAD box RNA helicases A number of structural studies have universally shown that members of the DEAD box family contain a highly conserved helicase core that harbours the binding sites for ATP and RNA 3,4,8 . The core is surrounded by variable auxiliary domains, which are thought to be critical for the diverse functions of these enzymes.Structure of the helicase core. DEAD box proteins belong to helicase superfamily 2 (SF2) 2 . Similarly to all SF2 helicases, DEAD box proteins are built around a highly conserved helicase core of two virtually identical domains that resemble the bacterial recombination protein recombinase A (RecA) 3,4,8 (FIG. 2a,b). Within this helicase core, at least 12 characteristic sequence motifs are located at conserved positions (FIG. 2a,b). Some of these motifs are conserved across the entire SF2 family, whereas others are found only in the DEAD box family 3 ....

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Dbp5, a DEAD-box protein required for mRNA export, is recruited to the cytoplasmic fibrils of nuclear pore complex via a conserved interaction with CAN/Nup159p

Cited by 257 publications

References 69 publications

Cell-specific and Hormone-regulated Expression of Gonadotropin-regulated Testicular RNA Helicase Gene (GRTH/Ddx25) Resulting from Alternative Utilization of Translation Initiation Codons in the Rat Testis

Cell-specific and Hormone-regulated Expression of Gonadotropin-regulated Testicular RNA Helicase Gene (GRTH/Ddx25) Resulting from Alternative Utilization of Translation Initiation Codons in the Rat Testis

Nup214-Nup88 Nucleoporin Subcomplex Is Required for CRM1-mediated 60 S Preribosomal Nuclear Export

From unwinding to clamping — the DEAD box RNA helicase family

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