scholarly journals PRH75, a new nucleus-localized member of the DEAD-box protein family from higher plants.

1997 ◽  
Vol 17 (4) ◽  
pp. 2257-2265 ◽  
Author(s):  
Z J Lorković ◽  
R G Herrmann ◽  
R Oelmüller
Keyword(s):  
Amino Acids ◽  
Rna Binding ◽  
Protein Family ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Rna Helicases ◽  
Nuclear Targeting ◽  
Dead Box ◽  
Cell Extracts ◽  
The Dead

The putative RNA helicases of the DEAD-box protein family are involved in pre-mRNA splicing, rRNA maturation, ribosome assembly, and translation. Members of this protein family have been identified in organisms from Escherichia coli to humans, but except for the translation initiation factor 4A, there have been no reports on the characterization of other DEAD-box proteins from plants. Here we report on a novel member of the DEAD-box protein family, the plant RNA helicase 75 (PRH75). PRH75 is localized in the nucleus and contains two domains for RNA binding. One is located at the C terminus and is similar to RGG RNA-binding domains of nucleus-localized RNA-binding proteins. The other one is located between amino acids 308 and 622, a region containing the conserved motif VI characteristic of DEAD-box proteins and known as the RNA-binding site of eIF-4A. The N-terminal 81 amino acids are sufficient for nuclear targeting of the protein. Northern and Western blot analyses show that PRH75 is mainly expressed in young and rapidly developing tissues. The purified recombinant PRH75 has a weak ATPase activity which is barely stimulated by RNA ligands. The fractionation of spinach whole-cell extracts by glycerol gradient centrifugation and gel filtration on a Superdex 200 column shows that the protein exists in a complex of about 500 kDa. Possible biological functions of PRH75 as well as structure-function relationships in the context of its modular primary structure are discussed.

The DEAD-box protein family of RNA helicases

Gene ◽  
2006 ◽  
Vol 367 ◽  
pp. 17-37 ◽  
Author(s):  
Olivier Cordin ◽  
Josette Banroques ◽  
N. Kyle Tanner ◽  
Patrick Linder
Keyword(s):  
Protein Family ◽  
Rna Helicases ◽  
Dead Box ◽  
The Dead

The HRIGRXXR region of the DEAD box RNA helicase eukaryotic translation initiation factor 4A is required for RNA binding and ATP hydrolysis

1993 ◽  
Vol 13 (11) ◽  
pp. 6789-6798 ◽  
Author(s):  
A Pause ◽  
N Méthot ◽  
N Sonenberg
Keyword(s):  
Atpase Activity ◽  
Rna Binding ◽  
Atp Hydrolysis ◽  
Rna Helicase ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cross Linking ◽  
Atp Binding ◽  
Eukaryotic Translation ◽  
The Dead

eIF-4A is a eukaryotic translation initiation factor that is required for mRNA binding to ribosomes. It exhibits single-stranded RNA-dependent ATPase activity, and in combination with a second initiation factor, eIF-4B, it exhibits duplex RNA helicase activity. eIF-4A is the prototype of a large family of proteins termed the DEAD box protein family, whose members share nine highly conserved amino acid regions. The functions of several of these conserved regions in eIF-4A have previously been assigned to ATP binding, ATPase, and helicase activities. To define the RNA-binding region of eIF-4A, a UV-induced cross-linking assay was used to analyze binding of mutant eIF-4A proteins to RNA. Mutants carrying mutations in the ATP-binding region (AXXXXGKT), ATPase region (DEAD), helicase region (SAT), and the most carboxy-terminal conserved region of the DEAD family, HRIGRXXR, were tested for RNA cross-linking. We show that mutations, either conservative or not, in any one of the three arginines in the HRIGRXXR sequence drastically reduced eIF-4A cross-linking to RNA. In addition, all the mutations in the HRIGRXXR region abrogate RNA helicase activity. Some but not all of these mutations affect ATP binding and ATPase activity. This is consistent with the hypothesis that the HRIGRXXR region is involved in the ATP hydrolysis reaction and would explain the coupling of ATPase and RNA-binding/helicase activities. Our results show that the HRIGRXXR region, which is QRXGRXXR or QXXGRXXR in the RNA and DNA helicases of the helicase superfamily II, is involved in ATP hydrolysis-dependent RNA interaction during unwinding. We also show that mutations in other regions of eIF-4A that abolish ATPase activity sharply decrease eIF-4A cross-linking to RNA. A model is proposed in which eIF-4A first binds ATP, resulting in a change in eIF-4A conformation which allows RNA binding that is dependent on the HRIGRXXR region. Binding of RNA induces ATP hydrolysis, leading to a more stable interaction with RNA. This process is then linked to unwinding of duplex RNA in the presence of eIF-4B.

scholarly journals DEAD-Box Helicases: Sensors, Regulators, and Effectors for Antiviral Defense

Viruses ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 181 ◽  
Author(s):  
Frances Taschuk ◽  
Sara Cherry
Keyword(s):  
Rna Binding ◽  
Viral Infections ◽  
Rna Binding Proteins ◽  
Large Family ◽  
Cellular Functions ◽  
Rna Helicases ◽  
Independent Manner ◽  
Dead Box ◽  
The Dead ◽  
Immune Sensing

DEAD-box helicases are a large family of conserved RNA-binding proteins that belong to the broader group of cellular DExD/H helicases. Members of the DEAD-box helicase family have roles throughout cellular RNA metabolism from biogenesis to decay. Moreover, there is emerging evidence that cellular RNA helicases, including DEAD-box helicases, play roles in the recognition of foreign nucleic acids and the modulation of viral infection. As intracellular parasites, viruses must evade detection by innate immune sensing mechanisms and degradation by cellular machinery while also manipulating host cell processes to facilitate replication. The ability of DEAD-box helicases to recognize RNA in a sequence-independent manner, as well as the breadth of cellular functions carried out by members of this family, lead them to influence innate recognition and viral infections in multiple ways. Indeed, DEAD-box helicases have been shown to contribute to intracellular immune sensing, act as antiviral effectors, and even to be coopted by viruses to promote their replication. However, our understanding of the mechanisms underlying these interactions, as well as the cellular roles of DEAD-box helicases themselves, is limited in many cases. We will discuss the diverse roles that members of the DEAD-box helicase family play during viral infections.

scholarly journals The DEAD-box helicase Ded1 from yeast is an mRNP cap-associated protein that shuttles between the cytoplasm and nucleus

2014 ◽  
Vol 42 (15) ◽  
pp. 10005-10022 ◽  
Author(s):  
Meriem Senissar ◽  
Agnès Le Saux ◽  
Naïma Belgareh-Touzé ◽  
Céline Adam ◽  
Josette Banroques ◽  
...  
Keyword(s):  
Cell Cycle Regulation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Start Codon ◽  
Yeast Protein ◽  
Enzymatic Assays ◽  
Dead Box ◽  
The Dead ◽  
Cycle Regulation

Abstract The DEAD-box helicase Ded1 is an essential yeast protein that is closely related to mammalian DDX3 and to other DEAD-box proteins involved in developmental and cell cycle regulation. Ded1 is considered to be a translation-initiation factor that helps the 40S ribosome scan the mRNA from the 5′ 7-methylguanosine cap to the AUG start codon. We used IgG pull-down experiments, mass spectrometry analyses, genetic experiments, sucrose gradients, in situ localizations and enzymatic assays to show that Ded1 is a cap-associated protein that actively shuttles between the cytoplasm and the nucleus. NanoLC-MS/MS analyses of purified complexes show that Ded1 is present in both nuclear and cytoplasmic mRNPs. Ded1 physically interacts with purified components of the nuclear CBC and the cytoplasmic eIF4F complexes, and its enzymatic activity is stimulated by these factors. In addition, we show that Ded1 is genetically linked to these factors. Ded1 comigrates with these proteins on sucrose gradients, but treatment with rapamycin does not appreciably alter the distribution of Ded1; thus, most of the Ded1 is in stable mRNP complexes. We conclude that Ded1 is an mRNP cofactor of the cap complex that may function to remodel the different mRNPs and thereby regulate the expression of the mRNAs.

scholarly journals PKR activity modulation by phosphomimetic mutations of serine residues located three aminoacids upstream of double-stranded RNA binding motifs

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Teresa Cesaro ◽  
Yohei Hayashi ◽  
Fabian Borghese ◽  
Didier Vertommen ◽  
Fanny Wavreil ◽  
...  
Keyword(s):  
Amino Acids ◽  
Rna Binding ◽  
Viral Infections ◽  
Phosphorylation Site ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Poly I:C ◽  
Binding Motif ◽  
Double Stranded Rna

AbstractEukaryotic translation initiation factor 2 alpha kinase 2 (EIF2AK2), better known as PKR, plays a key role in the response to viral infections and cellular homeostasis by regulating mRNA translation. Upon binding dsRNA, PKR is activated through homodimerization and subsequent autophosphorylation on residues Thr446 and Thr451. In this study, we identified a novel PKR phosphorylation site, Ser6, located 3 amino acids upstream of the first double-stranded RNA binding motif (DRBM1). Another Ser residue occurs in PKR at position 97, the very same position relative to the DRBM2. Ser or Thr residues also occur 3 amino acids upstream DRBMs of other proteins such as ADAR1 or DICER. Phosphoinhibiting mutations (Ser-to-Ala) introduced at Ser6 and Ser97 spontaneously activated PKR. In contrast, phosphomimetic mutations (Ser-to-Asp) inhibited PKR activation following either poly (I:C) transfection or virus infection. These mutations moderately affected dsRNA binding or dimerization, suggesting a model where negative charges occurring at position 6 and 97 tighten the interaction of DRBMs with the kinase domain, thus keeping PKR in an inactive closed conformation even in the presence of dsRNA. This study provides new insights on PKR regulation mechanisms and identifies Ser6 and Ser97 as potential targets to modulate PKR activity for therapeutic purposes.

Selective targeting of the DEAD-box RNA helicase eukaryotic initiation factor (eIF) 4A by natural products

2020 ◽  
Vol 37 (5) ◽  
pp. 609-616 ◽  
Author(s):  
Leo Shen ◽  
Jerry Pelletier
Keyword(s):  
Natural Products ◽  
Rna Binding ◽  
Rna Helicase ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Dead Box ◽  
Selective Targeting ◽  
Resident Time ◽  
The Dead

This highlight reviews natural products targeting of the eIF4A RNA helicase by interfering with RNA-binding or acting as interfacial inhibitors to increase RNA resident time.

scholarly journals The HRIGRXXR region of the DEAD box RNA helicase eukaryotic translation initiation factor 4A is required for RNA binding and ATP hydrolysis.

1993 ◽  
Vol 13 (11) ◽  
pp. 6789-6798 ◽  
Author(s):  
A Pause ◽  
N Méthot ◽  
N Sonenberg
Keyword(s):  
Atpase Activity ◽  
Rna Binding ◽  
Atp Hydrolysis ◽  
Rna Helicase ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cross Linking ◽  
Atp Binding ◽  
Eukaryotic Translation ◽  
The Dead

eIF-4A is a eukaryotic translation initiation factor that is required for mRNA binding to ribosomes. It exhibits single-stranded RNA-dependent ATPase activity, and in combination with a second initiation factor, eIF-4B, it exhibits duplex RNA helicase activity. eIF-4A is the prototype of a large family of proteins termed the DEAD box protein family, whose members share nine highly conserved amino acid regions. The functions of several of these conserved regions in eIF-4A have previously been assigned to ATP binding, ATPase, and helicase activities. To define the RNA-binding region of eIF-4A, a UV-induced cross-linking assay was used to analyze binding of mutant eIF-4A proteins to RNA. Mutants carrying mutations in the ATP-binding region (AXXXXGKT), ATPase region (DEAD), helicase region (SAT), and the most carboxy-terminal conserved region of the DEAD family, HRIGRXXR, were tested for RNA cross-linking. We show that mutations, either conservative or not, in any one of the three arginines in the HRIGRXXR sequence drastically reduced eIF-4A cross-linking to RNA. In addition, all the mutations in the HRIGRXXR region abrogate RNA helicase activity. Some but not all of these mutations affect ATP binding and ATPase activity. This is consistent with the hypothesis that the HRIGRXXR region is involved in the ATP hydrolysis reaction and would explain the coupling of ATPase and RNA-binding/helicase activities. Our results show that the HRIGRXXR region, which is QRXGRXXR or QXXGRXXR in the RNA and DNA helicases of the helicase superfamily II, is involved in ATP hydrolysis-dependent RNA interaction during unwinding. We also show that mutations in other regions of eIF-4A that abolish ATPase activity sharply decrease eIF-4A cross-linking to RNA. A model is proposed in which eIF-4A first binds ATP, resulting in a change in eIF-4A conformation which allows RNA binding that is dependent on the HRIGRXXR region. Binding of RNA induces ATP hydrolysis, leading to a more stable interaction with RNA. This process is then linked to unwinding of duplex RNA in the presence of eIF-4B.

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