Intrinsic Double-Stranded-RNA Processing Activity ofEscherichia coliRibonuclease III Lacking the dsRNA-Binding Domain†

Biochemistry ◽  
2001 ◽  
Vol 40 (49) ◽  
pp. 14976-14984 ◽  
Author(s):  
Weimei Sun ◽  
Eunjoo Jun ◽  
Allen W. Nicholson
Keyword(s):  
Rna Processing ◽  
Binding Domain ◽  
Double Stranded Rna ◽  
Dsrna Binding ◽  
Processing Activity

scholarly journals ADAR1 Interacts with NF90 through Double-Stranded RNA and Regulates NF90-Mediated Gene Expression Independently of RNA Editing

2005 ◽  
Vol 25 (16) ◽  
pp. 6956-6963 ◽  
Author(s):  
Yongzhan Nie ◽  
Li Ding ◽  
Peter N. Kao ◽  
Robert Braun ◽  
Jing-Hua Yang
Keyword(s):  
Gene Expression ◽  
Rna Editing ◽  
Nuclear Export ◽  
Fetal Liver ◽  
Regulation Of Gene Expression ◽  
Nuclear Export Signal ◽  
Binding Domain ◽  
Double Stranded Rna ◽  
Recognition Element ◽  
Dsrna Binding

ABSTRACT The RNA-editing enzyme ADAR1 modifies adenosines by deamination and produces A-to-I mutations in mRNA. ADAR1 was recently demonstrated to function in host defense and in embryonic erythropoiesis during fetal liver development. The mechanisms for these phenotypic effects are not yet known. Here we report a novel function of ADAR1 in the regulation of gene expression by interacting with the nuclear factor 90 (NF90) proteins, known regulators that bind the antigen response recognition element (ARRE-2) and have been demonstrated to stimulate transcription and translation. ADAR1 upregulates NF90-mediated gene expression by interacting with the NF90 proteins, including NF110, NF90, and NF45. A knockdown of NF90 with small interfering RNA suppresses this function of ADAR1. Coimmunoprecipitation and double-stranded RNA (dsRNA) digestion demonstrate that ADAR1 is associated with NF110, NF90, and NF45 through the bridge of cellular dsRNA. Studies with ADAR1 deletions demonstrate that the dsRNA binding domain and a region covering the Z-DNA binding domain and the nuclear export signal comprise the complete function of ADAR1 in upregulating NF90-mediated gene expression. These data suggest that ADAR1 has the potential both to change information content through editing of mRNA and to regulate gene expression through interacting with the NF90 family proteins.

scholarly journals Characterization of RNA Determinants Recognized by the Arginine- and Proline-Rich Region of Us11, a Herpes Simplex Virus Type 1-Encoded Double-Stranded RNA Binding Protein That Prevents PKR Activation

2002 ◽  
Vol 76 (23) ◽  
pp. 11971-11981 ◽  
Author(s):  
David Khoo ◽  
Cesar Perez ◽  
Ian Mohr
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Rna Binding ◽  
Binding Protein ◽  
Binding Domain ◽  
Double Stranded Rna ◽  
Dsrna Binding ◽  
Rna Ligands ◽  
Pkr Kinase ◽  
Simplex Virus

ABSTRACT The herpes simplex virus Us11 gene product inhibits activation of the cellular PKR kinase and associates with a limited number of unrelated viral and cellular RNA molecules via a carboxyl-terminal 68-amino-acid segment rich in arginine and proline. To characterize the determinants underlying the recognition of an RNA target by Us11, we employed an in vitro selection technique to isolate RNA ligands that bind Us11 with high affinity from a population of molecules containing an internal randomized segment. Binding of Us11 to these RNA ligands is specific and appears to occur preferentially on conformational isoforms that possess a higher-order structure. While the addition of unlabeled poly(I · C) reduced binding of Us11 to a selected radiolabeled RNA, single-stranded homopolymers were not effective competitors. Us11 directly associates with poly(I · C), and inclusion of an unlabeled selected RNA in the reaction reduces poly(I · C) binding, while single-stranded RNA homopolymers have no effect. Finally, Us11 binds to defined, double-stranded RNA (dsRNA) molecules that exhibit greater sequence complexity. Binding to these dsRNA perfect duplexes displays a striking dependence on length, as 39-bp or shorter duplexes do not bind efficiently. Furthermore, this interaction is specific for dsRNA as opposed to dsDNA, implying that the Us11 RNA binding domain can distinguish nucleic acid duplexes containing 2′ hydroxyl groups from those that do not. These results establish that Us11 is a dsRNA binding protein. The arginine- and proline-rich Us11 RNA binding domain is unrelated to known dsRNA binding elements and thus constitutes a unique recognition motif that interacts with dsRNA. The ability of Us11 to bind dsRNA may be important for inhibiting activation of the cellular PKR kinase in response to dsRNA.

Mechanism of Interaction of the Double-Stranded RNA (dsRNA) Binding Domain of Protein Kinase R with Short dsRNA Sequences†

Biochemistry ◽  
2007 ◽  
Vol 46 (1) ◽  
pp. 55-65 ◽  
Author(s):  
Jason W. Ucci ◽  
Yumiko Kobayashi ◽  
Gregory Choi ◽  
Andrei T. Alexandrescu ◽  
James L. Cole
Keyword(s):  
Protein Kinase ◽  
Binding Domain ◽  
Protein Kinase R ◽  
Double Stranded Rna ◽  
Dsrna Binding ◽  
Mechanism Of Interaction

scholarly journals Mutational Analysis of the Double-stranded RNA (dsRNA) Binding Domain of the dsRNA-activated Protein Kinase, PKR

1995 ◽  
Vol 270 (6) ◽  
pp. 2601-2606 ◽  
Author(s):  
Nigel A. J. McMillan ◽  
Bruce W. Carpick ◽  
Britton Hollis ◽  
W. Mark Toone ◽  
Maryam Zamanian-Daryoush ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Mutational Analysis ◽  
Binding Domain ◽  
Double Stranded Rna ◽  
Dsrna Binding

scholarly journals A Novel Murine Staufen Isoform Modulates the RNA Content of Staufen Complexes

2000 ◽  
Vol 20 (15) ◽  
pp. 5592-5601 ◽  
Author(s):  
Thomas Duchaîne ◽  
Hui-Jun Wang ◽  
Ming Luo ◽  
Sergey V. Steinberg ◽  
Ivan R. Nabi ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Rna Binding ◽  
Binding Domain ◽  
Binding Ability ◽  
Double Stranded Rna ◽  
Transfected Cells ◽  
Rna Content ◽  
Dsrna Binding ◽  
In Vivo Analysis

ABSTRACT Mouse Staufen (mStau) is a double-stranded RNA-binding protein associated with polysomes and the rough endoplasmic reticulum (RER). We describe a novel endogenous isoform of mStau (termed mStaui) which has an insertion of six amino acids within dsRBD3, the major double-stranded RNA (dsRNA)-binding domain. With a structural change of the RNA-binding domain, this conserved and widely distributed isoform showed strongly impaired dsRNA-binding ability. In transfected cells, mStaui exhibited the same tubulovesicular distribution (RER) as mStau when weakly expressed; however, when overexpressed, mStaui was found in large cytoplasmic granules. Markers of the RER colocalized with mStaui-containing granules, showing that overexpressed mStaui could still be associated with the RER. Cotransfection of mStaui with mStau relocalized overexpressed mStaui to the reticular RER, suggesting that they can form a complex on the RER and that a balance between these isoforms is important to achieve proper localization. Coimmunoprecipitation demonstrated that the two mStau isoforms are components of the same complex in vivo. Analysis of the immunoprecipitates showed that mStau is a component of an RNA-protein complex and that the association with mStaui drastically reduces the RNA content of the complex. We propose that this new isoform, by forming a multiple-isoform complex, regulates the amount of RNA in mStau complexes in mammalian cells.

scholarly journals Double-Stranded RNA Binding by Human Cytomegalovirus pTRS1

2005 ◽  
Vol 79 (12) ◽  
pp. 7311-7318 ◽  
Author(s):  
Morgan Hakki ◽  
Adam P. Geballe
Keyword(s):  
Human Cytomegalovirus ◽  
Mammalian Cells ◽  
Rna Binding ◽  
Binding Domain ◽  
Deletion Mutants ◽  
Double Stranded Rna ◽  
Antiviral Responses ◽  
Dsrna Binding ◽  
Free Translation ◽  
Carboxy Terminal

ABSTRACT The human cytomegalovirus (HCMV) TRS1 and IRS1 genes rescue replication of vaccinia virus (VV) that has a deletion of the double-stranded RNA binding protein gene E3L (VVΔE3L). Like E3L, these HCMV genes block the activation of key interferon-induced, double-stranded RNA (dsRNA)-activated antiviral pathways. We investigated the hypothesis that the products of these HCMV genes act by binding to dsRNA. pTRS1 expressed by cell-free translation or by infection of mammalian cells with HCMV or recombinant VV bound to dsRNA. Competition experiments revealed that pTRS1 preferentially bound to dsRNA compared to double-stranded DNA or single-stranded RNA. 5′- and 3′-end deletion analyses mapped the TRS1 dsRNA-binding domain to amino acids 74 through 248, a region of identity to pIRS1 that contains no homology to known dsRNA-binding proteins. Deletion of the majority of this region (Δ86-246) completely abrogated dsRNA binding. To determine the role of the dsRNA-binding domain in the rescue of VVΔE3L replication, wild-type or deletion mutants of TRS1 were transfected into HeLa cells, which were then infected with VVΔE3L. While full-length TRS1 rescued VVΔE3L replication, deletion mutants affecting a carboxy-terminal region of TRS1 that is not required for dsRNA binding failed to rescue VVΔE3L. Analyses of stable cell lines revealed that the carboxy-terminal domain is necessary to prevent the shutoff of protein synthesis and the phosphorylation of eIF2α after VVΔE3L infection. Thus, pTRS1 contains an unconventional dsRNA-binding domain at its amino terminus, but a second function involving the carboxy terminus is also required for countering host cell antiviral responses.

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