Double-stranded RNA-activated protein kinase PKR of fishes and amphibians: Varying the number of double-stranded RNA binding domains and lineage-specific duplications

ABSTRACTMaleless (MLE) is an evolutionary conserved member of the DExH family of helicases in Drosophila. Besides its function in RNA editing and presumably siRNA processing, MLE is best known for its role in remodelling non-coding roX RNA in the context of X chromosome dosage compensation in male flies. MLE and its human orthologue, DHX9 contain two tandem double-stranded RNA binding domains (dsRBDs) located at the N-terminal region. The two dsRBDs are essential for localization of MLE at the X-territory and it is presumed that this involves binding roX secondary structures. However, for dsRBD1 roX RNA binding has so far not been described. Here, we determined the solution NMR structure of dsRBD1 and dsRBD2 of MLE in tandem and investigated its role in double-stranded RNA (dsRNA) binding. Our NMR data show that both dsRBDs act as independent structural modules in solution and are canonical, non-sequence-specific dsRBDs featuring non-canonical KKxAK RNA binding motifs. NMR titrations combined with filter binding experiments document the contribution of dsRBD1 to dsRNA binding in vitro. Curiously, dsRBD1 mutants in which dsRNA binding in vitro is strongly compromised do not affect roX2 RNA binding and MLE localization in cells. These data suggest alternative functions for dsRBD1 in vivo.

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Functionally Distinct Double-stranded RNA-binding Domains Associated with Alternative Splice Site Variants of the Interferon-inducible Double-stranded RNA-specific Adenosine Deaminase

Journal of Biological Chemistry ◽

10.1074/jbc.272.7.4419 ◽

1997 ◽

Vol 272 (7) ◽

pp. 4419-4428 ◽

Cited By ~ 95

Author(s):

Yong Liu ◽

Cyril X. George ◽

John B. Patterson ◽

Charles E. Samuel

Keyword(s):

Adenosine Deaminase ◽

Alternative Splice ◽

Splice Site ◽

Rna Binding ◽

Double Stranded Rna ◽

Alternative Splice Site ◽

Binding Domains ◽

Rna Binding Domains

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Structure, dynamics and roX2-lncRNA binding of tandem double-stranded RNA binding domains dsRBD1,2 of Drosophila helicase Maleless

Nucleic Acids Research ◽

10.1093/nar/gkz125 ◽

2019 ◽

Vol 47 (8) ◽

pp. 4319-4333 ◽

Cited By ~ 4

Author(s):

Pravin Kumar Ankush Jagtap ◽

Marisa Müller ◽

Pawel Masiewicz ◽

Sören von Bülow ◽

Nele Merret Hollmann ◽

...

Keyword(s):

Rna Binding ◽

Double Stranded Rna ◽

Structure Dynamics ◽

Binding Domains ◽

Rna Binding Domains

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Deformability in the cleavage site of primary microRNA is not sensed by the double-stranded RNA binding domains in the microprocessor component DGCR8

Proteins Structure Function and Bioinformatics ◽

10.1002/prot.24810 ◽

2015 ◽

Vol 83 (6) ◽

pp. 1165-1179 ◽

Cited By ~ 3

Author(s):

Kaycee A. Quarles ◽

Durga Chadalavada ◽

Scott A. Showalter

Keyword(s):

Cleavage Site ◽

Rna Binding ◽

Double Stranded Rna ◽

Binding Domains ◽

Rna Binding Domains

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Distinct in vivo roles for double-stranded RNA-binding domains of the Xenopus RNA-editing enzyme ADAR1 in chromosomal targeting

The Journal of Cell Biology ◽

10.1083/jcb.200301034 ◽

2003 ◽

Vol 161 (2) ◽

pp. 309-319 ◽

Cited By ~ 18

Author(s):

Michael Doyle ◽

Michael F. Jantsch

Keyword(s):

Rna Editing ◽

Rna Binding ◽

Double Stranded Rna ◽

Binding Domains ◽

Initial Substrate ◽

Rna Binding Domains ◽

Editing Enzyme ◽

Deaminase Domain

The RNA-editing enzyme adenosine deaminase that acts on RNA (ADAR1) deaminates adenosines to inosines in double-stranded RNA substrates. Currently, it is not clear how the enzyme targets and discriminates different substrates in vivo. However, it has been shown that the deaminase domain plays an important role in distinguishing various adenosines within a given substrate RNA in vitro. Previously, we could show that Xenopus ADAR1 is associated with nascent transcripts on transcriptionally active lampbrush chromosomes, indicating that initial substrate binding and possibly editing itself occurs cotranscriptionally. Here, we demonstrate that chromosomal association depends solely on the three double-stranded RNA-binding domains (dsRBDs) found in the central part of ADAR1, but not on the Z-DNA–binding domain in the NH2 terminus nor the catalytic deaminase domain in the COOH terminus of the protein. Most importantly, we show that individual dsRBDs are capable of recognizing different chromosomal sites in an apparently specific manner. Thus, our results not only prove the requirement of dsRBDs for chromosomal targeting, but also show that individual dsRBDs have distinct in vivo localization capabilities that may be important for initial substrate recognition and subsequent editing specificity.

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