Activation of the protein kinase PKR by short double-stranded RNAs with single-stranded tails.
The human RNA-activated protein kinase PKR is an interferon-induced protein that is part of the innate immune response and inhibits viral replication. The action of PKR involves RNA-dependent autophosphorylation leading to inhibition of translation. PKR has an N-terminal dsRNA-binding domain that can interact non-sequence specifically with long (>33 bp) stretches of dsRNA leading to activation. In addition, certain viral and cellular RNAs containing non-Watson-Crick structures and multiple, shorter dsRNA sections can regulate PKR. In an effort to identify novel binders and possible activators of PKR, we carried out selections on a partially structured dsRNA library using truncated and full-length versions of PKR. A library with 10(11) sequences was constructed and aptamers that bound to His6-tagged proteins were isolated. Characterization revealed a novel minimal RNA motif for activation of PKR with the following unified structural characteristics: a hairpin with a nonconserved imperfect 16-bp dsRNA stem flanked by 10-15-nt single-stranded tails, herein termed a "ss-dsRNA motif." Boundary experiments revealed that the single-stranded tails flanking the dsRNA core provide the critical determinant for activation. The ss-dsRNA motif occurs in a variety of cellular and viral RNAs, suggesting possible novel functions for PKR in nature.
Zheng
Xiaofeng
X
Department of Chemistry, The Pennsylvania State University, University Park, PA 16802, USA.
Bevilacqua
Philip C
PC
D001483:Base Sequence; D004789:Enzyme Activation; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D012330:RNA, Double-Stranded / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000378:metabolism*
D012330:RNA, Double-Stranded; D019892:eIF-2 Kinase