Characterization of the direct physical interaction of nc886, a cellular non-coding RNA, and PKR

• Published in: FEBS letters Vol. 586; no. 19; pp. 3477 - 3484
• Main Authors: Jeon, Sung Ho, Lee, Kwanbok, Lee, Kwang Soo, Kunkeaw, Nawapol, Johnson, Betty H., Holthauzen, Luis Marcelo F., Gong, Bin, Leelayuwat, Chanvit, Lee, Yong Sun
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 21.09.2012
• Subjects: Base Sequence; Binding Sites; Binding, Competitive; calf-intestinal alkaline phosphatase; CIP; Double-stranded RNA; dsRBM; dsRNA; dsRNA binding motif; eIF-2 Kinase - chemistry; eIF-2 Kinase - metabolism; electrophoretic mobility shift assays; EMSA; Enzyme Activation; HEK293 Cells; Humans; IFN-β; interferon-β; Kinetics; messenger RNA; microRNA; miRNA; Molecular Sequence Data; mRNA; nc886; ncRNA; non-coding RNA; non-coding RNA-886; Nucleic Acid Conformation; PKR; PNK; polynucleotide kinase; Protein Kinase RNA-activated; Protein Structure, Tertiary; Protein–RNA interaction; response unit; RNA, Untranslated - chemistry; RNA, Untranslated - genetics; RNA, Untranslated - metabolism; single-stranded RNA; ssRNA; Vault RNA; vtRNA; wildtype; dsRNA; RU; Double-stranded RNA; Protein–RNA interaction; mRNA; CIP; ncRNA; PNK; ssRNA; Vault RNA; IFN-β; vtRNA; EMSA; PKR; dsRBM; miRNA; nc886; wt
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/j.febslet.2012.07.076

► First characterization of in vitro interaction between PKR and nc886. ► Direct binding of nc886 to PKR with an affinity comparable to that of dsRNA. ► Two RNA binding domains of PKR recognize the central region of nc886. ► PKR binds equally well to 5′-mono, tri- or de-phosphorylated nc886. ► nc886 sets a threshold for cellular PKR activation. We have recently shown that nc886 (pre-miR-886 or vtRNA2-1) is not a genuine microRNA precursor nor a vault RNA, but a novel type of non-coding RNA that represses PKR, a double-stranded RNA (dsRNA) dependent kinase. Here we have characterized their direct physical association. PKR’s two RNA binding domains form a specific and stable complex with nc886’s central portion, without any preference to its 5′-end structure. By binding to PKR with a comparable affinity, nc886 competes with dsRNA and attenuates PKR activation by dsRNA. Our data suggest that nc886 sets a threshold for PKR activation so that it occurs only during genuine viral infection but not by a minute level of fortuitous cellular dsRNA.