Details

Autor(en) / Beteiligte

• Kamei, Daisuke
• Yamada, Michiyuki

Titel

Interactions of heterogeneous nuclear ribonucloprotein D-like protein JKTBP and its domains with high-affinity binding sites

Ist Teil von

• Gene, 2002-09, Vol.298 (1), p.49-57

Ort / Verlag

Elsevier B.V

Erscheinungsjahr

2002

Link zum Volltext

Quelle

Elsevier Journal Backfiles on ScienceDirect (DFG Nationallizenzen)

Beschreibungen/Notizen

• JKTBP proteins consisting of two canonical RNA binding domains (RBDs) and a glycine-rich carboxyl domain are nucleocytoplasmic shuttling proteins. We studied in vivo and in vitro interactions between JKTBP and RNA. UV cross-linking experiments on HL-60 cells indicated that following RNA synthesis inhibition by actinomycin D, JKTBP1 accumulated in the cytoplasam is bound to poly(A) + RNAs. Recombinant JKTBP1 protein blots could bind poly(A) + RNAs, but not poly(A) − RNAs. For examination of RNA binding specificity of JKTBP, we enriched high binding sites from pools of 20 nt random sequence-containing RNAs by a selection/amplification method. After eight rounds of a selection and amplification, >20 sequences for each of JKTBPs 1 and 2 were identified. Their consensus high-affinity site was ACUAGC. Approximate K ds of JKTBPs 2 and 1 were estimated to be 6–12 nM for the selected sequences by filter binding assays. JKTBP deletion analysis indicated that not individual RBDs, both RBDs and the N-terminal 15 amino acids of the carboxyl domain are required for sequence-specific and high-affinity binding. These results indicate that JKTBP is a sequence-specific RNA binding protein differing from the related heterogeneous nuclear ribonucleoproteins A1 and D.