A 36-base hairpin within lncRNA DRAIC, which is modulated by alternative splicing, interacts with the IKKα coiled-coil domain and inhibits NF-κB and tumor cell phenotypes

Abstract

A tumor-suppressive long noncoding RNA (lncRNA) DRAIC (down-regulated RNA in cancers) inhibits NF-{kappa}B activity and physically interacts with IKK, a kinase component of the IKK complex, in several cancer types. Here we explore the precise molecular mechanism involved in this interaction and suppression. Using SHAPE-MaP, we identified a 36-nucleotide hairpin (A+B) within DRAIC that is necessary and sufficient for its anti-oncogenic function. RNA immunoprecipitation (RIP) and Electrophoretic mobility shift assays (EMSA) confirmed this hairpin physically interacts with the coiled coil domain of IKK. A+B RNA has a high binding affinity (KD [~]1-7 nM) to the coiled-coil domain of IKK. The binding of A+B disrupts the dimerization of NEMO and IKK coiled-coil domains, a critical step for IKK action. Consistent with this, A+B inhibits the phosphorylation of the NF-{kappa}B inhibitor I{kappa}B and suppresses NF-{kappa}B activity. Publicly available tumor RNAseq data revealed that alternative splicing modulates the presence of this critical hairpin: the inclusion of exon 4a (encoding one side of the A+B hairpin) in lung tumors correlates with reduced NF-{kappa}B activity. By demonstrating that the A+B hairpin is both necessary and sufficient to inhibit IKK and oncogenic phenotypes, this study underscores the centrality of IKK interaction and NF-{kappa}B inhibition in DRAIC-mediated cancer suppression and indicates that the activity of this lncRNA is regulated by alternative splicing. This study also reveals the first example of a short RNA disrupting coiled-coil dimerization, offering a new approach to disrupt such dimerization in cancer biology.