A 36-base hairpin within lncRNA DRAIC interacts with the IKKa coiled-coil domain, weakening the IKK complex and inhibiting NF-kappaB activation

Long noncoding RNAs have diverse functions, but not enough is known about the molecular mechanisms underlying their cellular functions. The 1.7 kb long down-regulated non-coding RNA in cancers (DRAIC) slows tumor growth and inhibits the activation of the transcription factor NF-kappaB through interactions with IKK subunits. Starting with SHAPE-MaP-based structure-function studies of DRAIC we have now identified the minimal regions of DRAIC and IKK involved in the interactions and the anti-cancer function of DRAIC. A hairpin structure comprised of bases 705-722/741-758 (referred to as A+B) within DRAIC is responsible for inhibiting NF-kappaB activity. A+B binds specifically with the IKKa coiled-coil domain with a KD of ~1-5 nM. The coiled-coil domain of IKKa (the kinase subunit) and NEMO (the substrate recognition subunit,) mediate the dimerization of the subunits in the functional kinase complex. Consistent with its interaction with the coiled-coil domain of IKKa, A+B weakens the interaction between NEMO and the IKKa coiled-coil domain in an in vitro complex formation assay. The 36 nt A+B RNA was sufficient and necessary for DRAIC to inhibit tumor cell migration, invasion, and clonal growth in multiple cancer cell-lines. Therefore, this minimal hairpin fragment of the growth-suppressive DRAIC lncRNA inhibits NF-kappaB activation by interacting with high affinity with the coiled-coil domain of IKKa, weakening the interaction between the kinase subunit and the substrate recognition subunit of IKK, thereby inhibiting I?B phosphorylation and NF-kappaB activation, and suppressing tumor phenotypes. Overall design: To discover functional structures within the DRAIC RNA, we performed SHAPE-MaP to obtain conformational flexibility measurements of the DRAIC active region from cell-extracted RNA.