Editors' ChoiceCancer

Bound by a lncRNA

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Science Signaling  24 Mar 2015:
Vol. 8, Issue 369, pp. ec69
DOI: 10.1126/scisignal.aab1668

Chronic inflammatory signaling contributes to the progression of cancer, and the transcription factor NF-κB is a key mediator of inflammatory signals. NF-κB activity is controlled by cytoplasmic sequestration through its interaction with IκB, and phosphorylation of IκB by the kinase IKK is one mechanism for releasing NF-κB, enabling it to translocate to the nucleus and regulate gene expression. Liu et al. identified NKILA (NF-κB interacting long noncoding RNA) in a screen for changes in the expression of long noncoding RNAs (lncRNAs) in response to inflammatory cytokine stimulation of the highly metastatic breast cancer cell line MDA-MB-231. The abundance of NKILA increased in response to cytokine stimulation. The basal abundance of this lncRNA was higher in breast cancer cell lines with low invasive behavior (low metastatic cells), which also had lower amounts of active NF-κB, than in the breast cancer cell lines with a high metastatic phenotype and higher amount of active NF-κB, which had higher amounts of active NF-κB. Overexpression of NKILA in MDA-MB-231 (high metastatic) cells reduced basal NF-κB activity and IκB phosphorylation, inhibited cytokine-induced stimulation of NF-κB activity, increased apoptosis in culture, and reduced metastasis in xenografts; whereas silencing NKILA had the opposite effects. Microscopic analysis indicated that the p65 subunit of NF-κB and NKILA colocalized in the cytoplasm, and NKILA coimmunoprecipitated with the p65 or p50 subunits of NF-κB or with IκBα, but not with IKKβ, and the interaction required p65. In vitro analysis of the interaction of NKILA deletion constructs with p65 and IκBα indicated that hairpin A of NKILA interacted with p65 and that both hairpin A and B were necessary for the formation of a stable complex with p65 and IκBα. In an in vitro kinase assay NKILA added prior to the addition of IKK prevented IKK-mediated phosphorylation of IκBα in an IκΒα-p65 complex. Mass spectrometry analysis of IκBα biotinylated at the sites phosphorylated by IKK and in vitro kinase assays with mutated NKILA indicated that hairpin C of NKILA physically blocked the phosphorylation sites. A microRNA (miRNA) screen identified several miRNAs that inhibited NKILA expression in MCF7 (low metastatic) cells, two of which—miR-103 and miR-107—exhibited an opposite expression pattern to NKILA. Expression of oligonucleotide mimics of these two miRNAs reduced the abundance of NKILA when introduced into MCF7 cells or antisense oligonucleotides, for these two miRNAs increased the abundance of NKILA when introduced into MDA-MB-231 cells. Analysis of breast cancer patient samples and normal breast tissue or primary cells derived from such tissues confirmed that NKILA was most abundant in normal breast tissue, reduced in cancer cells and tissue, and lowest in metastatic cancers. Low NKILA abundance correlated with poor patient survival. The induction of NKILA by inflammatory cytokines observed in the cancer cell lines was confirmed in the primary cultures, and silencing NKILA in the normal breast epithelial cells increased active NF-κB. Thus, this regulatory lncRNA represents an important link between inflammation and cancer.

B. Liu, L. Sun, Q. Liu, C. Gong, Y. Yao, X. Lv, L. Lin, H. Yao, F. Su, D. Li, M. Zeng, E. Song, A cytoplasmic NF-κB interacting long noncoding RNA blocks IκB phosphorylation and suppresses breast cancer metastasis. Cancer Cell 27, 370–381 (2015). [PubMed]

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