TWIST1 associates with NF-κB subunit RELA via carboxyl-terminal WR domain to promote cell autonomous invasion through IL8 production

Abstract

BackgroundMetastasis is the primary cause of death for cancer patients. TWIST1, an evolutionarily conserved basic helix-loop-helix (bHLH) transcription factor, is a strong promoter of metastatic spread and its expression is elevated in many advanced human carcinomas. However, the molecular events triggered by TWIST1 to motivate dissemination of cancer cells are largely unknown.ResultsHere we show that TWIST1 induces the production of interleukin 8 (IL8), which activates matrix metalloproteinases and promotes invasion of breast epithelial and cancer cells. In this novel mechanism, TWIST1-mediated IL8 transcription is induced through the TWIST1 carboxy-terminal WR (Trp-Arg) domain instead of the classic DNA binding bHLH domain. Co-immunoprecipitation analyses revealed that the WR domain mediates the formation of a protein complex comprised of TWIST1 and the nuclear factor-kappaB (NF-κB) subunit RELA (p65/NF-κB3), which synergistically activates the transcriptional activity of NF-κB. This activation leads to increased DNA binding affinity of RELA to the IL8 promoter and thus induces the expression of the cytokine. Blockage of IL8 signaling by IL8 neutralizing antibodies or receptor inhibition reduced the invasiveness of both breast epithelial and cancer cells, indicating that TWIST1 induces autonomous cell invasion by establishing an IL8 antocrine loop.ConclusionsOur data demonstrate that the TWIST1 WR domain plays a critical role in TWIST1-induced IL8 expression through interactions with and activation of NF-κB. The produced IL8 signals through an autocrine loop and promotes extracellular matrix degradation to enable cell invasion across the basement membrane.

A breast cancer stem cell niche supported by juxtacrine signalling from monocytes and macrophages

“…The Twist EMT-TF has been shown to mediate IL8 transcription in cooperation with RelA NF-κB subunit 50 . Indeed, we detected the association of the p50 NF-κB subunit with the endogenous Twist protein in the HMLER90hi cells by co-immunoprecipitation (Fig.…”

TWIST1 drives cisplatin resistance and cell survival in an ovarian cancer model, via upregulation of GAS6, L1CAM, and Akt signalling

“…We then transfected Ov8GFP cells with either TWIST1 or sh492, a previously validated shRNA against TWIST12425, using the pCI-Neo G418-selectable plasmid vector system. Following G418 selection of cells with stably integrated plasmid, we verified that TWIST1 was differentially expressed in the two cell lines – referred to hereafter as Ov8GFP-TWIST1 and Ov8GFP-sh492 – via western blot (Fig.…”

An evolutionarily conserved DNA architecture determines target specificity of the TWIST family bHLH transcription factors

“…Instead, the double E-box-binding model described here could possibly be the mechanistic explanation for the requirement of the WR domain in TWIST-mediated transcription regulation. The WR domain was also reported to bind to RUNX2 (Bialek et al 2004), SOX9 (Gu et al 2012), the PPA E3 ligase (Lander et al 2011), p53 (Piccinin et al 2012), and RELA (Li et al 2012) in certain biological settings. It is possible that the WR domain has pleiotropic functions as a homotypic or heterotypic protein-protein interaction module in different cells or species or during different developmental programs, which could greatly expand the flexibility of target gene regulation by TWIST complexes.…”