Annexin-1 interacts with NEMO and RIP1 to constitutively activate IKK complex and NF-κB: implication in breast cancer metastasis

Abstract

The molecular mechanisms underlying constitutive nuclear factor-jB (NF-jB) activation in solid tumors has not been elucidated. We show that Annexin-1 (ANXA1) is involved in this process, and suppression of ANXA1 in highly metastatic breast cancer cells impedes migration and metastasis capabilities in vitro and in vivo. ANXA1 expression correlates with NF-jB activity, suggesting that ANXA1 may be required for the constitutive activity of IjB kinase (IKK) and NF-jB in highly metatstatic breast cancer. Gel-filtration analysis demonstrated that ANXA1 co-elutes with the members of the IKK complex and NF-jB signaling pathway, and immunoprecipitation confirmed that ANXA1 can bind to and interact with IKKc or NEMO, but not IKKa or IKKb. Importantly, silencing of ANXA1 prevents the interaction of NEMO and RIP1, which indicates that ANXA1 is required for the recruitment of RIP1 to the IKK complex, which may be important for the activation of NF-jB. Downstream targets of NF-jB include uPA and CXCR4, which can be modulated by ANXA1 silencing. CXCR4-mediated migration of breast cancer cell lines in response to CXCL12 was significantly modulated by ANXA1, indicating its importance in the tissue-specific migration of breast cancer cells. Chromatin immunoprecipitation experiments confirmed that in ANXA1 overexpressed cells, NF-jB was recruited to CXCR4 promoter without external stimulation, indicating that ANXA1 is critical for the constitutive activation of NF-jB in breast cancer to promote metastasis. Finally, we show that ANXA1 overexpression enhances metastasis and reduces survival in an intracardiac metastasis model, while ANXA1-deficient mice crossed with MMTV-PyMT mice display significantly less metastasis than their heterozygous littermates, indicating that ANXA1 is an important gene in breast cancer metastasis. Our data reveal that ANXA1 can constitutively activate NF-jB in breast cancer cells through the interaction with the IKK complex, and suggests that modulating ANXA1 levels has therapeutic potential to suppress breast cancer metastasis.

Quantitative Proteomics Profiling of Murine Mammary Gland Cells Unravels Impact of Annexin-1 on DNA Damage Response, Cell Adhesion, and Migration

“…Down-regulation of ANXA1 has also been shown in carcinogenic models to inhibit cell migration/invasion (15,48), suggesting that ANXA1 could possibly be pro-migratory in carcinogenesis. Recent studies from our lab and others have demonstrated that ANXA1 is indeed pro-migratory in cancer cells, whereas silencing ANXA1 reduces metastatic capability (15). Altogether, cell motility plays a central part in coordinating normal physiological and pathological processes including wound healing and metastasis.…”

“…One of the mechanisms proposed in this adhesion inhibition is the down- regulation and shedding of a cell adhesion molecule L-selectin on the leukocytes (46,47). Down-regulation of ANXA1 has also been shown in carcinogenic models to inhibit cell migration/invasion (15,48), suggesting that ANXA1 could possibly be pro-migratory in carcinogenesis. Recent studies from our lab and others have demonstrated that ANXA1 is indeed pro-migratory in cancer cells, whereas silencing ANXA1 reduces metastatic capability (15).…”

“…ANXA1 was reported as an important modulator for an epithelial-to-mesenchymal-like phenotypic switch via the transforming growth factor ␤ signaling pathway (14). Furthermore, our recent study demonstrates that ANXA1 is required for constitutive NF-B activity in basal cell carcinoma cell lines, which is of utmost importance to metastatic potential (15). Moreover, genomics approaches to studying molecular signatures associated with transformation and progression to breast cancer highlighted an up-regulation of ANXA1 in cellular transformation (16).…”

Annexin-A1 Regulates TLR-Mediated IFN-β Production through an Interaction with TANK-Binding Kinase 1

“…Our present findings on ANXA1 as a regulator of the TRIF pathway by binding to TRIF and TBK1 provides a mechanistic basis linking GC and TRIF pathway through ANXA1. We have previously reported that ANXA1 can bind to NEMO and RIP1 (40) to enhance the activation of NF-kB. It is possible that the same regions of ANXA1 bind to TBK1 as well to regulate IRF3 activity.…”

Pontin, a new mutant p53-binding protein, promotes gain-of-function of mutant p53

“…The gene products of ADRB1, RIPK1 belong to the gap junction signaling, which is important for cell motility. [29][30][31] STMN1, which has a role in cell migration, was reported to be upregulated by mutp53. 32 It is possible that some of these genes mediate the promoting effect of Pontin on mutp53 GOF.…”