Zonula Occludens-1/Nf-Kappa B/Cxcl8: A New Regulatory Axis For Tumor Angiogenesis

Zonula occludens-1 (ZO-1) is a submembrane scaffolding protein thatmay display proinvasive functions when it relocates from tight junctions into the cytonuclear compartment. This article examines the functional involvement of ZO-1 in CXCL8/ IL-8 chemokine expression in lung and breast tumor cells. ZO-1 small interfering RNA and cDNA transfection experiments emphasized regulation of CXCL8/ IL-8 expression via a cytonuclear pool of ZO-1. Luciferase reporter assays highlighted a 173-bp region of CXCL8/ IL-8 promoter that responded to ZO-1. Moreover, by using mutated promoter constructs, we identified a NF-kappa B site as critical in this activation. Furthermore, NF-kappa Bpathwaysignaling analysis revealedboth IkBaandp65phosphorylationinZO-1-overexpressingcells, and subsequent p65 silencing validated its requirement for CXCL8/ IL-8 induction. Investigation of the functional implication of this regulatory axis next showed the proangiogenic activity of ZO-1 in both ex vivo and in vivo angiogenesis assays. Finally, we found that non-small-cell lung carcinoma that presented a cytonuclear ZO-1 pattern was significantly more angiogenic that that without detectable cytonuclear ZO-1 expression. Taken together, our results demonstrate that ZO-1 regulatesCXCL8/IL-8 expression via theNF-kappa B signaling pathway and its p65 subunit, which subsequently modulates the transcription of IL-8. We also provide evidence of a newly identified regulatory pathway that could promote angiogenesis. Thus, our results support the concept that the ZO-1 shuttle from the cell junction to the cytonuclear compartment may affect both the intrinsic invasive properties of tumor cells and the establishment of the protumoral microenvironment.-Lesage, J., Suarez-Carmona, M., Neyrinck-Leglantier, D., Grelet, S., Blacher, S., Hunziker, W., Birembaut, P., No " el, A., Nawrocki-Raby, B., Gilles, C., Polette, M. Zonula occludens-1/ NF-kappa B/ CXCL8: a new regulatory axis for tumor angiogenesis. FASEB J. 31, 1678-1688 (2017). www.fasebj.org