Nox4 involvement in TGF-beta and SMAD3-driven induction of the epithelial-to-mesenchymal transition and migration of breast epithelial cells

Wednesday, October 10, 2012 — Poster Session II
Noon – 2:00 p.m	Natcher Conference Center, Building 45	NIAID	CANCER-2

Authors

• H.E. Boudreau
• B.W. Casterline
• B. Rada
• A. Korzeniowska
• T.L. Leto

Abstract

The epithelial-to-mesenchymal transition (EMT) is the development of increased cell plasticity that occurs normally during wound healing and embryonic development and can be coopted for cancer metastasis. TGF-beta induces EMT but the mechanism is unclear. Our studies suggest NADPH oxidase 4 (Nox4) is a source of reactive oxygen species (ROS) affecting cell migration and fibronectin expression, an EMT marker, in normal and metastatic breast epithelial cells. We found TGF-beta induces Nox4 expression and ROS generation in MCF10A and MDA-MB-231 breast epithelial cells. Conversely, cells expressing a dominant-negative form of Nox4 or Nox4 shRNA showed lower ROS upon TGF-beta treatment. Expression of a constitutively active TGF-beta receptor type I increased Nox4 promoter activity, mRNA and protein expression, and ROS. Additionally, a constitutively active SMAD3 mutant increased Nox4 promoter activity, whereas dominant-negative SMAD3 or SIS3, a SMAD3-specific inhibitor, had the opposite effect. Inhibition of Nox4 or SMAD3 blunted TGF-beta induced wound healing and cell migration. Finally, Nox4 plays a role in TGF-beta regulation of fibronectin expression, as dominant-negative Nox4 reduced fibronectin mRNA in TGF-beta treated cells. Collectively, these data indicate Nox4-derived ROS production may be critical for progression of the EMT in breast epithelial cells, and thereby has therapeutic implications.

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