Abstract 5406: Transcriptional upregulation of NADPH Oxidase 1 (NOX1) by Interleukin-4 (IL-4) enhances tumor cell proliferation and reactive oxygen production in human colon cancer cells.
Authors: Liu Han , Juhasz Agnes , Jiang Guojian , Antony Smitha , Meitzler Jennifer L. , Wu Yongzhong , Lu Jiamo , Roy Krishnendu , Doroshow James H. 
Published: August 2014

Reactive oxygen species (ROS) produced by the family of NOX genes play a critical role in cytokine-mediated signal transduction in both normal and tumor tissue. Human colon cancers, compared to adjacent normal tissues, overexpress NOX1; and premalignant, chronic inflammatory conditions of the colon (ulcerative colitis, Crohn's disease) are associated with NOX up-regulation. However, a mechanistic understanding of how cytokines might increase ROS production and NOX1 expression in colon cancer remains unclear. We screened a panel of human colon cancer cells with a series of pro-inflammatory cytokines to evaluate their effects on NOX1 expression and function, as well as cell proliferation. Cytokine IL-4 significantly up-regulated NOX1 expression (but not other members of the NOX family) in HT-29 (as well as other) colon cancer cells while increasing O2• production and growth; decreasing NOX1 expression by 90% in stable, NOX1-shRNA-containing HT-29 clones blocked IL-4-related enhancement of cell growth, NOX1 expression, or ROS production. IL-4 up-regulated and activated the GATA3 transcription factor in addition to the Jak1-Stat6 pathway in HT-29 cells. GATA3 played an important role in the transcriptional up-regulation of NOX1 by IL-4; RNAi-mediated silencing of Jak1, Stat6, or GATA3 expression attenuated IL-4-induced NOX1 up-regulation in HT-29 cells. Transient overexpression of GATA3 cDNA in the presence of IL-4 resulted in a further increase in NOX1 expression with enhanced ROS activity beyond IL-4 treatment alone. Using an 1139 bp segment of upstream sequence from the transcription start site of the human NOX1 promoter as a promoter-reporter assay, we found that IL-4 treatment increased NOX1 reporter gene transcription 4-fold in HT-29 cells. DNA sequence analysis of this upstream promoter suggests the presence of four putative GATA3 binding, cis-active elements that may be sufficient for maximal IL-4-enahanced activity of the NOX1 promoter. Ongoing mutational analysis of the NOX1 promoter will further define the role of GATA3 in the control of NOX1 transcriptional up-regulation by IL-4. In summary, our results support a role for IL-4 in producing a NOX1-dependent pro-oxidant milieu that could increase the growth potential of colon cancer cells.

Citation Format: Han Liu, Agnes Juhasz, Guojian Jiang, Smitha Antony, Jennifer L. Meitzler, Yongzhong Wu, Jiamo Lu, Krishnendu Roy, James H. Doroshow. Transcriptional upregulation of NADPH Oxidase 1 (NOX1) by Interleukin-4 (IL-4) enhances tumor cell proliferation and reactive oxygen production in human colon cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5406. doi:10.1158/1538-7445.AM2013-5406