Nur77 regulates hypoxia induced epithelial to mesenchymal transition in colon cancer cells

S. Deng1, J. Zeng2, A. Wong1. 1university of Hong Kong, Hong Kong, HONG KONG, 2Xiamen University, Xiamen, CHINA

Introduction: Colorectal cancer is one of the most common malignancies worldwide, and over 50% of patients develop metastasis. Epithelial-to-mesenchymal transition (EMT) is a process by which tumor cells lose cell-cell adhesive contact, and exhibit migratory/invasive mesenchymal-like phenotype. Hypoxia, an important tumor microenvironmental factor that contributes to cancer progression and metastasis, has been demonstrated to promote EMT [1]. Nur77 (also known as NR4A1, TR3, or NGFIB) is a unique member of the orphan nuclear receptor superfamily. It has been reported as a growth factor-inducible immediate early gene that regulates cellular proliferation, apoptosis, and metastasis [2]. In this study, we aim to characterize the underlying mechanisms of Nur77 regulation in EMT under hypoxia.

Method: Colorectal cancer cells HCT116 and SW480 were transfected with Nur77 siRNA for 30 h, followed by 4 h hypoxic incubation. Cell morphologic changes were examined under phase contrast microscopy and cell migration was analyzed using a scratch assay. Western blot analysis was performed to assess EMT marker (Snail, Slug, E- and N-cadherin) and stem cell marker (CD44, CD133) expression. Data were shown as mean ± SD and statistical analysis was performed using one way ANOVA followed by unpaired Student’s t-test.

Results: We found that under hypoxic condition, HCT116 and SW480 cells transfected with Nur77 siRNA about 60% of scattered colonies exhibited increased cell-cell contact and compact epithelial structure, suggesting that EMT was reversed. Consistently, depletion of Nur77 also significantly abolished the cell migration induced by hypoxia. These changes were confirmed by a decrease of N-cadherin, Snail, Slug, and an increase of E-cadherin expression and CD44 and CD133 expression. In addition, we showed that a non-genomic signaling via mTOR is critical for the hypoxia-induced EMT and subsequent stem cell characteristics.

Conclusion: These results provide insights into the roles and mechanisms of Nur77 in EMT and metastasis under hypoxia (This work was supported by NSFC/RGCN_HKU740/14).

References:

1. Gilkes DM, et al. (2014) Nat Rev Cancer 14: 430-439.

2. Wang JR, et al. (2014) Carcinogenesis. 35: 2474-2484.