BMP4 depletion by miR-200 inhibits tumorigenesis and metastasis of lung adenocarcinoma cells

Jeong Seon Kim, Jonathan M. Kurie, Young-Ho Ahn

MicroRNA-200 (miR-200) suppresses the epithelial-mesenchymal transition of various cancer cells, including lung adenocarcinoma cells. We found that bone morphogenetic protein 4 (BMP4) was decreased in miR-200-overexpressing cells and epithelial-like lung cancer cells. In this study, we investigated the mechanism and role of BMP4 depletion by miR-200 in murine lung adenocarcinoma cells. BMP4 expression levels in murine lung cancer cells were measured by quantitative reverse transcription-PCR (qRT-PCR) and Western blotting. Promoter and 3'-untranslated region (UTR) luciferase reporter assays were performed to discover the mechanism of regulation of BMP4 by miR-200. Murine lung cancer cells were transfected with Bmp4 shRNAs, which were then injected into syngeneic mice to measure their tumorigenic and metastatic potential and cultured on Matrigel to study the influence of BMP4 on 3-D acinus formation. miR-200 down-regulated BMP4 via direct targeting of the GATA4 and GATA6 transcription factors that stimulate Bmp4 transcription. BMP4 up-regulated JAG2, an upstream factor of miR-200; therefore, JAG2, miR-200, and BMP4 form a regulatory loop. Bmp4 knockdown suppressed cancer cell growth, migration, and invasion and inhibited tumorigenesis and metastasis of lung cancer cells when injected into syngeneic mice. In addition, BMP4 was required for normal acinus formation in Matrigel 3-D culture of murine lung cancer cells, which may be mediated by MYH10, a downstream target of BMP4. BMP4 functions as a pro-tumorigenic factor in a murine lung cancer model, and its transcription is regulated by miR-200 and GATA4/6. Thus, we propose that BMP4 and its antagonists may be suitable therapeutic targets for the treatment of lung cancer.