Although X chromosome transfer experiments indicated that tumor suppressor genes are present on the X chromosome, they have not been previously identified. In this report, we show that the ETS transcription factor MEF (ELF4), which is located on chromosome Xq26.1, possesses tumor suppressive capability. MEF expression was up-regulated by 5-azacytidine in some cancer cell lines. MEF overexpression induced morphological changes, such as the conversion of normally loose cell-cell contacts to strong interactions similar to those seen in the presence of matrix metalloproteinase (MMP) inhibitor BB94. In the colony formation assay, A549 cells, but not MEF-overexpressing cells, formed colonies in soft agar culture. Furthermore, MEF-overexpressing cells s.c. injected in the nude mice did not grow, whereas the control cells did. The A549 tumors were poorly differentiated, whereas the MEF-overexpressing tumors were well differentiated. By immunostaining with CD31, a marker on vascular endothelial cells, we found that tumor angiogenesis was significantly suppressed in the tumors formed from MEF-overexpressing cells. In addition, the conditioned media from A549 cell cultures stimulated the migration of human umbilical vein endothelial cells, whereas conditioned media from MEF-overexpressing cell cultures had less of an effect. By gelatin zymography, Western blotting analysis, and immunohistochemistry, we found that the expression levels of MMP-9 and MMP-2 were significantly reduced in MEF-overexpressing tumors. Immunohistochemical analyses showed that interleukin (IL)-8 expression was reduced in the MEF-overexpressing tumors in nude mice. Furthermore, IL-8 mRNA expression in vitro was significantly down-regulated in MEF-overexpressing cells, compared with A549 cells. MEF suppressed the transcription and promoter activities of the genes encoding MMP-9 and IL-8, whereas ETS-2 up-regulated these activities. Therefore, we propose that MEF is a candidate tumor suppressor gene on the X chromosome with activities that are opposite to those of ETS-2.