The 200-amino-acid repeat (HIN-200) gene family with the hematopoietic interferon (IFN)-inducible nuclear protein encodes highly homologous proteins involved in cell growth, differentiation, autoimmunity, and tumor suppression. IFIX is the newest member of the human HIN-200 family and is often downregulated in breast tumors and breast cancer cell lines. The expression of the longest isoform of IFIX gene products, IFIXalpha1, is associated with growth inhibition, suppression of transformation, and tumorigenesis. However, the mechanism underlying the tumor suppression activity of IFIXalpha1 is not well understood. Here, we show that IFIXalpha1 downregulates HDM2, a principal negative regulator of p53, at the posttranslational level. IFIXalpha1 destabilizes HDM2 protein and promotes its ubiquitination. The E3 ligase activity of HDM2 appears to be required for this IFIXalpha1 effect. Importantly, HDM2 downregulation is required for the IFIXalpha1-mediated increase of p53 protein levels, transcriptional activity, and nuclear localization, suggesting that IFIXalpha1 positively regulates p53 by acting as a negative regulator of HDM2. We found that IFIXalpha1 interacts with HDM2. Interestingly, the signature motif of the HIN-200 gene family, i.e., the 200-amino-acid HIN domain of IFIXalpha1, is sufficient not only for binding HDM2 but also for downregulating it, leading to p53 activation. Finally, we show that IFIX mediates HDM2 downregulation in an IFN-inducible system. Together, these results suggest that IFIXalpha1 functions as a tumor suppressor by repressing HDM2 function.