PRIP (peroxisome proliferator-activator receptor interacting protein) is a nuclear receptor coactivator required for mammary gland development. To understand the function of PRIP in breast tumorigenesis, we established a mammary tumor cell line with the PRIP(Loxp/Loxp) genotype. By knocking out the PRIP gene in the tumor cell line, we demonstrated that PRIP deficiency led to inhibited tumor formation without affecting tumor cell proliferation. The PRIP deficiency was associated with decreased cell invasion and migration capabilities. We found that PRIP deficiency substantially reduced FOS gene expression. A chromatin immunoprecipitation assay revealed that PRIP was recruited to the FOS promoter. In addition, we demonstrated that PRIP also directly up-regulated the FOS gene expression in human breast cancer cells. Promoter analysis showed that PRIP acted through serum-responsive factor to regulate FOS gene expression. Finally, by re-expressing the FOS gene, we confirmed that the inhibited tumor formation of PRIP-deficient tumor cells was due to reduced expression of the FOS gene.