Maspin, a member of the serine protease inhibitor (serpin) family, is a tumor suppressor in breast and prostate cancer. To address molecular mechanisms underlying maspin's activity, we restored its expression in invasive carcinoma cells and analyzed the resulting changes by shotgun proteomics. Using a mass spectrometry-based multidimensional proteomic method, we observed changes to the expression of approximately 27% of the detectable proteome. In particular, we noted changes to the expression of proteins that regulate cytoskeletal architecture, cell death, and protein turnover. In each case, changes in protein expression were accompanied by measurable changes in tumor cell phenotype. Thus, maspin-expressing cells exhibit a more prominent actin cytoskeleton, a reduced invasive capacity, an increased rate of spontaneous apoptosis, and an altered proteasome function. These observations reveal for the first time the far reaching effects of maspin on multiple protein networks and a new hypothesis of maspin function based on the regulation of proteasome function.