Peroxisome proliferator–activated receptor (PPAR) α is a ligand‐activated transcription factor that has been linked with rodent hepatocarcinogenesis. It has been suggested that PPARα mRNA expression levels are an important determinant of rodent hepatic tumorigenicity. Previous work in rat mammary gland epithelial cells showed significantly increased PPARα mRNA expression in carcinomas, suggesting the possible role of this isoform in rodent mammary gland carcinogenesis. In this study we sought to determine whether PPARα is expressed and dynamically regulated in human breast cancer MCF‐7 and MDA‐MB‐231 cells. Having established the presence of PPARα in both cell types, we then examined the consequence of PPARα activation, by its ligands Wy‐14,643 and clofibrate, on proliferation. With real‐time reverse transcriptase–polymerase chain reaction, we showed that PPARα mRNA was dynamically regulated in MDA‐MB‐231 cells and that PPARα activation significantly increased proliferation of the cell line. In contrast, PPARα expression in MCF‐7 cells did not change with proliferation during culture and was present at significantly lower levels than in MDA‐MB‐231 cells. However, PPARα ligand activation still significantly increased the proliferation of MCF‐7 cells. The promotion of proliferation in breast cancer cell lines following PPARα activation was in stark contrast to the effects of PPARγ‐activating ligands that decrease proliferation in human breast cancer cells. Our results established the presence of PPARα in human breast cancer cell lines and showed for the first time that activation of PPARα in human breast cancer cells promoted proliferation. Hence, this pathway may be significant in mammary gland tumorigenesis. © 2002 Wiley‐Liss, Inc.