TARGETED EXPRESSION OF MDM2 UNCOUPLES S PHASE FROM MITOSIS AND INHIBITS MAMMARY-GLAND DEVELOPMENT INDEPENDENT OF P53

MDM2 is a cellular protein that binds to and inactivates the p53 tumor suppressor protein. Although mdm2 has been shown to function as an on cogene in vitro, all studies to date have assessed MDM2 activities in the presence of p53, implicating p53 inactivation in MDM2-directed tra nsformation. To determine the role of MDM2 in the cell cycle and in tu morigenesis and whether or not this role is dependent on p53, an MDM2 minigene was expressed during gestation and lactation in the mammary g land of both wild-type p53 (p53+/+) and p53 knockout (p53-/-) mice usi ng the bovine beta-lactoglobulin promoter. In six different transgenic mouse lines, deregulated expression of MDM2 inhibited normal developm ent and morphogenesis of the mammary gland, and caused cellular hypert rophy and nuclear abnormalities. These abnormalities included both mul tinucleated cells and enlarged cells with giant nuclei. Although there were fewer epithelial cells present in the transgenic mammary gland, no apoptosis was observed. Instead, BrdU incorporation and PCNA staini ng showed that 12%-27% of the transgenic mammary epithelial cells were in S phase at a time when normal cells were terminally differentiated . Analysis of DNA content showed that 30%-45% of the cells were polypl oid, with DNA contents up to 16N, indicating that overexpression of MD M2 caused mammary epithelial cells to undergo multiple rounds of S pha se without cell division. This phenotype was similar in the p53+/+ and p53-/- background, demonstrating a role for MDM2 in the regulation of DNA synthesis that is independent of the ability of MDM2 to inhibit p 53 activity. Additionally, multiple lines of BLGMDM2 transgenic mice d eveloped mammary tumors, confirming that overproduction of MDM2 contri butes to tumorigenesis in epithelial cells in vivo.