Fetal alcohol exposure (FAE) increases the risk of mammary tumorigenesis in adult offspring; however, the underlying mechanism remains unknown. This study tested the hypothesis that FAE shifts the mammary epithelial cell (MEC) composition toward one that promotes tumorigenesis. Pregnant Friend Virus BNIH Jacksondams bred to MMTV-Wnt1 male mice were given ad libitum access to 5% alcohol in 0.2% saccharin solution from GD9-10 and 10% alcohol in 0.2% saccharin from GD11-GD19 or 0.2% saccharin solution from GD9-GD19. Thoracic and inguinal mammary glands from wild-type (WT) and transgenic (Tg) female offspring were harvested at 5 and 10weeks of age and dissociated to yield a single cell suspension enriched for MECs for flow cytometry, mammosphere assay, and gene analysis. A subset of Tg offspring was followed for tumor formation. WT glands of FAE animals exhibited a decreased basal cell population and increased luminal: basal ratio at 10weeks of age. qRT-PCR analysis of total MECs found that Hey1 mRNA expression was increased in the WT FAE group at 10weeks of age. In Tg glands, FAE increased the luminal progenitor cell population at 5weeks of age but did not alter MEC composition at 10weeks of age. Tertiary mammosphere-forming efficiency was greater in the WT glands of FAE animals at 10weeks of age. Tumor latency was decreased in the FAE group. Flow cytometry analysis indicated that FAE females developed tumors with an increased basal cell population. These data indicate that FAE can shift MEC subpopulations, increasing the proportion of cells that are potentially vulnerable to transformation and affecting cancer risk.