Estrogen replacement therapy in postmenopausal women is associated with a reduction in colorectal cancer risk, potentially via interactions between 17beta-estradiol (E(2)) and the estrogen receptors (ER) alpha and beta. To study the role of E(2) in intestinal tumor inhibition, we separately crossed C57BL/6J-Min/+ (Min/+) mice with Eralpha(+/-) and Erbeta(+/-) mice to generate ER-deficient Min/+ progeny. We found an increased incidence of visible colon tumors and dysplastic microadenomas in ER-deficient Min/+ relative to Er(+/+)Min/+ controls. Small intestinal tumor numbers were unaffected. Invasive carcinomas were found only in Eralpha(+/-)Min/+ mice, suggesting that ERalpha plays additional non-cell autonomous roles that limit tumor progression. Histologic analyses of ER-deficient Min/+ colons, as well as colons from ovariectomized Min/+ mice (OvxMin/+) and E(2)-treated OvxMin/+ mice (OvxMin/+ +E(2)), revealed significant differences in crypt architecture, enterocyte proliferation, and goblet cell differentiation relative to Min/+ and Er(+/+)Apc(+/+) (wild-type) controls. The expression of ERalpha and ERbeta was regionally compartmentalized along the colonic crypt axis, suggesting functional antagonism. Our results indicate that ERalpha and ERbeta are inhibitory modifiers of Apc-dependent colon tumorigenesis. As a result, loss of E(2) and ER signaling in postmenopausal women may contribute to colorectal cancer development.