The estrogen receptor (ER) is thought to play a crucial role in the regulation of many life processes, including development, reproduction and normal physiology. Because there have been no known mutations of the estrogen receptor in normal tissue of humans and animals, its presence and tissue distribution is thought to be essential for survival. Using the techniques of homologous recombination, we have disrupted the ER gene and have produced a line of transgenic mice possessing the altered ER gene (ERKO). The mouse ER gene was disrupted by inserting a 1.8 kb PGK-Neomycin sequence into exon 2, approximately 280 bp downstream of the transcription start codon. The correct targeting of the disruption was demonstrated by Southern blot analysis and PCR. Western blot analysis of uterine preparations from ERKO females showed no detectable ER protein. Heterozygotes had one half the level of ER protein compared to wild-type animals. Estrogen insensitivity was confirmed using estrogen agonists, estradiol, hydroxy tamoxifen, diethylstilbestrol treatment for 3 days which resulted in a 3-4-fold increase in uterine wet weight and vaginal cornification in wild-type females, while ERKO mice were totally unresponsive. These data were further supported by the failure of estrogen or EGF treatment to induce DNA synthesis in uterine tissue of similarly treated mice. Lactoferrin, an estrogen-responsive gene in the uterus, was also assayed by Northern blot. Wild-type mice treated with a single estradiol injection showed a 350-fold induction in lactoferrin mRNA. while ERKO females showed no detectable response. Both male and female animals survive to adulthood with normal gross external phenotypes. As expected, females are infertile and demonstrate hypoplastic uteri and hyperemic ovaries with no apparent corpora lutea. Males are also infertile, with atrophy of the testes and seminiferous tubule dysmorphogenesis. Although the reproductive capabilities have been altered with a dramatic effect on the gonads, prenatal development of the reproductive tracts of both sexes appear to be independent of an ER-mediated response. Analysis of the mammary glands of the ERKO females at 4 months of age showed a primitive ductal rudiment rather than the fully developed ductal tree seen in wild-type siblings. Also absent were the terminal end buds seen during normal ductal morphogenesis. Both sexes show a decrease in skeletal bone density, supporting a direct role for ER action in bone. A single patient is described who is homozygous for a point mutation in the human ER gene at codon 157. The mutation produces a truncation of the ER protein and results in estrogen insensitivity syndrome. Most significant of the clinical findings are effects on skeletal bone density and retarded bone age. Findings from the patient and mice suggest that the absence of functional ER is not lethal. Mutation in the ER gene is present in the human population. Further characterization of the mice and identification of additional patients will be required to more fully understand the consequences of ER gene mutations.