α-Class GST (Gsta) represents an essential component of cellular antioxidant defense mechanisms in both the liver and the kidney. Estrogens and thyroid hormones (TH) play central roles in animal development, physiology, and behavior. Evidence of the overlapping functions of thyroid hormones and estrogens has been shown, although the molecular mechanisms are not always clear. We evaluated an interaction between TH and estradiol in regulating kidney Gsta expression and function. First, we observed that female mice expressed greater amounts of Gsta compared with males and showed an opposite pattern of expression in TRβ knock-in mice. To further investigate these sex differences, hypothyroidism was induced by a 5-propyl-2-thiouracil diet, and hyperthyroidism was induced by daily T₃ injections. Hypothyroidism increased kidney Gsta expression in male mice but not in female mice, indicating that sex hormones could be influencing the regulation of Gsta by thyroid hormones. To analyze this hypothesis, ovariectomized females were subjected to hypo- and hyperthyroidism, which led to a male profile of Gsta expression. When hypo- or hyperthyroid ovariectomized mice were treated with 17β-estradiol benzoate, we were able to confirm that estradiol was interfering with TH modulation; Gsta expression is increased by T₃ when estradiol is present and decreased by T₃ when estradiol is absent. Using proximal tubule cells, we also showed that estradiol and T₃ worked together to modulate Gsta expression in an overlapping fashion. In summary, 1) the sex difference in the basal expression of Gsta impacts the detoxification process, 2) kidney Gsta expression is regulated by TH in males and females but in opposite directions, and 3) T₃ and estradiol interact directly in renal proximal cells to regulate Gsta expression in females.