The PvuII and XbaI polymorphisms of the estrogen receptor α (ER1) gene have been variably associated with type 2 diabetes (T2D) in several populations. However, this association has not been studied in Iranian subjects and we hypothesized that the ER1 variants might be associated with T2D and related metabolic traits in this population. The PvuII and XbaI genotypes were determined by PCR-RFLP in 377 normoglycemic controls and 155 T2D patients. Bonferroni correction was applied for the correction of multiple testing. No significant association was found between the allele and genotype frequencies of PvuII and XbaI variants with T2D in females. In a dominant model (PP vs. Pp+pp), the frequency of the Pp+pp genotype was higher in normoglycemic subjects compared to T2D patients [85.5% vs. 66.7%, OR 0.22 (0.08-0.55), P=0.001]. Four possible haplotypes were observed in the population, whereas haplotype TA had a higher frequency in male T2D subjects than the controls. Furthermore, non-diabetic male subjects carrying the genotype of PP had a higher fasting glucose levels than the individuals with the genotype of Pp+pp (P=0.013). Multivariate logistic regression analysis showed that PvuII polymorphism was the independent determinants of T2D in males [OR 4.37 (1.61-11.86), P=0.004]. No association was found between the XbaI polymorphism and diabetes in male group. Our results suggest that the ER1 polymorphisms might associate with T2D and fasting glucose among Iranian male subjects.