There is evidence that transforming growth factor (TGF)beta acts as a suppressor of tumor initiation but also as a promoter of tumor progression when the antiproliferative effect of the TGFbeta signaling pathway has been overridden by other oncogenic mutations. Several somatic mutations that disrupt the TGFbeta-SMAD signaling pathway have been reported in human breast tumors. We have examined the association between single nucleotide polymorphisms (SNPs) in the TGFbeta1 gene and the incidence of invasive breast cancer in three case-control series, with a maximum of 3987 patients and 3867 controls, median age approximately 50 years, and range 22-92 years. The promoter SNP, C-509T, and the T +29C signal-peptide SNP (encoding Leu10Pro) are in strong linkage disequilibrium. They are both significantly associated with increased incidence of invasive breast cancer in a recessive manner [odds ratios: (TT versus C-carrier), 1.25; 95% confidence intervals 1.06-1.48; P = 0.009 and (ProPro versus Leu-carrier), 1.21; 95% confidence intervals 1.05-1.37; P = 0.01]. The G-800A SNP was not significantly associated with incidence of breast cancer. The C-509T SNP is not contained within a known consensus sequence for a promoter regulatory element and therefore unlikely to affect TGFbeta1 expression, whereas the Leu10Pro signal peptide substitution potentially affects TGFbeta1 secretion. Transfections of HeLa cells with constructs encoding either the Pro or Leu forms of TGFbeta1 and driven by the cytomegalovirus promoter indicate that the signal peptide with Pro at residue 10 causes a 2.8-fold increase in secretion compared with the Leu form. These data indicate that the allele encoding Pro10 is associated with increased rates of TGFbeta1 secretion and with increased incidence of invasive breast cancer for the population samples described. It is estimated that 3% of all breast cancer cases may be attributable to Pro10 homozygosity.