The Smad3 gene is a member of the Smad family, vertebrate homologues of Drosophila Mad, and its gene product is a cytoplasmic element in the TGF-beta signaling pathway. Smad2 and Smad4/DPC4, other members of the Smad family, are possibly tumor suppressor genes because alterations of these genes occurred in various carcinomas. We determined the genomic structure of human Smad3 which consists of nine exons. Then we examined whether or not Smad3 gene mutations exist in sporadic and hereditary non-polyposis colorectal cancers and found no mutations in the entire coding region in 50 cancers. Loss of heterozygosity of Smad3 was observed in two of the 17 (11.8%) informative cases using a polymorphism found in intron 2. These findings suggest that the Smad3 gene may not play an important role in the tumorigenesis of colorectal cancers.