A novel SMAD4 gene mutation in seminoma germ cell tumors.

Transforming growth factor (TGF)-beta is known as an antiproliferative factor in the majority of mammalian cells, including stem germ cells. Lack of TGF-beta-induced growth inhibition has been associated with disruptions of TGF-beta receptors and SMADs. In the present study, we performed a mutational analysis of the TGF-beta signaling system, including TGF-beta receptor type I and type II and SMADs (SMAD1-SMAD7), in 20 seminoma germ cell tumors. Using reverse transcription-PCR, single-strand conformational polymorphism, and sequencing analysis, the COOH-terminal domain of SMAD4 was found to be mutated: a single thymine was inserted between nt 1521 and 1522 in 2 of 20 tumors analyzed. This addition of a thymine creates a frameshift and a new stop signal at codon 492, which leads to premature termination of the encoded protein. Such a mutation potentially abrogates signaling from TGF-beta as well as the other TGF-beta family members, including activin and bone morphogenetic protein, which all use the SMAD pathway. Immunohistological analysis confirmed the loss of expression of SMAD4 protein in the seminoma tissues with the insertional mutation. To our knowledge, this is the first description of a novel SMAD4 insertional mutation in seminoma testicular germ cell tumors. This mutational inactivation of SMAD4/COOH-terminal domain may cause TGF-beta unresponsiveness. It could thus provide a basis for understanding the potential role of the TGF-beta system in germ cell tumorigenesis.