We have identified a new member of the transforming growth factor-beta (TGF-beta) superfamily, growth/differentiation factor-10 (GDF-10), which is highly related to bone morphogenetic protein-3 (BMP-3). The nucleotide sequence of GDF-10 encodes a predicted protein of 476 amino acids with a molecular weight of approximately 52,000. The GDF-10 polypeptide contains a potential signal sequence for secretion, a putative RXXR proteolytic processing site, and a carboxy-terminal domain with considerable homology to other known members of the TGF-beta superfamily. In the mature carboxy-terminal domain GDF-10 is more homologous to BMP-3 (83% amino acid sequence identity) than to any other previously identified TGF-beta family member. GDF-10 also shows significant homology to BMP-3 (approximately 30% amino acid sequence identity) in the pro- region of the molecule. Based on these sequence comparisons, GDF-10 and BMP-3 define a new subgroup within the larger TGF-beta superfamily. By Northern analysis, GDF-10 mRNA was detected primarily in murine uterus, adipose tissue, and brain and to a lesser extent in liver and spleen. In addition, GDF-10 mRNA was present in both neonatal and adult bone samples, with higher levels being detected in calvaria than in long bone. These results suggest that GDF10 may play multiple roles in regulating cell differentiation events, including those involved in skeletal morphogenesis. Gdf10 was mapped to the proximal region of mouse chromosome 14 close to a region known to contain a spontaneous recessive mutation that is associated with a craniofacial defect.