SPAG11C immunoreactivity is exclusively found in the Wolffian duct mesenchyme. Androgen receptor signaling regulates SPAG11C expression in developing Wolffian duct. Recombinant hSPAG11C protein affects Wolffian duct morphogenesis ex vivo. The hSPAG11C mechanism relies on regulation of ductal epithelial cell proliferation. The β-defensin SPAG11C is suggested as a new player in Wolffian duct morphogenesis.