R-Spondin 4 (RSPO4, roof plate-specific spondin 4), also called cysteine-rich and single thrombospondin domain containing-4 (Cristin 4), is an ~33 kDa secreted heparin-binding protein that shares ~35% amino acid (aa) identity with three other R-Spondin family members. All are positive modulators of Wnt/beta-Catenin signaling, but vary in activity. R-Spondins regulate Wnt/beta-Catenin by competing with the Wnt antagonist DKK-1 for binding to the Wnt co-receptors LRP-6 and Kremen, reducing their DKK-1-mediated internalization. Like other R-spondins, human R-Spondin 4 (228 aa) contains a signal sequence (aa 1-19), two adjacent cysteine-rich furin-like domains (aa 85-128) with one potential tyrosine phosphorylation site (aa 114), followed by a thrombospondin (TSP-1) motif (aa 137-197) and a region rich in basic residues (aa 199-228). Mature human R-Spondin 4 shares 81%, 81%, 84%, 84% and 86% aa identity with mouse, rat, equine, canine and bovine R-Spondin 4, respectively. Of two potential isoforms, one lacks the TSP-1 domain, while another terminates at aa 224. Each R-Spondin has a distinct expression pattern. In the mouse, R-Spondin 4 mRNA is found during development of limb bud mesenchyme, nail beds, heart and teeth. In humans, mutations of R-Spondin 4 have been found to cause anonychia, a condition in which fingernails and toenails are absent.