We previously identified the novel gene, periodontal ligament-associated protein-1 (PLAP-1)/asporin and reported that PLAP-1/asporin inhibited bone morphogenetic protein-2 (BMP-2)-induced cytodifferentiation of periodontal ligament (PDL) cells probably by direct interaction with BMP-2. Here, we elucidated the detailed regulatory mechanism of this protein on BMP-2-induced cytodifferentiation of PDL cells. Recombinant PLAP-1 /asporin inhibited BMP-2-induced cytodifferentiation of PDL cells and competitively prevented BMP-2 from binding to the BMP receptor-IB (BMPR-IB), resulting in inhibition of BMP-dependent activation of Smad proteins. The induction of mutation to the leucine-rich repeat (LRR) motif, especially LRR5, within PLAP-1 /asporin rescued the inhibitory effect of PLAP-I /asporin on BMP-2. By contrast, a 26-amino acid peptide in the PLAP-1/asporin LRR5 sequence inhibited BMP-2 activity. Our findings indicate that PLAP-1 /asporin inhibits BMP-2-induced differentiation of PDL cells resulting from inactivation of the BMP-2 signaling pathway and that LRR, especially LRR5 of PLAP-1 /asporin, plays an important role in the PLAP-1/asporin-BMP-2 interaction. (C) 2008 Elsevier Inc. All rights reserved.