Influence of an LRP5 cytoplasmic SNP on Wnt signaling and osteoblastic differentiation

The low density lipoprotein receptor-related protein 5 (LRP5) is a key determinant of bone mass, via the Wnt signaling pathway control of osteoblast function. This study examined human LRP5 signaling and the effects of an intracellular domain single nucleotide polymorphism (SNP: p.V1525A) on osteoblast differentiation and mineralization. Constitutively active LRP5 was constructed by deletion of the extracellular domain of LRP5 (LRP5ΔN). Expression of LRP5ΔN-V, which carries the allele p.1525V, induced higher β-catenin/TCF-LEF activity compared to LRP5ΔN-A, which carries the allele p.1525A. In a yeast two-hybrid assay, LRP5ΔN-V also demonstrated a stronger interaction with AXIN than LRP5ΔN-A. Expression of either of the alleles did not change cell proliferation. However, cells expressing LRP5ΔN-V showed increased alkaline phosphatase activity and bone nodule formation compared to cells transfected with empty vector or LRP5ΔN-A after osteogenic supplement (OS: β-glycerophosphate and l-ascorbic acid) treatment. Cells expressing LRP5ΔN-V revealed significantly increased bone sialoprotein (BSP) expression after 7 days of OS treatment and maintained elevated expression until day 21. Osteocalcin (OCN) mRNA levels were increased after 14–21 days of OS treatment in LRP5ΔN-V expressing cells. LRP5ΔN-V expressing cells demonstrated positive interaction with BMP-2 signaling of transcription at the SBE-luc promoter. LRP5 signaling is affected by the cytoplasmic SNP, p.V1525A. mRNA levels of Runx2 and Osterix were not affected by this SNP.