Catalysis-Independent ENPP1 Protein Signaling Regulates Mammalian Bone Mass

Area of Science:

• Biochemistry
• Molecular Biology
• Skeletal Biology

Background:

• Biallelic ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) deficiency causes generalized arterial calcification of infancy (GACI) and hypophosphatemic rickets.
• ENPP1 deficiency presents with reduced skeletal mineral accrual and excess heterotopic mineralization, a paradox not explained by its role in generating pyrophosphate (PPi).
• ENPP1 generates extracellular pyrophosphate (PPi), an inhibitor of mineralization, and its deficiency leads to vascular calcification due to low plasma PPi.

Purpose of the Study:

• To investigate the catalysis-independent pathways of ENPP1 that regulate bone mass.
• To differentiate the roles of ENPP1's catalytic activity versus signaling in phosphate homeostasis and bone mineralization.
• To develop a murine model that uncouples ENPP1 protein signaling from its catalytic function.

Main Methods:

• Developed a murine model (Enpp1^T238A) with uncoupled ENPP1 signaling and catalysis.
• Compared Enpp1^T238A mice with Enpp1^asj mice (lacking ENPP1) and wild-type (WT) controls.
• Analyzed bone microarchitecture, biomechanics, plasma phosphate (Pi) and PPi levels, fibroblast growth factor 23 (FGF23), and calvarial cell cultures.

Main Results:

• Enpp1^T238A mice exhibited normal bone structure, unlike Enpp1^asj mice.
• Both Enpp1^T238A and Enpp1^asj mice showed low plasma Pi and PPi, elevated FGF23, and osteomalacia, indicating equivalent phosphate wasting.
• Calvarial cells from Enpp1^asj mice showed reduced calcification, increased Sfrp1, and decreased β-catenin signaling, which were restored by Sfrp1 knockout.

Conclusions:

• Catalysis-independent ENPP1 signaling regulates bone mass through pathways involving secreted frizzled-related protein 1 (SFRP1), a Wnt inhibitor.
• Catalytic activity of ENPP1 regulates phosphate homeostasis via plasma FGF23 levels.
• ENPP1 has distinct roles in bone health, with separate mechanisms controlling mineralization and phosphate balance.