Abstract
Raine syndrome is caused by mutations in FAM20C, which had been reported to encode a secreted component of bone and teeth. We found that FAM20C encodes a Golgi-localized protein kinase, distantly related to the Golgi-localized kinase Four-jointed. Drosophila also encode a Golgi-localized protein kinase closely related to FAM20C. We show that FAM20C can phosphorylate secreted phosphoproteins, including both Casein and members of the SIBLING protein family, which modulate biomineralization, and we find that FAM20C phosphorylates a biologically active peptide at amino acids essential for inhibition of biomineralization. We also identify autophosphorylation of FAM20C, and characterize parameters of FAM20C’s kinase activity, including its Km, pH and cation dependence, and substrate specificity. The biochemical properties of FAM20C match those of an enzymatic activity known as Golgi casein kinase. Introduction of point mutations identified in Raine syndrome patients into recombinant FAM20C impairs its normal localization and kinase activity. Our results identify FAM20C as a kinase for secreted phosphoproteins and establish a biochemical basis for Raine syndrome.
Highlights
The main structural component of bone is a composite of secreted extracellular proteins and the mineral hydroxyapatite
When we compared the relative activity of FAM20C, Casein kinase 1 (CK1), and Casein Kinase 2 (CK2) on these peptides, we found that FAM20C exhibited a strong activity on the Golgi casein kinase (G-CK) peptide, and only very weak activity on the other peptides, further supporting its identification as a protein responsible for G-CK activity (Fig. 3a)
Our combined results indicate that FAM20C kinase activity is essential for limiting bone formation in humans, and identify Small Integrin-Binding Ligand Nlinked Glycoproteins (SIBLING) proteins as an important class of FAM20C substrates
Summary
The main structural component of bone is a composite of secreted extracellular proteins and the mineral hydroxyapatite. Insufficient bone density is a significant health concern for a majority of the human population as they age. Excess mineralization is implicated in pathological conditions. Human genetic diseases can identify proteins that modulate biomineralization. Raine syndrome (lethal osteosclerotic bone dysplasia) is associated with increased ossification resulting in skeletal malformation [1,2,3]. Raine syndrome is caused by mutations in FAM20C [4,5,6], which has been reported to encode a secreted component of bone and teeth [7,8]
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