Genetics and Genomics of SOST: Functional Analysis of Variants and Genomic Regulation in Osteoblasts.

Núria Martínez-Gil,Daniel Grinberg,Mónica Cozar,Neus Roca-Ayats,Susanna Balcells,Natàlia Garcia-Giralt,Xavier Nogués,Diana Ovejero

doi:10.3390/ijms22020489

Abstract

SOST encodes the sclerostin protein, which acts as a key extracellular inhibitor of the canonical Wnt pathway in bone, playing a crucial role in skeletal development and bone homeostasis. The objective of this work was to assess the functionality of two variants previously identified (the rare variant rs570754792 and the missense variant p.Val10Ile) and to investigate the physical interactors of the SOST proximal promoter region in bone cells. Through a promoter luciferase reporter assay we show that the minor allele of rs570754792, a variant located in the extended TATA box motif, displays a significant decrease in promoter activity. Likewise, through western blot studies of extracellular and intracellular sclerostin, we observe a reduced expression of the p.Val10Ile mutant protein. Finally, using a circular chromosome conformation capture assay (4C-seq) in 3 bone cell types (MSC, hFOB, Saos-2), we have detected physical interactions between the SOST proximal promoter and the ECR5 enhancer, several additional enhancers located between EVT4 and MEOX1 and a distant region containing exon 18 of DHX8. In conclusion, SOST presents functional regulatory and missense variants that affect its expression and displays physical contacts with far reaching genomic sequences, which may play a role in its regulation within bone cells.

Highlights

Sclerostin is a potent antagonist of the canonical Wnt pathway and of bone formation, exerting its function by blocking LRP5/6 co-receptors [1,2]
SOST mutations are associated with inherited high bone mass conditions characterized by excessive bone formation, with a sclerosis phenotype prominent in the skull, jaw and long bones
The clinical spectrum of these conditions ranges from severe craniodiaphyseal dysplasia to non-pathological high bone mass, with the severity of the condition being inversely proportional to sclerostin abundance

Summary

Introduction

Sclerostin (encoded by the SOST gene) is a potent antagonist of the canonical Wnt pathway and of bone formation, exerting its function by blocking LRP5/6 co-receptors [1,2]. To perform this function, sclerostin binds to LRP4, which enhances its suppressive effect [3,4,5]. Craniodiaphyseal dysplasia (CDD, MIM 122860) is the most severe sclerostin bone dysplasia, characterized by an absence of extracellular sclerostin and a dominant negative effect It is an extremely rare autosomal dominant condition, with less than 20 cases reported [9,10].

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