Characterization of the different oligomeric states of the DAN family antagonists SOSTDC1 and SOST.

Gregory R Gipson,Thomas B Thompson,Magdalena Czepnik,Chandramohan Kattamuri

doi:10.1042/bcj20200552

Gregory R Gipson, Thomas B Thompson + Show 2 more

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https://doi.org/10.1042/bcj20200552

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Journal: Biochemical Journal	Publication Date: Sep 4, 2020
Citations: 9	License type: CC BY 4.0

Affiliation: University of Cincinnati

Abstract

The DAN (differential screening-selected gene aberrative in neuroblastoma) family are a group of secreted extracellular proteins which typically bind to and antagonize BMP (bone morphogenetic protein) ligands. Previous studies have revealed discrepancies between the oligomerization state of certain DAN family members, with SOST (a poor antagonist of BMP signaling) forming a monomer while Grem1, Grem2, and NBL1 (more potent BMP antagonists) form non-disulfide linked dimers. The protein SOSTDC1 (Sclerostin domain containing protein 1) is sequentially similar to SOST, but has been shown to be a better BMP inhibitor. In order to determine the oligomerization state of SOSTDC1 and determine what effect dimerization might have on the mechanism of DAN family antagonism of BMP signaling, we isolated the SOSTDC1 protein and, using a battery of biophysical, biochemical, and structural techniques, showed that SOSTDC1 forms a highly stable non-covalent dimer. Additionally, this SOSTDC1 dimer was shown, using an in vitro cell based assay system, to be an inhibitor of multiple BMP signaling growth factors, including GDF5, while monomeric SOST was a very poor antagonist. These results demonstrate that SOSTDC1 is distinct from paralogue SOST in terms of both oligomerization and strength of BMP inhibition.

Highlights

Bone morphogenetic proteins (BMPs) are the largest subclass of the transforming growth factor-β (TGF-β) family of extracellular signaling proteins, containing ∼15 of the 33 family members identified in mammals [1,2,3,4]
To reconcile the cross-linking data that SOSTDC1 exists as a dimer with the computational evidence that SOSTDC1 lacks the molecular features of other differential screening selected gene aberrative in neuroblastoma (DAN) family dimers required for dimerization, this study characterized the oligomeric state of recombinant SOSTDC1 using a number of orthogonal biophysical approaches and compared these results to purified SOST
The DAN family consists of seven extracellular proteins, generally categorized as inhibitors of BMP signaling

Summary

Introduction

Bone morphogenetic proteins (BMPs) are the largest subclass of the transforming growth factor-β (TGF-β) family of extracellular signaling proteins, containing ∼15 of the 33 family members identified in mammals [1,2,3,4]. Phosphorylation of the BMP-specific intracellular Smads (Smad-1, -5, and -9) allows them to bind the partnering co-Smad, Smad-4 [4,15,16,17,19,20,21] This Smad complex associates with other DNA-binding proteins and accumulates in the nucleus to induce transcriptional activation of target genes [4,15,16,17,18,21,22,23]. While differing greatly in terms of structure, these antagonists generally function by directly binding to BMP proteins blocking the receptor binding sites, to prevent the formation of the signaling complexes. The most common BMP antagonists are noggin, chordin, follistatin, and members of the DAN family [26,27,28,29,30,31]

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