Endofin acts as a Smad anchor for receptor activation in BMP signaling

Weibin Shi,Xu Cao,Shuyi Nie,Mei Wan,Shutao Xie,Chenbei Chang

doi:10.1242/jcs.03400

Abstract

Signaling through receptors of the transforming growth factor beta (TGFbeta) superfamily is mediated by cytoplasmic Smad proteins. It has been demonstrated that Smad anchor for receptor activation (SARA) facilitates TGFbeta and activin/nodal signaling by recruiting and presenting Smad2/3 to the receptor complex. SARA does not bind Smad1 and hence does not enhance bone morphogenetic protein (BMP) signaling. Here we report for the first time that the endosome-associated FYVE-domain protein endofin acts as a Smad anchor for receptor activation in BMP signaling. We demonstrate that endofin binds Smad1 preferentially and enhances Smad1 phosphorylation and nuclear localization upon BMP stimulation. Silencing of endofin by RNAi resulted in a reduction in BMP-dependent Smad1 phosphorylation. Moreover, disruption of the membrane-anchoring FYVE motif by point mutation led to a reduction of BMP-responsive gene expression in cell culture and Xenopus ectodermal explants. Furthermore, we demonstrate that endofin contains a protein-phosphatase-binding motif, which functions to negatively modulate BMP signals through receptor dephosphorylation. Taken together, our results suggest that endofin plays an important role in both positive and negative feedback regulation of the BMP signaling pathway.

Highlights

Growth factors of the TGF␤ superfamily regulate cell proliferation, differentiation, migration and apoptosis and are crucial for the development and maintenance of many different tissues (Cheifetz, 1999; Francis-West et al, 1999)
Endofin interacts with Smad1 and PP1c Endofin has been shown to localize to early endosomes, as does Smad anchor for receptor activation (SARA); but it does not bind to Smad2 or influence TGF␤ signals (Seet and Hong, 2001)
The relatively high divergence of the Smad-binding domain suggested that endofin might bind R-Smads other than the TGF␤-specific Smad-2, such as the bone morphogenetic protein (BMP)-specific Smad1

Summary

Introduction

Growth factors of the TGF␤ superfamily regulate cell proliferation, differentiation, migration and apoptosis and are crucial for the development and maintenance of many different tissues (Cheifetz, 1999; Francis-West et al, 1999). Bone morphogenetic proteins (BMPs) in this superfamily induce bone marrow mesenchymal stem cell differentiation into osteoblasts and enhance periodontal regeneration in surgically created defects. BMP ligands, as well as other ligands of the TGF␤ superfamily, initiate cellular signaling by binding to type I and type II receptors, both of which are serine/threonine kinase receptors, thereby inducing the formation of a heterooligomeric receptor complex. The type II receptor phosphorylates and activates the type I receptor, which subsequently transiently associates with and phosphorylates a subclass of a unique family of intracellular signaling molecules – the receptor-regulated Smads or R-Smads (Godin et al, 1999; Ghosh-Choudhury et al, 1994). The recruitment of R-Smads to the receptor complex is the initial step of intracellular TGF␤ signal transduction and plays a critical role in regulating signal transduction by the TGF␤ receptor family

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