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Quantitative kinetic analysis of BMP2 uptake into cells and its modulation by BMP-antagonists

Abstract:

Bone morphogenetic proteins (BMPs) are members of the TGFβ-family of signaling proteins and play an important role during development and in tissue formation. BMP signaling is a well studied process, which is initiated through binding of cognate receptors and processed via activation of Smad downstream mediators. A hallmark of BMP signaling is its modulation on the extracellular level through specific antagonists.Although it had been shown that BMP- and TGFβ-receptors are internalized following activation, little is known about the fate of BMP ligands. We prepared biologically active fluorescently labeled BMP2 and quantitatively analyzed its binding and uptake in cells using flow cytometry and confocal microscopy.Exogenous BMP2 was rapidly bound to the cell surface and subsequently internalized in a time dependent manner and accumulated in the cell center. While binding to the cell surface was limited by binding sites at the beginning, internalization continuously increased after a short delay at later time points. Using different inhibitors we found that internalization of BMP2 through endosomal particles occurred in a clathrin dependent pathway. Furthermore, uptake of BMP2 was modulated in strikingly different ways by BMP2 antagonists. While Noggin and Gremlin increased BMP2 uptake, Chordin blocked BMP2 uptake, which was concentration dependent in both cases.In conclusion, our findings present interesting mechanisms for the modulation of BMP signaling by concentration gradients of BMP ligands and antagonists in a dose and time dependent manner, which can provide an explanation of some properties of the BMP-regulatory network.

PubMed ID: 23077176

Projects: A3.2: Cross-talk of signaling pathways and endocytic machinery in hepato...

Journal: J. Cell. Sci.
Citation: J. Cell. Sci. 126(Pt 1): 117-27
Date Published: 19th Oct 2012

Authors: Hamed Alborzinia, Hannah Schmidt-Glenewinkel, Iryna Ilkavets, Katja Breitkopf-Heinlein, Xinlai Cheng, Peter Hortschansky, Steven Dooley, Stefan Wölfl

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