Regulating developmental signaling by sorting at the plasma membrane

Dr. Giorgos Pyrowolakis (BIOSS and Institute of Biology I, University of Freiburg)

Signalling requires a high degree of regulation for proper tissue development and homeostasis. This project addresses the molecular mechanisms that regulate BMP/TGFß signalling at the level of the plasma membrane. Our previous genetic data on graded BMP signalling in a Drosophila epithelium identified key players and interactions that globally adjust the levels of signaling across a cellular field and critically influence the profile of the activity gradient. We have identified, Pentagone (Pent), an evolutionarily conserved extracellular protein that affects signalling activity through interactions with glypicans, GPI-anchored heparin-sulfate proteoglycans that bind ligands and act as co-receptors in BMP signal transduction. We could show that binding of Pent induces internalization and membrane-depletion of glypicans through endocytosis and lysosomal degradation. Our data suggest that Pent “compartmentalizes” glypican activity at the plasma membrane to regulate signaling: Pent shunts a fraction of the glypican pool into endocytosis, thus making them unavailable for productive interactions with the ligand/receptor complex.

In this project we wish to define the molecular events at - and proximal to - the plasma membrane by which Pent divert glypicans from signaling to endocytosis. To this end, and in collaboration with the Römer group (BIOSS), we will seek to re-build Pent-induced endocytosis of glypicans in heterologous cell culture systems that will allow following the process by high-end microscopy. We will address whether and how Pent binding induces redistribution (such as clustering) of glypicans. Using such systems, we will address the requirements of GPI-anchoring of glypicans in the process, as well as the endocytic components and pathways involved in co-receptor endocytosis. Findings of the cell culture studies will be confirmed and complemented by in vivo genetic studies to address the impact of identified components and endocytic routes in BMP gradient formation and in, ultimately, in tissue and organ development.