Reconstitution of morphogen shuttling circuits.

Developing tissues form spatial patterns by establishing concentration gradients of diffusible signaling proteins called morphogens. The bone morphogenetic protein (BMP) morphogen pathway uses a family of extracellular modulators to reshape signaling gradients by actively "shuttling" ligands to different locations. It has remained unclear what circuits are sufficient to enable shuttling, what other patterns they can generate, and whether shuttling is evolutionarily conserved. Here, using a synthetic, bottom-up approach, we compared the spatiotemporal dynamics of different extracellular circuits. Three proteins-Chordin, Twsg, and the BMP-1 protease-successfully displaced gradients by shuttling ligands away from the site of production. A mathematical model explained the different spatial dynamics of this and other circuits. Last, combining mammalian and Drosophila components in the same system suggests that shuttling is a conserved capability. Together, these results reveal principles through which extracellular circuits control the spatiotemporal dynamics of morphogen signaling.

Gγ and Gα Identity Dictate a G-Protein Heterotrimer Plasma Membrane Targeting.

Heterotrimeric G-proteins along with G-protein-coupled receptors (GPCRs) regulate many biochemical functions by relaying the information from the plasma membrane to the inside of the cell. The lipid modifications of Gα and Gγ subunits, together with the charged regions on the membrane interaction surface, provide a peculiar pattern for various heterotrimeric complexes. In a previous study, we found that Gαs and Gαi3 prefer different types of membrane-anchor and subclass-specific lipid domains. In the present report, we examine the role of distinct Gγ subunits in the membrane localization and spatiotemporal dynamics of Gαs and Gαi3 heterotrimers. We characterized lateral diffusion and G-protein subunit interactions in living cells using fluorescence recovery after photobleaching (FRAP) microscopy and fluorescence resonance energy transfer (FRET) detected by fluorescence lifetime imaging microscopy (FLIM), respectively. The interaction of Gγ subunits with specific lipids was confirmed, and thus the modulation of heterotrimeric G-protein localization. However, the Gα subunit also modulates trimer localization, and so the membrane distribution of heterotrimeric G-proteins is not dependent on Gγ only.