Solution structure and biophysical characterization of the multifaceted signalling effector protein growth arrest specific-1

Background: The protein growth arrest specific-1 (GAS1) was discovered based on its ability to stop the cell cycle. During development it is involved in embryonic patterning, inhibits cell proliferation and mediates cell death, and has therefore been considered as a tumor suppressor. GAS1 is known to signal through two different cell membrane receptors: Rearranged during transformation (RET), and the sonic hedgehog receptor Patched-1. Sonic Hedgehog signalling is important in stem cell renewal and RET mediated signalling in neuronal survival. Disorders in both sonic hedgehog and RET signalling are connected to cancer progression. The neuroprotective effect of RET is controlled by glial cell-derived neurotrophic factor family ligands and glial cell-derived neurotrophic factor receptor alphas (GFR alpha s). Human Growth arrest specific-1 is a distant homolog of the GFRas. Results: We have produced and purified recombinant human GAS1 protein, and confirmed that GAS1 is a monomer in solution by static light scattering and small angle X-ray scattering analysis. The low resolution solution structure reveals that GAS1 is more elongated and flexible than the GFRas, and the homology modelling of the individual domains show that they differ from GFR alpha s by lacking the amino acids for neurotrophic factor binding. In addition, GAS1 has an extended loop in the N-terminal domain that is conserved in vertebrates after the divergence of fishes and amphibians. Conclusions: We conclude that GAS1 most likely differs from GFRas functionally, based on comparative structural analysis, while it is able to bind the extracellular part of RET in a neurotrophic factor independent manner, although with low affinity in solution. Our structural characterization indicates that GAS1 differs from GFR alpha's significantly also in its conformation, which probably reflects the functional differences between GAS1 and the GFR alpha s.