Abstract
UNC119 is widely expressed among vertebrates and other phyla. We found that UNC119 recognized the acylated N terminus of the rod photoreceptor transducin α (Tα) subunit and Caenorhabditis elegans G proteins ODR-3 and GPA-13. The crystal structure of human UNC119 at 1.95-Å resolution revealed an immunoglobulin-like β-sandwich fold. Pulldowns and isothermal titration calorimetry revealed a tight interaction between UNC119 and acylated Gα peptides. The structure of co-crystals of UNC119 with an acylated Tα N-terminal peptide at 2.0 Å revealed that the lipid chain is buried deeply into UNC119′s hydrophobic cavity. UNC119 bound Tα-GTP, inhibiting its GTPase activity, thereby providing a stable UNC119–Tα-GTP complex capable of diffusing from the inner segment back to the outer segment after light-induced translocation. UNC119 deletion in both mouse and C. elegans led to G protein mislocalization. Thus, UNC119 is a Gα subunit cofactor essential for G protein trafficking in sensory cilia.
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Acknowledgements
We thank R. Abramowitz and J. Schwanof for access to the X4A beamline at the National Synchrotron Light Source and G. DeTitta of Hauptman Woodward Research Institute for crystallization screening. This work was supported by US National Institutes of Health grants EY08123 (W.B.), EY019298 (W.B.), EY014800-039003 (National Eye Institute core grant), EY10848 (G.I.) and NS034307 (E.M.J.), by the Howard Hughes Medical Institute (E.M.J.), by a grant from the Protein Structure Initiative of the US National Institutes of Health (U54 GM074958), by the University of Utah Macromolecule Crystallography Core Facility, by a Center Grant of the Foundation Fighting Blindness to the University of Utah, and unrestricted grants to the Departments of Ophthalmology at the University of Utah from Research to Prevent Blindness. W.B. is a recipient of a Research to Prevent Blindness Senior Investigator Award.
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H.Z. generated pulldown/light-induced translocation results and is responsible for C. elegans immunostaining and imaging. R.C. generated ITC results. R.C., F.G.W. and C.P.H. generated human UNC119/acylated Tα-peptide co-crystals and determined the structure. S.V., Y.C., J.S., Y.J.H., R.X., G.T.M. and L.T. determined the unbound human UNC119 structure. R.C., C.D.G. and W.B. isolated ROS membranes, transducin and determined GTPase activity. M.W.D. and E.M.J. generated transgenic C. elegans mutants. G.I. generated the Unc119 knockout mouse. H.Z., R.C., C.P.H., L.T. and W.B. wrote the manuscript.
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Zhang, H., Constantine, R., Vorobiev, S. et al. UNC119 is required for G protein trafficking in sensory neurons. Nat Neurosci 14, 874–880 (2011). https://doi.org/10.1038/nn.2835
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DOI: https://doi.org/10.1038/nn.2835
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