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Structural and functional studies of the Ras-associating and pleckstrin-homology domains of Grb10 and Grb14

Nature Structural & Molecular Biology volume 16, pages 833–839 (2009)Cite this article

An Erratum to this article was published on 01 December 2009

This article has been updated

Abstract

Growth factor receptor–binding proteins Grb7, Grb10 and Grb14 are adaptor proteins containing a Ras-associating (RA) domain, a pleckstrin-homology (PH) domain, a family-specific BPS (between PH and SH2) region and a C-terminal Src-homology-2 domain. Previous structural studies showed that the Grb14 BPS region binds as a pseudosubstrate inhibitor in the tyrosine kinase domain of the insulin receptor to suppress insulin signaling. Here we report the crystal structure of the RA and PH domains of Grb10 at 2.6-Å resolution. The structure reveals that these two domains, along with the intervening linker, form an integrated, dimeric structural unit. Biochemical studies demonstrated that Grb14 binds to activated Ras, which may serve as a timing mechanism for downregulation of insulin signaling. Our results illuminate the membrane-recruitment mechanisms not only of Grb7, Grb10 and Grb14 but also of MIG-10, Rap1-interacting adaptor molecule, lamellipodin and Pico, proteins involved in actin-cytoskeleton rearrangement that share a structurally related RA-PH tandem unit.

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Figure 1: Crystal structure of human Grb10 RA-PH.
Figure 2: Structure-based sequence alignments.
Figure 3: Binding of phosphoinositides to Grb10 and Grb14 RA-PH.
Figure 4: Interaction between Grb7, Grb10 and Grb14 and Ras.
Figure 5: Inhibition of insulin signaling by Grb14 and Grb10.
Figure 6: Model of the interaction between Grb14, Ras and the insulin receptor.

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Change history

  • 22 September 2009

    In the version of this article initially published, the Grp1 structure described in Figure 3c was incorrectly labeled Grb1. The error has been corrected in the HTML and PDF versions of the article.

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Acknowledgements

This work was supported by US National Institutes of Health grant DK052916 (to S.R.H.) We thank R. Ghirlando for sedimentation velocity measurements, M. Philips (New York University School of Medicine) for hemagglutinin-tagged Ras vectors and discussions, F. McCormick (University of California, San Francisco) for the GST-Ras and GST-Rap vectors, D. Lambright (University of Massachusetts Medical School) for the Grp1 PH domain construct, D. Ceccarelli for assistance in the fluorescence-polarization experiments, T. Miller for critical reading of the manuscript and N. Hiremath and J. Burrill for technical assistance. Beamline X4A at the National Synchrotron Light Source, Brookhaven National Laboratory, a Department of Energy facility, is supported by the New York Structural Biology Consortium.

Author information

Author notes

  1. Rafael S Depetris

    Present address: Present address: Department of Microbiology and Immunology, Weill Medical College of Cornell University, New York, New York, USA.,

  2. Rafael S Depetris and Jinhua Wu: These authors contributed equally to this work.

Authors and Affiliations

  1. and Department of Pharmacology, Structural Biology Program, Kimmel Center for Biology and Medicine of the Skirball Institute, New York University School of Medicine, New York, New York, USA

    Rafael S Depetris, Jinhua Wu & Stevan R Hubbard

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  1. Rafael S Depetris
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Contributions

R.S.D. performed crystallographic studies, in-cell biochemical experiments and manuscript preparation; J.W. performed structure refinement, fluorescence-polarization assays, in-cell biochemical experiments and manuscript preparation; S.R.H. supervised the project and was the principal manuscript author.

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Correspondence to Stevan R Hubbard.

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Depetris, R., Wu, J. & Hubbard, S. Structural and functional studies of the Ras-associating and pleckstrin-homology domains of Grb10 and Grb14. Nat Struct Mol Biol 16, 833–839 (2009). https://doi.org/10.1038/nsmb.1642

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