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SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes.

Publication ,  Journal Article
Okada, H; Uezu, A; Mason, FM; Soderblom, EJ; Moseley, MA; Soderling, SH
Published in: Sci Signal
November 29, 2011

Rho family GAPs [guanosine triphosphatase (GTPase) activating proteins] negatively regulate Rho family GTPase activity and therefore modulate signaling events that control cytoskeletal dynamics. The spatial distribution of these GAPs and their specificity toward individual GTPases are controlled by their interactions with various proteins within signaling complexes. These interactions are likely mediated through the Src homology 3 (SH3) domain, which is abundant in the Rho family GAP proteome and exhibits a micromolar binding affinity, enabling the Rho family GAPs to participate in transient interactions with multiple binding partners. To capture these elusive GAP signaling complexes in situ, we developed a domain-based proteomics approach, starting with in vivo phototrapping of SH3 domain-binding proteins and the mass spectrometry identification of associated proteins for nine representative Rho family GAPs. After the selection of candidate binding proteins by cluster analysis, we performed peptide array-based high-throughput in vitro binding assays to confirm the direct interactions and map the SH3 domain-binding sequences. We thereby identified 54 SH3-mediated binding interactions (including 51 previously unidentified ones) for nine Rho family GAPs. We constructed Rho family GAP interactomes that provided insight into the functions of these GAPs. We further characterized one of the predicted functions for the Rac-specific GAP WRP and identified a role for WRP in mediating clustering of the postsynaptic scaffolding protein gephyrin and the GABA(A) (γ-aminobutyric acid type A) receptor at inhibitory synapses.

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Published In

Sci Signal

DOI

EISSN

1937-9145

Publication Date

November 29, 2011

Volume

4

Issue

201

Start / End Page

rs13

Location

United States

Related Subject Headings

  • src Homology Domains
  • Signal Transduction
  • Receptors, GABA-A
  • Proteomics
  • Models, Biological
  • Humans
  • HEK293 Cells
  • GTPase-Activating Proteins
  • 3101 Biochemistry and cell biology
  • 0601 Biochemistry and Cell Biology
 

Citation

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Okada, H., Uezu, A., Mason, F. M., Soderblom, E. J., Moseley, M. A., & Soderling, S. H. (2011). SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes. Sci Signal, 4(201), rs13. https://doi.org/10.1126/scisignal.2002189
Okada, Hirokazu, Akiyoshi Uezu, Frank M. Mason, Erik J. Soderblom, M Arthur Moseley, and Scott H. Soderling. “SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes.Sci Signal 4, no. 201 (November 29, 2011): rs13. https://doi.org/10.1126/scisignal.2002189.
Okada H, Uezu A, Mason FM, Soderblom EJ, Moseley MA, Soderling SH. SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes. Sci Signal. 2011 Nov 29;4(201):rs13.
Okada, Hirokazu, et al. “SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes.Sci Signal, vol. 4, no. 201, Nov. 2011, p. rs13. Pubmed, doi:10.1126/scisignal.2002189.
Okada H, Uezu A, Mason FM, Soderblom EJ, Moseley MA, Soderling SH. SH3 domain-based phototrapping in living cells reveals Rho family GAP signaling complexes. Sci Signal. 2011 Nov 29;4(201):rs13.

Published In

Sci Signal

DOI

EISSN

1937-9145

Publication Date

November 29, 2011

Volume

4

Issue

201

Start / End Page

rs13

Location

United States

Related Subject Headings

  • src Homology Domains
  • Signal Transduction
  • Receptors, GABA-A
  • Proteomics
  • Models, Biological
  • Humans
  • HEK293 Cells
  • GTPase-Activating Proteins
  • 3101 Biochemistry and cell biology
  • 0601 Biochemistry and Cell Biology