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Proteomic identification of the cerebral cavernous malformation signaling complex.

Publication ,  Journal Article
Hilder, TL; Malone, MH; Bencharit, S; Colicelli, J; Haystead, TA; Johnson, GL; Wu, CC
Published in: J Proteome Res
November 2007

Cerebral cavernous malformations (CCM) are sporadic or inherited vascular lesions of the central nervous system characterized by dilated, thin-walled, leaky vessels. Linkage studies have mapped autosomal dominant mutations to three loci: ccm1 (KRIT1), ccm2 (OSM), and ccm3 (PDCD10). All three proteins appear to be scaffolds or adaptor proteins, as no enzymatic function can be attributed to them. Our previous results demonstrated that OSM is a scaffold for the assembly of the GTPase Rac and the MAPK kinase kinase MEKK3, for the hyperosmotic stress-dependent activation of p38 MAPK. Herein, we show that the three CCM proteins are members of a larger signaling complex. To define this complex, epitope-tagged wild type OSM or OSM harboring the mutation of F217-->A, which renders the OSM phosphotyrosine binding (PTB) domain unable to bind KRIT1, were stably introduced into RAW264.7 mouse macrophages. FLAG-OSM or FLAG-OSMF217A and the associated complex members were purified by immunoprecipitation using anti-FLAG antibody. OSM binding partners were identified by gel-based methods combined with electrospray ionization-MS or by multidimensional protein identification technology (MudPIT). Previously identified proteins that associate with OSM including KRIT1, MEKK3, Rac, and the KRIT1-binding protein ICAP-1 were found in the immunoprecipitates. In addition, we show for the first time that PDCD10 binds to OSM and is found in cellular CCM complexes. Other prominent proteins that bound the CCM complex include EF1A1, RIN2, and tubulin, with each interaction disrupted with the OSMF217A mutant protein. We further show that PDCD10 binds phosphatidylinositol di- and triphosphates and OSM binds phosphatidylinositol monophosphates. The findings define the targeting of the CCM complex to membranes and to proteins regulating trafficking and the cytoskeleton.

Duke Scholars

Published In

J Proteome Res

DOI

ISSN

1535-3893

Publication Date

November 2007

Volume

6

Issue

11

Start / End Page

4343 / 4355

Location

United States

Related Subject Headings

  • rac GTP-Binding Proteins
  • Spectrometry, Mass, Electrospray Ionization
  • Signal Transduction
  • Proto-Oncogene Proteins
  • Proteomics
  • Protein Structure, Tertiary
  • Phosphatidylinositols
  • Phosphates
  • Mice
  • Membrane Proteins
 

Citation

APA
Chicago
ICMJE
MLA
NLM
Hilder, T. L., Malone, M. H., Bencharit, S., Colicelli, J., Haystead, T. A., Johnson, G. L., & Wu, C. C. (2007). Proteomic identification of the cerebral cavernous malformation signaling complex. J Proteome Res, 6(11), 4343–4355. https://doi.org/10.1021/pr0704276
Hilder, Thomas L., Michael H. Malone, Sompop Bencharit, John Colicelli, Timothy A. Haystead, Gary L. Johnson, and Christine C. Wu. “Proteomic identification of the cerebral cavernous malformation signaling complex.J Proteome Res 6, no. 11 (November 2007): 4343–55. https://doi.org/10.1021/pr0704276.
Hilder TL, Malone MH, Bencharit S, Colicelli J, Haystead TA, Johnson GL, et al. Proteomic identification of the cerebral cavernous malformation signaling complex. J Proteome Res. 2007 Nov;6(11):4343–55.
Hilder, Thomas L., et al. “Proteomic identification of the cerebral cavernous malformation signaling complex.J Proteome Res, vol. 6, no. 11, Nov. 2007, pp. 4343–55. Pubmed, doi:10.1021/pr0704276.
Hilder TL, Malone MH, Bencharit S, Colicelli J, Haystead TA, Johnson GL, Wu CC. Proteomic identification of the cerebral cavernous malformation signaling complex. J Proteome Res. 2007 Nov;6(11):4343–4355.
Journal cover image

Published In

J Proteome Res

DOI

ISSN

1535-3893

Publication Date

November 2007

Volume

6

Issue

11

Start / End Page

4343 / 4355

Location

United States

Related Subject Headings

  • rac GTP-Binding Proteins
  • Spectrometry, Mass, Electrospray Ionization
  • Signal Transduction
  • Proto-Oncogene Proteins
  • Proteomics
  • Protein Structure, Tertiary
  • Phosphatidylinositols
  • Phosphates
  • Mice
  • Membrane Proteins