A protein interaction network links GIT1, an enhancer of huntingtin aggregation, to Huntington's disease

Mol Cell. 2004 Sep 24;15(6):853-65. doi: 10.1016/j.molcel.2004.09.016.

Abstract

Analysis of protein-protein interactions (PPIs) is a valuable approach for characterizing proteins of unknown function. Here, we have developed a strategy combining library and matrix yeast two-hybrid screens to generate a highly connected PPI network for Huntington's disease (HD). The network contains 186 PPIs among 35 bait and 51 prey proteins. It revealed 165 new potential interactions, 32 of which were confirmed by independent binding experiments. The network also permitted the functional annotation of 16 uncharacterized proteins and facilitated the discovery of GIT1, a G protein-coupled receptor kinase-interacting protein, which enhances huntingtin aggregation by recruitment of the protein into membranous vesicles. Coimmunoprecipitations and immunofluorescence studies revealed that GIT1 and huntingtin associate in mammalian cells under physiological conditions. Moreover, GIT1 localizes to neuronal inclusions, and is selectively cleaved in HD brains, indicating that its distribution and function is altered during disease pathogenesis.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Amino Acid Sequence
  • Animals
  • Antibodies, Monoclonal / metabolism
  • Binding Sites
  • COS Cells
  • Cell Cycle Proteins / chemistry
  • Cell Cycle Proteins / metabolism*
  • Chlorocebus aethiops
  • GTPase-Activating Proteins / chemistry
  • GTPase-Activating Proteins / metabolism*
  • Glutathione / metabolism
  • Humans
  • Huntingtin Protein
  • Huntington Disease / metabolism*
  • Huntington Disease / pathology
  • Mice
  • Mice, Transgenic
  • Nerve Tissue Proteins / metabolism*
  • Nuclear Proteins / metabolism*
  • PC12 Cells
  • Phosphoproteins / chemistry
  • Phosphoproteins / metabolism*
  • Precipitin Tests
  • Proline / chemistry
  • Protein Binding
  • Protein Structure, Tertiary
  • RNA Interference
  • Rats
  • Recombinant Fusion Proteins / metabolism
  • Tissue Distribution
  • Two-Hybrid System Techniques

Substances

  • Adaptor Proteins, Signal Transducing
  • Antibodies, Monoclonal
  • Cell Cycle Proteins
  • GIT1 protein, human
  • GTPase-Activating Proteins
  • Git1 protein, mouse
  • Git1 protein, rat
  • HTT protein, human
  • Htt protein, mouse
  • Htt protein, rat
  • Huntingtin Protein
  • Nerve Tissue Proteins
  • Nuclear Proteins
  • Phosphoproteins
  • Recombinant Fusion Proteins
  • Proline
  • Glutathione