Use of G-protein fusions to monitor integral membrane protein-protein interactions in yeast

Nat Biotechnol. 2000 Oct;18(10):1075-9. doi: 10.1038/80274.

Abstract

The control of protein-protein interactions is a fundamental aspect of cell regulation. Here we describe a new approach to detect the interaction of two proteins in vivo. By this method, one binding partner is an integral membrane protein whereas the other is soluble but fused to a G-protein gamma-subunit. If the binding partners interact, G-protein signaling is disrupted. We demonstrate interaction between known binding partners, syntaxin 1a with neuronal Sec1 (nSec1), and the fibroblast-derived growth factor receptor 3 (FGFR3) with SNT-1. In addition, we describe a genetic screen to identify nSec1 mutants that are expressed normally, but are no longer able to bind to syntaxin 1a. This provides a convenient method to study interactions of integral membrane proteins, a class of molecules that has been difficult to study by existing biochemical or genetic methods.

Publication types

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

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Antigens, Surface / genetics
  • Antigens, Surface / metabolism
  • Cell Membrane / chemistry
  • Cell Membrane / metabolism*
  • Cytoplasm / chemistry
  • Cytoplasm / metabolism
  • GTP-Binding Protein gamma Subunits*
  • Genes, Reporter / genetics
  • Heterotrimeric GTP-Binding Proteins / genetics*
  • Heterotrimeric GTP-Binding Proteins / metabolism*
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism*
  • Munc18 Proteins
  • Mutation / genetics
  • Nerve Tissue Proteins / genetics
  • Nerve Tissue Proteins / metabolism
  • Pheromones / pharmacology
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism
  • Protein Binding / drug effects
  • Protein Transport
  • Protein-Tyrosine Kinases*
  • Receptor, Fibroblast Growth Factor, Type 3
  • Receptors, Fibroblast Growth Factor / genetics
  • Receptors, Fibroblast Growth Factor / metabolism
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae / cytology
  • Saccharomyces cerevisiae / drug effects
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae Proteins*
  • Signal Transduction / drug effects
  • Solubility
  • Syntaxin 1
  • Two-Hybrid System Techniques*
  • Vesicular Transport Proteins*

Substances

  • Adaptor Proteins, Signal Transducing
  • Antigens, Surface
  • FRS2 protein, human
  • GTP-Binding Protein gamma Subunits
  • Membrane Proteins
  • Munc18 Proteins
  • Nerve Tissue Proteins
  • Pheromones
  • Phosphoproteins
  • Receptors, Fibroblast Growth Factor
  • Recombinant Fusion Proteins
  • STE18 protein, S cerevisiae
  • STX1A protein, human
  • Saccharomyces cerevisiae Proteins
  • Syntaxin 1
  • Vesicular Transport Proteins
  • Protein-Tyrosine Kinases
  • Receptor, Fibroblast Growth Factor, Type 3
  • Heterotrimeric GTP-Binding Proteins