Use of G-protein fusions to monitor integral membrane protein-protein interactions in yeast.
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.
Ehrhard
K N
KN
Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06536, USA.
Jacoby
J J
JJ
Fu
X Y
XY
Jahn
R
R
Dohlman
H G
HG
D048868:Adaptor Proteins, Signal Transducing; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D003593:Cytoplasm / Q000737:chemistry / Q000378:metabolism; D044388:GTP-Binding Protein gamma Subunits; D017930:Genes, Reporter / Q000235:genetics; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051938:Munc18 Proteins; D009154:Mutation / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D010675:Pheromones / Q000494:pharmacology; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases; D051498:Receptor, Fibroblast Growth Factor, Type 3; D017468:Receptors, Fibroblast Growth Factor / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000187:drug effects / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D015398:Signal Transduction / Q000187:drug effects; D012995:Solubility; D050827:Syntaxin 1; D020798:Two-Hybrid System Techniques; D033921:Vesicular Transport Proteins
D048868:Adaptor Proteins, Signal Transducing; D000954:Antigens, Surface; C106534:FRS2 protein, human; D044388:GTP-Binding Protein gamma Subunits; D008565:Membrane Proteins; D051938:Munc18 Proteins; D009419:Nerve Tissue Proteins; D010675:Pheromones; D010750:Phosphoproteins; D017468:Receptors, Fibroblast Growth Factor; D011993:Recombinant Fusion Proteins; C084294:STE18 protein, S cerevisiae; C494212:STX1A protein, human; D029701:Saccharomyces cerevisiae Proteins; D050827:Syntaxin 1; D033921:Vesicular Transport Proteins; D011505:Protein-Tyrosine Kinases; D051498:Receptor, Fibroblast Growth Factor, Type 3; D020962:Heterotrimeric GTP-Binding Proteins