Adaptation of the Ras-recruitment system to the analysis of interactions between membrane-associated proteins.
Interactions of membrane-associated proteins play important roles in many cellular processes. The yeast two-hybrid assay is of limited utility for the analysis of such interactions, due to the need for soluble protein partners, whose interaction is assessed in the nucleus. The advent of the Ras-recruitment system (RRS) has enabled the study of membrane-associated proteins interacting with cytoplasmic proteins fused to Ras. Constitutive membrane association of the Ras fusion protein is expected to complement the growth defect of the yeast strain CDC25-2, assayed in the RRS, independent from the interaction with a membrane-bound partner. We describe the adaptation of the RRS to the analysis of interactions between two membrane-associated proteins using a model system. These results may facilitate the study of protein-protein interactions between membrane-bound proteins and further increase the utility of the RRS.
Köhler
Fabian
F
Institut für Biologie III, Albert-Ludwigs-Universität, Schänzlestrasse 1, 79104 Freiburg, Germany. fabian.koehler@biologie.uni-freiburg.de
Müller
Kristian M
KM
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D018797:Cell Cycle Proteins / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics; D051380:GRB2 Adaptor Protein; D005816:Genetic Complementation Test / Q000379:methods; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008954:Models, Biological; D009154:Mutation; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins; D013696:Temperature; D020798:Two-Hybrid System Techniques; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*; D020666:ras-GRF1 / Q000235:genetics
D048868:Adaptor Proteins, Signal Transducing; C542351:CDC25 protein, S cerevisiae; D018797:Cell Cycle Proteins; D005656:Fungal Proteins; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C495949:Grb2 protein, mouse; D008565:Membrane Proteins; D011506:Proteins; D029701:Saccharomyces cerevisiae Proteins; D020666:ras-GRF1; D018631:ras Proteins