TY - JOUR
T1 - Toward a protein-protein interaction map of the budding yeast
T2 - A comprehensive system to examine two-hybrid interactions in all possible combinations between the yeast proteins
AU - Ito, Takashi
AU - Tashiro, Kosuke
AU - Muta, Shigeru
AU - Ozawa, Ritsuko
AU - Chiba, Tomoko
AU - Nishizawa, Mayumi
AU - Yamamoto, Kiyoshi
AU - Kuhara, Satoru
AU - Sakaki, Yoshiyuki
PY - 2000/2/1
Y1 - 2000/2/1
N2 - Protein-protein interactions play pivotal roles in various aspects of the structural and functional organization of the cell, and their complete description is indispensable to thorough understanding of the cell. As an approach toward this goal, here we report a comprehensive system to examine two-hybrid interactions in all of the possible combinations between proteins of Saccharomyces cerevisiae. We cloned all of the yeast ORFs individually as a DNA-binding domain fusion ('bait') in a MATa strain and as an activation domain fusion ('prey') in a MATα strain, and subsequently divided them into pools, each containing 96 clones. These bait and prey clone pools were systematically mated with each other, and the transformants were subjected to strict selection for the activation of three reporter genes followed by sequence tagging. Our initial examination of ≃4 x 106 different combinations, constituting ≃10% of the total to be tested, has revealed 183 independent two-hybrid interactions, more than half of which are entirely novel. Notably, the obtained binary data allow us to extract more complex interaction networks, including the one that may explain a currently unsolved mechanism for the connection between distinct steps of vesicular transport. The approach described here thus will provide many leads for integration of various cellular functions and serve as a major driving force in the completion of the protein-protein interaction map.
AB - Protein-protein interactions play pivotal roles in various aspects of the structural and functional organization of the cell, and their complete description is indispensable to thorough understanding of the cell. As an approach toward this goal, here we report a comprehensive system to examine two-hybrid interactions in all of the possible combinations between proteins of Saccharomyces cerevisiae. We cloned all of the yeast ORFs individually as a DNA-binding domain fusion ('bait') in a MATa strain and as an activation domain fusion ('prey') in a MATα strain, and subsequently divided them into pools, each containing 96 clones. These bait and prey clone pools were systematically mated with each other, and the transformants were subjected to strict selection for the activation of three reporter genes followed by sequence tagging. Our initial examination of ≃4 x 106 different combinations, constituting ≃10% of the total to be tested, has revealed 183 independent two-hybrid interactions, more than half of which are entirely novel. Notably, the obtained binary data allow us to extract more complex interaction networks, including the one that may explain a currently unsolved mechanism for the connection between distinct steps of vesicular transport. The approach described here thus will provide many leads for integration of various cellular functions and serve as a major driving force in the completion of the protein-protein interaction map.
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U2 - 10.1073/pnas.97.3.1143
DO - 10.1073/pnas.97.3.1143
M3 - Article
C2 - 10655498
AN - SCOPUS:0033974688
VL - 97
SP - 1143
EP - 1147
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 3
ER -