In vitro evidence for the recognition of 8-oxoGTP by Ras, a small GTP-binding protein.
Oxygen radicals attack guanine bases in DNA but they also attack cytoplasmic GTP forming 8-oxoGTP. The presence of 8-oxoGTP in cytoplasm is evidenced by the fact that cells contain MutT/MTH1 which hydrolyze 8-oxoGTP into 8-oxoGMP. In this study, the interaction between 8-oxoGTP and Ras, a small GTP-binding protein, was tested in vitro, and the action of 8-oxoGTP was compared to that of GTP. When purified Ras was treated with 8-oxoGTPgammaS, Ras was activated, as indicated by the enhanced binding of Ras with Raf-1. GTPgammaS also activated Ras but 8-oxoGTPgammaS had a much more potent effect. In lysates of human embryo kidney 293 cells, 8-oxoGTPgammaS activated not only Ras but also the downstream effectors of the Ras-ERK pathway, i.e., Raf-1 and ERK1/2. In contrast to Ras, other small GTP-binding proteins, Rac1 and Cdc42, were inactivated by 8-oxoGTPgammaS, whereas both of these proteins were activated by GTPgammaS, indicating that the biological natures of 8-oxoGTP and GTP differ. These results suggest the possibility that 8-oxoGTP is not a simple by-product but a functional molecule transmitting an oxidative signal to small GTP-binding proteins like Ras.
Yoon
Sun-Hee
SH
Department of Pharmacology, Seoul National University College of Medicine, 28 Yongon-dong, Chongno-gu, Seoul 110-799, Republic of Korea.
Hyun
Jin-Won
JW
Choi
Jinhee
J
Choi
Eun-Young
EY
Kim
Hee-Joon
HJ
Lee
Su-Jae
SJ
Chung
Myung-Hee
MH
D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology*; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D018631:ras Proteins / Q000378:metabolism*
C057674:8-hydroxyguanosine triphosphate; D006160:Guanosine Triphosphate; D019908:Proto-Oncogene Proteins c-raf; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; D018631:ras Proteins