Interaction of the Ras-related protein associated with diabetes Rad and the putative tumor metastasis suppressor NM23 provides a novel mechanism of GTPase regulation

Rad is the prototypic member of a new class of Ras-related GTPases, Purific ation of the GTPase-activating protein (CAP) for Rad revealed nm23, a putat ive tumor metastasis suppressor and a development gene in Drosophila. Antib odies against nm23 depleted Rad-CAP activity from human skeletal muscle cyt osol, and bacterially expressed nm23 reconstituted the activity. The GAP ac tivity of nm23 was specific for Rad, was absent with the S105N putative dom inant negative mutant of Rad, and was reduced with mutations of nm23, In th e presence of ATP, GDP Rad was also reconverted to CTP Rad by the nucleosid e diphosphate (NDP) kinase activity of nm23. Simultaneously, Rad regulated nm23 by enhancing its NDP kinase activity and decreasing its autophosphoryl ation. Melanoma cells transfected with wild-type Rad, but not the S105N-Rad , showed enhanced DNA synthesis in response to serum; this effect was lost with coexpression of nm23, Thus, the interaction of nm23 and Rad provides a potential novel mechanism for bidirectional, bimolecular regulation in whi ch nm23 stimulates both CTP hydrolysis and GTP loading of Rad whereas Rad r egulates activity of nm23. This interaction may play important roles in the effects of Rad on glucose metabolism and the effects of nm23 on tumor meta stasis and developmental regulation.