Nm23-H2 interacts with a G protein-coupled receptor to regulate its endocytosis through an Rac1-dependent mechanism

G protein-coupled receptors (GPCRs) represent a vast family of transmembrane proteins involved in the regulation of several physiological responses. The thromboxane A2 receptor (present as two isoforms: TPalpha and TPbeta) is a GPCR displaying diverse pharmacological effects. As seen for many other GPCRs, TPbeta is regulated by agonist-induced internalization. In the present study, we report the identification by yeast two-hybrid screening of Nm23-H2, a nucleoside diphosphate kinase, as a new interacting molecular partner with the C-terminal tail of TPbeta. This interaction was confirmed in a cellular context when Nm23-H2 was co-immunoprecipitated with TPbeta in HEK293 cells, a process dependent on agonist stimulation of the receptor. We observed that agonist-induced internalization of TPbeta was regulated by Nm23-H2 through modulation of Rac1 signaling. Immunofluorescence microscopy in HEK293 cells revealed that Nm23-H2 had a cytoplasmic and nuclear localization but was induced to translocate to the plasma membrane upon stimulation of TPbeta to show extensive co-localization with the receptor. Our findings represent the first demonstration of an interaction of an Nm23 protein with a membrane receptor and constitute a novel molecular regulatory mechanism of GPCR endocytosis.