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: TP␣ and TP␤) is a GPCR displaying diverse pharmacological effects. As seen for many other GPCRs, TP␤ is regulated by agonistinduced 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 TP␤. This interaction was confirmed in a cellular context when Nm23-H2 was co-immunoprecipitated with TP␤ in HEK293 cells, a process dependent on agonist stimulation of the receptor. We observed that agonist-induced internalization of TP␤ 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 TP␤ 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.

Lang (2019) Cancer vaccines: designing artificial synthetic long peptides to improve presentation of class I and class II T cell epitopes by dendritic cells, OncoImmunology, 8:4, e1560919,