A SERINE PHOSPHORYLATION OF NM23, AND NOT ITS NUCLEOSIDE DIPHOSPHATE KINASE-ACTIVITY, CORRELATES WITH SUPPRESSION OF TUMOR METASTATIC POTENTIAL

We describe a serine phosphorylation of the putative metastasis suppre ssor protein Nm23, and present evidence of its relevance to the signal transduction and tumor metastatic processes. Nm23 was previously demo nstrated to exhibit nucleoside diphosphate kinase (NDPK) activity, whi ch transfers a phosphate among nucleoside tri- and diphosphates via an Nm23-phosphohistidine intermediate. Recent data have dissociated the NDPK activity of Nm23 from its phenotypic effects; therefore we have a sked whether Nm23 possesses additional biochemical functions. An acid- stable (nonhistidine) phosphorylation was identified on autophosphoryl ated purified recombinant Nm23 proteins and [P-32]orthophosphate-label ed human breast carcinoma and murine melanoma Nm23. Phosphoamino acid analysis identified serine as the acid-stable phosphorylation and seri ne 44 as the major site of phosphorylation. The acid stable phosphoryl ation (serine) of Nm23 was inhibited by cAMP in vitro and forskolin in vivo, suggesting that this phosphorylation pathway is regulated in si gnal transduction. No effect of cAMP was observed on Nm223 NDPK activi ty. Once phosphorylated, Nm23-phosphoserine can release free phosphate in vitro. The biological relevance of the novel phosphorylation ident ified herein is suggested by the direct correlation of in vivo Nm23 ac id-stable phosphorylation levels, but not Nm23 NDPK activity, with sup pression of tumor metastatic potential among control and nm23-1 transf ected murine melanoma cells.