The NM23/NME gene was identified as a
metastasis suppressor. It's re-expression inhibited
cancer cell motility and suppressed
metastasis, without effecting primary
tumor size in multiple model systems. The mechanisms of NME suppression of motility and
metastasis are incompletely known. Of particular interest, has been NME
histidine 118 phosphorylation, involved in
nucleoside diphosphate kinase (NDPK) and
histidine protein kinase (HPK) activities. Using recently developed
monoclonal antibodies to
phosphohistidine, we have addressed the correlation of NME
phosphohistidine with motility suppression, and distinguished the NDPK and HPK contributions. While general levels of NME correlated with its
1-phosphohistidine form in two cell line model systems, two exceptions were noted:
Tumor cells actively migrating in scratch assays, even if expressing high levels of NME1, were low in its
1-phosphohistidine form. Site-directed mutagenesis of NME1
histidine 118 and
proline 96 was examined by transfection experiments and partial purification of
recombinant proteins. NME1P96S overexpressing
tumor cells exhibited high motility and migration phenotypes despite high
1-phosphohistidine content and NDPK activity; HPK activity using
succinate thiokinase as a substrate was poor. The data suggest the importance of NME
1-phosphohistidine levels in potential mechanistic pathways of
metastasis suppression and point toward the HPK activity of NME1 downstream of autophosphorylation.