A putative pH-dependent nuclear localization signal in the juxtamembrane region of c-Met

Abstract

The C-terminal fragment of the c-Met receptor tyrosine kinase is present in the nuclei of some cells irrespective of ligand stimulation, but the responsible nuclear localization signal (NLS) has not been previously reported. Here, we report that two histidine residues separated by a 10-amino-acid spacer (H1068–H1079) located in the juxtamembrane region of c-Met function as a putative novel NLS. Deletion of these sequences significantly abolished the nuclear translocation of c-Met, as did substitution of the histidines with alanines. This substitution also decreased the association of c-Met fragment with importin β. The putative NLS of c-Met is unique in that it relies on histidines, whose positive charge changes depending on pH, rather than the lysines or arginines, commonly found in classical bipartite NLSs, suggesting the possible ‘pH-dependency’ of this NLS. Indeed, decreasing the cytosolic pH either with nigericin, an Na+/H+ exchanger or pH 6.5 KRB buffer significantly increased the level of nuclear c-Met and the interaction of the c-Met fragment with importin β, indicating that low pH itself enhanced nuclear translocation. Consistent with this, nigericin treatment also increased the nuclear level of endogenous c-Met in HeLa cells. The putative aberrant bipartite NLS of c-Met seems to be the first example of what we call a ‘pH-dependent’ NLS. An unconventional mechanism for regulating protein localization within the cell may give tumors an advantage. The c-Met protein primarily acts at the cell membrane, transmitting signals that promote proliferation, and many cancers exhibit overactive c-Met. One fragment of c-Met enters the nucleus, however, where it directly controls growth-related genes, and researchers led by Jae-Ho Lee at South Korea's Ajou University have uncovered how this migration occurs. Most nuclear proteins contain a ‘nuclear localization signal’ (NLS), which is enriched with positively-charged amino acids. c-Met lacks such an NLS but contains a novel localization sequence containing two histidines, an amino acid that is only positively charged in acidic conditions. The researchers demonstrate that c-Met transport requires this acidic environment—commonly found in cancer cells—which may explain this protein's role in tumor growth.