Hyperactivation of mitogen-activated protein kinase increases phospho-tau immunoreactivity within human neuroblastoma: additive and synergistic influence of alteration of additional kinase activities

Abstract

Mitogen-activated protein (MAP) kinase phosphorylates tau in cell-free analyses, but whether or not it does so within intact cells remains controversial. In the present study, microinjection of MAP kinase into SH-SY-5Y human neuroblastoma cells increased tau immunoreactivity toward the phosphodependent antibodies PHF-1 and AT-8. In contrast, treatment with a specific inhibitor of MAP kinase (PD98059) did not diminish "basal" levels of these immunoreactivities in otherwise untreated cells. These findings indicate that hyperactivation of MAP kinase increases phospho-tau levels within cells, despite that MAP kinase apparently does not substantially influence intracellular tau phosphorylation under normal conditions. These findings underscore that results obtained following inhibition of kinase activities do not necessarily provide an indication of the consequences accompanying hyperactivation of that same kinase. Several studies conducted in cell-free systems indicate that exposure of tau to multiple kinases can have synergistic effects on the nature and extent of tau phosphorylation. We therefore examined whether or not such effects could be demonstrated within these cells. Site-specific phospho-tau immunoreactivity was increased in additive and synergistic manners by treatment of injected cells with TPA (which activates PKC), calcium ionophore (which activates calcium-dependent kinases), and wortmannin (which inhibits PIP3 kinase). Alteration in total tau levels was insufficient to account for the full extent of the increase in phospho-tau immunoreactivity. These additional results indicate that multiple kinase activities modulate the influence of MAP kinase on tau within intact cells.