ERK5 MEDIATES THE MACROPHAGE COLONY-STIMULATING FACTOR SIGNALLING IN A SRC-DEPENDENT AND PKC- INDEPENDENT FASHION

The Macrophage Colony-Stimulating factor (M-CSF) sustains the survival and proliferation as well cell spreading and adhesion of monocyte/ macrophage cells. ERK5 (also known as big MAPK1-BMK1) a member of the mitogen-activated protein kinase (MAPK) family, is a 98-kDa molecule sharing homology with ERK1/2. ERK5 is activated by a kinase cascade involving the upstream kinases MEK5, MEKK2/3 and Tpl-2. ERK5 plays a role as a redox-sensitive protein kinase, and may be activated in response to growth factor stimulation. This study was undertaken to characterize the involvement of ERK5 in M-CSF elicited signaling in the murine macrophage cell line BAC1.2F5 and in human bone marrow derived macrophages. Exposure to M-CSF resulted in a rapid and transient induction of ERK5 phosphorylation in activation-specific residues. Moreover, upon exposure to M-CSF, ERK5 showed a rapid and transient translocation from the cytosol to the nucleus, as indicated by immunofluorescence and cell fractionation. Induction of ERK5 activation was also observed in response to H2O2, a well known activator of ERK5, while macrophage activators, such as LPS or Interferon-g, failed to activate ERK5. Activation of ERK5 following M-CSF administration was peculiar of macrophages as M-CSF failed to activate ERK5 in NIH/3T3 fibroblasts expressing ectopic M-CSF receptor (M-CSF.R). To assess the contribution of pathways downstream of the M-CSF.R, activation of ERK5 was measured in the presence of different pharmacologic inhibitors. Inhibition of PKC isoforms or the serino-phosphatases PP1 and PP2A did not have any effects on ERK5 activation, while several molecules interfering with Src-family kinases activation inhibited MCSF- dependent ERK5 phosphorylation. Preliminary data performed with siRNA targeting ERK5 seem to indicate that ERK5 is involved in M-CSFinduced macrophage proliferation.