Protein phosphatase 2A is expressed in response to colony-stimulating factor 1 in macrophages and is required for cell cycle progression independently of extracellular signal-regulated protein kinase activity

Colony-stimulating factor 1 (CSF-1) is required for the development of mono cytes/macrophages from progenitor cells and for the survival and activation of mature macrophages. The receptor for CSF-1 is the product of the c-fms proto-oncogene, which, on binding ligand, can stimulate a mitogenic respons e in the appropriate cells. To investigate which genes are regulated in res ponse to CSF-l-stimulation in murine bone-marrow-derived macrophages (BMM), we employed mRNA differential display reverse transcriptase-mediated PCR t o identify cDNA species induced by CSF-I. Both Northern and Western blot an alyses confirmed the increased expression of one of the cDNA species identi fied as coding for the catalytic subunit of protein phosphatase 2A (PP2A), an observation not previously reported during the response to a growth fact or. To determine the significance of the increased expression of PP2A in re sponse to CSF-1, the PP2A inhibitor okadaic acid (OA) was added to CSF-l-tr eated BMM and found to inhibit DNA synthesis in a dose-dependent manner. Fu rther analysis with flow cytometry in the presence of OA led to the novel c onclusion that PP2A activity is critical for CSF-l-driven BMM cell cycle pr ogression in both early G(1) and S phases. Surprisingly, in the light of pr evious studies with other cells, the PP2A-dependent proliferation could be dissociated from activation by extracellular signal-regulated protein kinas e (ERK) in macrophages because OA did not affect either the basal or CSF-l- induced ERK activity in BMM. Two-dimensional SDS/PAGE analysis of lysates o f P-32-labelled BMM, which had been treated with CSF-1 in the presence or a bsence of OA, identified candidate substrates for PP2A.