Regulation of cell cycle entry and G1 progression by CSF-1

Proliferation, differentiation, and survival of monocytes, macrophages, and their immediate progenitors is regulated by the macrophage colony-stimulating factor (CSF-1). CSF-1 initiates a mitogenic response by binding to its receptor (CSF-1R), thereby activating the receptor's intrinsic tyrosine kinase activity and initiating signaling via multiple effector-mediated pathways. CSF-1 is required throughout G1 to ensure entry of bone marrow-derived macrophages into S phase, and persistent CSF-1R kinase activity is necessary to the expression of both immediate early (e.g., c-fos, c-jun, and c-myc) and delayed early (e.g., D-type cyclins) response genes. Ectopic expression of human CSF-1R in different mouse cell lines, including fibroblasts, IL-3-dependent myeloid cells, and early pre-B cells, confers CSF-1 responsiveness by replacing the cells' requirements for other mitogenic growth factors. NIH-3T3 fibroblasts engineered to express a human CSF-1 receptor point mutant (CSF-1R [Y809F]) fail to proliferate in response to CSF-1 and remain arrested in the early G1 phase of the cell cycle. Despite CSF-1-dependent transcription of fos and jun family members, c-myc, D-type, and E-type G1 cyclin mRNAs are not expressed in the latter cells in response to growth factor stimulation. However, enforced expression of c-myc or D-type cyclins, but not cyclin E, resensitizes cells bearing CSF-1R (Y809F) to the mitogenic effects of CSF-1, enabling them to proliferate continuously in liquid culture and to form colonies in agar in response to the growth factor. D-type cyclin mutants defective in binding to the retinoblastoma protein (pRB) were unable to rescue mutant receptor signaling, suggesting that the ability of D-type cyclin-dependent kinases to cancel pRB's growth-suppressive function is necessary for CSF-1-induced G1 exit. By contrast, cyclin E must function in a different pathway. Cells rescued by c-myc were prevented from entering S phase by microinjection of antibodies to cyclin D1. Conversely, cyclin D1-rescued cells were inhibited from forming CSF-1-dependent colonies in agar when challenged with either a dominant-negative c-myc mutant or mad, a transcription factor which competes with myc for max, its requisite heterodimeric partner. Thus, although the expression of c-myc and D-type cyclins is rate limiting for G1 phase progression, their functions are interdependent, with both activities being required for mitogenicity.