A chimeric growth factor receptor (CSF1R/IR) was constructed by splicing cDNA sequences encoding the extracellular ligand binding domain of the human colony stimulating factor-1 (CSF-1) receptor to sequences encoding the transmembrane and cytoplasmic domains of the human insulin receptor. The addition of CSF-1 to cells transfected with the CSF1R/IR chimera cDNA stimulated the tyrosine phosphorylation of a protein that was immunoprecipitated by an antibody directed against the carboxyl terminus of the insulin receptor. Phosphopeptide maps of the 32P-labeled CSF1R/IR protein revealed the same pattern of phosphorylation observed in 32P-labeled insulin receptor beta subunits. CSF-1 stimulated the tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) and Shc in cells expressing the CSF1R/IR chimera. Lipid accumulation and the expression of a differentiation-specific marker demonstrated that 3T3-L1 preadipocytes undergo CSF-1-dependent differentiation when transfected with the CSF1R/IR chimera cDNA but not when transfected with the expression vector alone. A 12-amino acid deletion within the juxtamembrane region of the CSF1R/IR (CSF1R/IRDelta960) blocked CSF-1-stimulated phosphorylation of IRS-1 and Shc but did not inhibit CSF-1-mediated differentiation of 3T3-L1 preadipocytes. These observations indicate that adipocyte differentiation can be initiated by intracellular pathways that do not require tyrosine phosphorylation of IRS-1 or Shc.