The crystal structure of the signaling complex of human granulocyte colony-stimulating factor (GCSF) and ligand binding region of human GCSF receptor (GCSF-R) has been determined to 2.8 Å resolution. The GCSF : GCSF-R complex formed a 2 : 2 stoichiometry via a crossover interaction between the Ig-like domains of GCSF-R and GCSF. The conformation of the complex is quite different to that between human GCSF and the CRH domain of mouse GCSF-R. This cross-over homodimerization necessary for GCSF-R activation is consistent with previously reported thermodynamic and mutational studies.