These studies demonstrate that CSF-1 and IL-34 are conserved in birds, and uses evolutionary comparisons to infer structure function relationships among vertebrate animals.

Macrophages are involved in many aspects of development, host defense, pathology, and homeostasis. Their normal differentiation, proliferation, and survival are controlled by CSF-1 via the activation of the CSF1R. A recently discovered cytokine, IL-34, was shown to bind the same receptor in humans. Chicken is a widely used model organism in developmental biology, but the factors that control avian myelopoiesis have not been identified previously. The CSF-1, IL-34, and CSF1R genes in chicken and zebra finch were identified from respective genomic/cDNA sequence resources. Comparative analysis of the avian CSF1R loci revealed likely orthologs of mammalian macrophage-specific promoters and enhancers, and the CSF1R gene is expressed in the developing chick embryo in a pattern consistent with macrophage-specific expression. Chicken CSF-1 and IL-34 were expressed in HEK293 cells and shown to elicit macrophage growth from chicken BM cells in culture. Comparative sequence and co-evolution analysis across all vertebrates suggests that the two ligands interact with distinct regions of the CSF1R. These studies demonstrate that there are two separate ligands for a functional CSF1R across all vertebrates.