SATURATION MUTAGENESIS OF THE BETA-SUBUNIT OF THE HUMAN GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR-RECEPTOR SHOWS CLUSTERING OF CONSTITUTIVE MUTATIONS, ACTIVATION OF ERK MAP KINASE AND STAT PATHWAYS, ANDDIFFERENTIAL BETA-SUBUNIT TYROSINE PHOSPHORYLATION

The high-affinity receptors for human granulocyte-macrophage colony-st imulating factor (GM-CSF), interleukin-1 (IL-3), and IL-5 are heterodi meric complexes consisting of cytokine-specific alpha subunits and a c ommon signal-transducing beta subunit (h beta c). We have previously d emonstrated the oncogenic potential of this group of receptors by iden tifying constitutively activating point mutations in the extracellular and transmembrane domains of h beta c. We report here a comprehensive screen of the entire h beta c molecule that has led to the identifica tion of additional constitutive point mutations by virtue of their abi lity to confer factor independence on murine FDC-P1 cells. These mutat ions were clustered exclusively in a central region of h beta c that e ncompasses the extracellular membrane-proximal domain, transmembrane d omain, and membrane-proximal region of the cytoplasmic domain. Interes tingly, most h beta c mutants exhibited cell type-specific constitutiv e activity, with only two transmembrane domain mutants able to confer factor independence on both murine FDC-P1 and BAF-B03 cells. Examinati on of the biochemical properties of these mutants in FDC-P1 cells indi cated that MAP kinase (ERK1/2), STAT, and JAK2 signaling molecules wer e constitutively activated. In contrast, only some of the mutant beta subunits were constitutively tyrosine phosphorylated. Taken together; these results highlight key regions involved in h beta c activation, d issociate h beta c tyrosine phosphorylation from MAP kinase and STAT a ctivation, and suggest the involvement of distinct mechanisms by which proliferative signals can be generated by h beta c. (C) 1998 by The A merican Society of Hematology.