A somatic cell genetic system for dissecting hemopoietic cytokine signal transduction

Somatic cell genetics has proven to be a powerful tool for the dissection of cytokine signal transduction pathways. Here we describe a system in which interleukin-6 (IL-6) signaling may be dissected using myeloid leukemic M1 cells. We utilized two properties of M1 cell differentiation to isolate IL-6-unresponsive mutants. First, M1 differentiation is associated with cessation of cell division. Second, differentiated M1 cells migrate rapidly and form dispersed colonies in agar. Mutant clones that are unresponsive to IL-6 are, therefore, large and compact as compared with clones derived from IL-6-responsive wild type M1 cells. Following spontaneous or chemically induced mutagenesis and selection in a high dose of IL-6, we isolated 27 M1 clones unresponsive to IL-6. Three harbored mutations that acted in a dominant manner, whereas 24 contained recessive mutations. gp130, an IL-6 receptor component, was affected in many mutant clones. We show that these clones display IL-6 and leukemia inhibitory factor receptors with reduced binding affinities and express gp130 at reduced levels. The IL-6-unresponsive phenotype of these mutant clones was fully rescued by the transfection of exogenous gp130 DNA. Therefore, this approach targets components of the IL-6 signaling pathway and may be suitable to study signaling from a variety of cytokines.