Mitogenicity and transforming activity of the insulin-like growth factor-I receptor with mutations in the tyrosine kinase domain.

We have investigated the effect of mutations in tyrosines 1131, 1135, and 1136 of the human insulin-like growth factor-I receptor (IGF-IR) on the growth and transformation of mammalian cells. We have used for this purpose R- cells, which are 3T3-like fibroblasts derived from mouse embryos with a targeted disruption of the IGF-IR genes. These cells have no IGF-IR, do not grow in serum-free medium supplemented with the growth factors that sustain the growth of 3T3 cells, and cannot be transformed by simian virus 40 large tumor antigen or other oncogenes. The R- cells were transfected with plasmids expressing: 1) a wild type human IGF-IR cDNA; 2) a receptor with a triple mutation in the above mentioned tyrosines; and 3) receptors with single tyrosine mutations. Cells expressing the wild type or the single tyrosine mutants Y1 (Y1131F) and Y2 (Y1135F) grew in serum-free medium supplemented solely with IGF-I. Cells expressing the triple tyrosine mutant YF or the single mutant Y3 (Y1136F) failed to grow in response to IGF-I only. All mutants, though, failed to form colonies in soft agar, indicating that a fully functional IGF-IR is more critical for anchorage-independent growth than for monolayer growth. The triple mutant expression plasmid also functioned as a dominant negative, inhibiting the growth of wild type cells transformed by the simian virus tumor antigen.