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) o n the growth and transformation of mammalian cells. We have used for t his purpose R(-) cells, which are 3T3-like fibroblasts derived from mo use embryos with a targeted disruption of the IGF-IR genes. These cell s have no IGF-IR, do not grow in serum-free medium supplemented with t he 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 t ype human IGF-IR cDNA; 2) a receptor with a triple mutation in the abo ve mentioned tyrosines; and 3) receptors with single tyrosine mutation s. Cells expressing the wild type or the single tyrosine mutants Y1 (Y 1131F) and Y2 (Y1135F) grew in serum-free medium supplemented solely w ith IGF-I. Cells expressing the triple tyrosine mutant YF or the singl e mutant Y3 (Y1136F) failed to grow in response to IGF-I only. Ah muta nts, though, failed to form colonies in soft agar, indicating that a f ully functional IGF-IR is more critical for anchorage-independent grow th than for monolayer growth. The triple mutant expression plasmid als o functioned as a dominant negative, inhibiting the growth of wild typ e cells transformed by the simian virus tumor antigen.