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.
Li
S
S
Jefferson Cancer Institute, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Ferber
A
A
Miura
M
M
Baserga
R
R
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002448:Cell Adhesion; D002455:Cell Division; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D016895:Culture Media, Serum-Free; D004247:DNA / Q000096:biosynthesis; D017931:DNA Primers; D005799:Genes, Dominant; D055504:Insulin Receptor Substrate Proteins; D051379:Mice; D018345:Mice, Knockout; D008934:Mitogens / Q000502:physiology*; D008969:Molecular Sequence Data; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000235:genetics*; D017526:Receptor, IGF Type 1 / Q000235:genetics / Q000502:physiology*
D016895:Culture Media, Serum-Free; D017931:DNA Primers; C526292:IRS1 protein, human; D055504:Insulin Receptor Substrate Proteins; C526295:Irs1 protein, mouse; D008934:Mitogens; D010750:Phosphoproteins; D004247:DNA; D011505:Protein-Tyrosine Kinases; D017526:Receptor, IGF Type 1