D'Ambrosio C, Hongo A, Li S, Baserga R
Jefferson Cancer Center, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Oncogene. 1996 Jan 18;12(2):371-8.
An overexpressed insulin-like growth factor I receptor (IGF-IR) allows cells to grow in IGF-I only and to form colonies in soft agar. Conversely, cells with a targeted disruption of the IGF-IR genes, R- cells, are refractory to transformation by several oncoproteins and growth factor receptors, that readily transform their wild type counterparts, W cells. Grb2 is an SH2-SH3 domains protein that links tyrosine kinase receptors to ras signalling. In order to determine its role in mitogenesis and transformation, we have transfected a plasmid expressing Grb2 into R- and W cells, and their derivatives already expressing the SV40 large T antigen. In addition, we have used loss-of-function mutants of Grb2 to inquire whether they would act as dominant negatives. Our results show that: (1) an overexpressed Grb2 cannot replace the IGF-IR in IGF-I-mediated mitogenesis; (2) Grb2 also fails to transform either W or R- cells; (3) Grb2 and SV40 T antigen, singly transfected, cannot transform R- cells, but can do so when combined; and (4) SH3 domain mutants of Grb2 act as dominant negatives, causing reversion of the transformed phenotype. We conclude that Grb2 is necessary but not sufficient for transformation.
过度表达的胰岛素样生长因子I受体（IGF-IR）使细胞仅在IGF-I中生长并在软琼脂中形成集落。相反，IGF-IR基因靶向破坏的细胞，即R-细胞，对几种癌蛋白和生长因子受体的转化具有抗性，而这些蛋白和受体能轻易地转化其野生型对应物W细胞。Grb2是一种SH2-SH3结构域蛋白，它将酪氨酸激酶受体与ras信号传导联系起来。为了确定其在有丝分裂和转化中的作用，我们将表达Grb2的质粒转染到R-细胞和W细胞及其已表达SV40大T抗原的衍生物中。此外，我们使用了Grb2的功能丧失突变体来探究它们是否会作为显性负性物起作用。我们的结果表明：（1）过度表达的Grb2不能替代IGF-IR在IGF-I介导的有丝分裂中的作用；（2）Grb2也不能转化W细胞或R-细胞；（3）单独转染的Grb2和SV40 T抗原不能转化R-细胞，但联合转染时可以；（4）Grb2的SH3结构域突变体作为显性负性物起作用，导致转化表型的逆转。我们得出结论，Grb2对于转化是必要的，但不是充分的。
