J Biol Chem, Vol. 274, Issue 19, 13118-13126, May 7, 1999

Diminished G₁ Checkpoint after -Irradiation and Altered Cell Cycle Regulation by Insulin-like Growth Factor II Overexpression

Lijuan Zhang, Min Kim^§, Yung Hyun Choi^¶, Bianca Goemans, Choh Yeung, Zongyi Hu, Shili Zhan, Prem Seth^§, and Lee J. Helman

From the  Molecular Oncology Section, ^§ Medical Breast Cancer Section, ^¶ Cell Signaling and Oncogenesis Section, NCI,  Liver Diseases Section, NIDDK, National Institutes of Health, Bethesda, Maryland 20892-1928

High levels of insulin-like growth factor II (IGFII) mRNA expression are detected in many human tumors of different origins including rhabdomyosarcoma, a tumor of skeletal muscle origin. To investigate the role of IGFII in tumorigenesis, we have compared the mouse myoblast cell line C2C12-2.7, which was stably transfected with human IGFII cDNA and expressed high and constant amounts of IGFII, to a control cell line C2C12-1.1. A rhabdomyosarcoma cell line, RH30, which expresses high levels of IGFII and contains mutated p53, was also used in these studies. IGFII overexpression in mouse myoblast C2C12 cells causes a reduced cycling time and higher growth rate. After -irradiation treatment, C2C12-1.1 cells were arrested mainly in G₀/G₁ phase. However, C2C12-2.7 and RH30 cells went through a very short G₁ phase and then were arrested in an extended G₂/M phase. To verify further the effect of IGFII on the cell cycle, we developed a Chinese hamster ovary (CHO) cell line with tetracycline-controlled IGFII expression. We found that CHO cells with high expression of IGFII have a shortened cycling time and a diminished G₁ checkpoint after treatment with methylmethane sulfonate (MMS), a DNA base-damaging agent, when compared with CHO cells with very low IGFII expression. It was also found that IGFII overexpression in C2C12 cells was associated with increases in cyclin D1, p21, and p53 protein levels, as well as mitogen-activated protein kinase activity. These studies suggest that IGFII overexpression shortens cell cycling time and diminishes the G₁ checkpoint after DNA damage despite an intact p53/p21 induction. In addition, IGFII overexpression is also associated with multiple changes in the levels and activities of cell cycle regulatory components following -irradiation. Taken together, these changes may contribute to the high growth rate and genetic alterations that occur during tumorigenesis.