ANALYSIS OF GENE EXPRESSION IN NORMAL AND ONCOGENETRANSFECTED CELLS OF RAT MODEL

Abstract

Cells in culture offer many opportunities to study the characteristics of both the normal and cancer cells. In order to understand the molecular mechanisms of cell proliferation, an attempt has been made to study the proteins which are at different phases of the cell cycle. To achieve this objective, rat embryo fibroblasts were synchronized at Go phase by serum starvation for 72h and then stimulated with serum mitogens or purified growth factors. The newly synthesized proteins were labeled with [35S] methionine at different phases of the cell cycle after stimulation and the secreted proteins were analyzed by SDS- polyacrylamide gels. One of the proteins which has shown to be involved in growth regulation was purified and bioassays were carried out to determine its function. The protein 48 KDa was found to be a major compound of the extracellular matrix (ECM) whereas the protein 26 KDa was not a matrix associated protein. When the cells were arrested at G1/S boundary with hydroxyurea (HOU), high levels of protein with 45KDa protein was observed in the medium. Normal rat embro fibroblasts were transfected with myc and ras oncogenes and the transformed colonies were cultured and purified 45KDa protein inhibited the DNA synthesis of myc and ras oncogene transformed cells in which the 45KDa protein secretion was down regulated, indicating the inhibition of DNA synthesis.