Objective: To investigate the role and molecular mechanism of surviving, an anti-apoptosis gene, in cell growth and transformation.
Methods: Coding sequence of surviving was amplified from Daudi cell mRNA by RT-PCR and then cloned into prokaryotic and eukaryotic vectors. The vectors with surviving were transfected into BL21 cells of Escherichia coli and human embryonic kidney 293 cells. The cells were cultured. Protein was extracted from the cells and examined by gel electrophoresis. Suspension of 293 cells was cultured and the number of cells was counted every other day, thus a growth curve was drawn. Another suspension of 293 cells was cultured in soft agar to observe the number of colonies. Cells transfected with plasmids void of surviving were used as controls.
Results: The anti-apoptosis gene surviving was well expressed in BL21 cells and 293 cells. The growth curve showed that the proliferation rate of 293 cells was slightly faster than that of control cells, however, without significant difference. Soft agar assay showed that the colonies formed by surviving-transfected 293 cells were of greater size and with greater number. Western blotting showed overexpression of cyclin D1 and c-myc, two important cancer proteins, in cells transfected with surviving.
Conclusion: The anti-apoptosis gene surviving promotes cell transformation. These effects may depend on the functions of cyclin D1 and c-myc.