CELL-CYCLE ANALYSIS OF FOREIGN GENE (BETA-GALACTOSIDASE) EXPRESSION IN RECOMBINANT MOUSE CELLS UNDER CONTROL OF MOUSE MAMMARY-TUMOR VIRUS PROMOTER

The cell cycle dependency of foreign gene expression in recombinant mo use L cells was investigated. Two different recombinant mouse L cell l ines having the glucocorti cold receptor-encoding gene and the lacZ re porter gene were used in this study. The lacZ gene expression was cont rolled by the glucocorticoid-inducible mouse mammary tumor virus (MMTV ) promoter in both cell lines. In ''M4'' cells the gr gene was under t he control of another MMTV promoter, but in ''R2'' cells it was under the control of the constitutive Rous sarcoma virus promoter. These nor mally attachment-grown cells were adapted to suspension culture, and a dual-laser flow cytometer was used to simultaneously determine the DN A and foreign protein (beta-galactosidase) content of single living ce lls. Expression of beta-galactosidase as a function of cell cycle phas e was evaluated for cells in exponential growth without any addition o f the glucocorticoid inducer, dexamethasone. Cell cycle positions in t he S phase were estimated on the basis of DNA content per cell, and po sition in the G1 phase was estimated on the basis of cell size as meas ured by pulse-width time of flight. The results showed that beta-galac tosidase synthesis occurred through all cell cycle phases, but the exp ression rate in the G1 phase was much lower than that in the S and G2/ M phases in both cell lines. On the basis of cell size analysis, beta- galactosidase expression in M4 cells (with autoinducible promoter) was found to be higher than that in R2 cells (with inducible promoter) du ring the G1 phase. (C) 1996 John Wiley & Sons, Inc.