Protein profiles of the Chinese hamster ovary cells in the resting and proliferating stages

Identification and characterization of the proteins that regulate the trans ition from the resting stage (G0) through G1 to S phase of the cell cycle a re of central importance to understand the control of cell proliferation an d chromosome replication. Unlike in lower organisms, where relatively small numbers of key factors are involved in this process, the factors involved in the same control mechanisms in mammalian systems are much more complex. Furthermore, accumulating lines of evidence now suggest that the nuclear ma trix and chromatin organization also play an essential role for the cell cy cle control in mammalian cells. To gain a better understanding of the overa ll dynamics and changes of the protein factors in the context of matrix/chr omatin organization, we examined the protein profiles of the Chinese hamste r ovary (CHO) cells in different cell cycle compartments. The methods used in this study included subcellular fractionations (cytosol, nuclear extract ion, chromatin, and nuclear matrix), two-dimensional polyacrylamide gel ele ctrophoresis (2-D PAGE), silver staining, and immunoblotting. As expected, significant changes of protein profiles were observed when cells entered in to proliferating stages from GO. Among approximately 1200 protein spots ana lyzed by 2-D PAGE, at least 12 showed marked increase or decrease at this t ransitional period. Further cell-cycle progression from G1 to S phase showe d less dramatic changes of overall protein protile. However, the profile of certain proteins showed rather dramatic changes of their subcellular local ization during this transitional period. In particular, the levels of proli ferating cell nuclear antigen (PCNA) in the nuclear matrix and chromatin dr amatically increased in mid-G1 and in the beginning of S phase, respectivel y, while the overall PCNA level was relatively constant throughout the cell cycle.