Changes in the Level of Usp28 Deubiquitinase in the Cell Cycle of HCT116 Intestinal Adenocarcinoma Cells Indicate Its Functional Role in the Regulation of G 1 /S Transition

Usp28 is a deubiquitinating enzyme that removes ubiquitin from its conjugates with substrates and prevents their degradation in proteasomes. Modern publications show that Usp28 and the Fbw7 ubiquitin ligase create a functional pair of proteins that controls ubiquitin-mediated degradation of key regulators of cellular functions, including Myc, Jun, Nicd, and Hif1. In this pair of proteins, Usp28 counteracts the destructive activity of Fbw7 and plays the role of a factor promoting tumor growth. Since the Usp28 targets are the Myc and Jun proteins associated with the cell cycle, we suggested that Usp28 regulates cell division and its level may change during the cell cycle. The aim of this work was to assess changes in the level of Usp28 during the cell cycle in HCT116 human intestinal carcinoma cells. HCT116 cells were synchronized by 72-h cultivation in a growth medium with a low serum content. In asynchronously dividing cells, the level of Usp28 was low and its localization was detected as nuclear–cytoplasmic. The general level of Usp28 and the degree of its nuclear localization increased in cells from the state of asynchronous growth to the late phase G 1 followed by a decrease and redistribution of protein from the nucleus to the cytoplasm after the completion of phase G 1 . According to immunoblot data, the fluctuations in the levels of Usp28 and Cdc25A phosphatase in synchronized HCT116 cells in the cell cycle coincided. The immunofluorescence data directly corresponded to the results of immunoblotting. The results suggest that Usp28 can regulate the level and functional activity of the Cdc25A protein that controls the entry of cells into the DNA replication phase.