CELL-CYCLE REGULATION AND SUBCELLULAR-LOCALIZATION OF THE MAJOR HUMAN URACIL-DNA GLYCOSYLASE

The subcellular localization of the human DNA-repair enzyme uracil-DNA glycosylase from the UNG gene has been studied using how cytometry and laser scanning confocal microscopy of freely cycling HeLa S-3 cells. A two-parameter how cytometric analysis using propidium iodide and UNG-specific antibodies demonstrated that total cellular UNG increased during the G(1)-phase and was approximately doubled in early S-phase compared to early G(1). The UNG level was stable during the S-phase and increased further during G(2), reaching a 2.8-fold level compared to early G(1). This factor included differences in cell size and staining variabilities. These findings were confirmed using two-parameter confocal analysis of UNG/DNA and UNG/mitochondria at different stages of the cell cycle. Although the major fp action of UNG was associated with nuclei, we also observed distinctive staining associated with mitochondria and a more diffuse staining probably reflecting UNG in the cytosol. Furthermore, very little UNG staining was observed in nucleoli. The UNG level in different cell compartments varied at different stages of the cell cycle, and this variation was most pronounced in the nuclei. These results demonstrate that the gene product from the UNG gene is located within three subcellular compartments and that the distribution between these compartments varies during the cell cycle. (C) 1995 Academic Press, Inc.