Variations in DNA Methylation Patterns During the Cell Cycle of HeLa Cells

Prospects for Epigenetic Epidemiology

“…The primary DNA sequence is generally fixed at conception, but epigenetic marks are dynamic and modifiable, probably throughout the life course. Recent work with human cell lines has shown evidence of dynamic reprogramming of epigenetic markings during the cell cycle (15)(16)(17). The expression of genetic risk is therefore likely to show varying penetrance over time in response to epigenetic profile.…”

Dynamic epigenetic states of ribosomal RNA promoters during the cell cycle

“…This method has been widely used to study a range of different processes in the cell cycle, 15,[28][29][30] including epigenetic changes. 14,31 This method halts the cells early in S phase, with the subsequent S phase being slightly shorter than normal, 32 and cells are able to recover fully following the double thymidine block. 33 Once the block is released, the cells cycle through S phase as a synchronized population, thus enabling examination of the epigenetic changes at specific points during S phase, G 2 and G 1 .…”

“…It is unlikely that these observations are due to the synchronization procedure as we have previously shown that global DNA methylation levels in HeLa cells at different stages of the cell cycle are similar in unsynchronized cells sorted into different phases of the cell cycle and cells synchronized using a double thymidine block. 14 Furthermore, double thymidine block was previously demonstrated to result in increased methylation of the genome, 31 therefore the decrease in DNA methylation at the rRNA promoter is unlikely to be the result of the synchronization procedure. To our knowledge, no previous work has shown selective hypomethylation of a promoter as the result of a double thymidine block synchronization procedure.…”

“…However, it is unclear whether changes in epigenetic states are involved in transient physiological responses such as the response in gene expression to the transition through the different phases of the cell cycle. We have recently shown that changes in global and sequence specific DNA methylation occur during one cell cycle in HeLa cells 14 however it was not clear whether these cell cycle associated changes in DNA methylation play any regulatory role. In this study we addressed the question of whether the epigenetic state of rRNA genes is stable or dynamic during different phases of the cell cycle of human HeLa cells.…”

“…Cells were synchronized at G 1 using serum starvation, or at G 1 /S using a double thymidine block, as previously described. 14,15 Briefly, HeLa cells were treated with 2.5 mM thymidine (Sigma) for 16 h, washed with PBS and then cultured in fresh DMEM + 10% FCS for 8 h. Next, the cells were again treated with 2.5 mM thymidine for 16 h. Cells were released from the block by washing with PBS and replacing the medium with fresh DMEM + 10% FCS. Cells at different stages of the cell cycle were obtained by harvesting them at varying lengths of time following cell cycle induction.…”