Regulation of the cell cycle via mitochondrial gene expression and energy metabolism in HeLa cells.
ABSTRACT Human cervical cancer HeLa cells have functional mitochondria. Recent studies have suggested that mitochondrial metabolism plays an essential role in tumor cell proliferation. Nevertheless, how cells coordinate mitochondrial dynamics and cell cycle progression remains to be clarified. To investigate the relationship between mitochondrial function and cell cycle regulation, the mitochondrial gene expression profile and cellular ATP levels were determined by cell cycle progress analysis in the present study. HeLa cells were synchronized in the G0/G1 phase by serum starvation, and re-entered cell cycle by restoring serum culture, time course experiment was performed to analyze the expression of mitochondrial transcription regulators and mitochondrial genes, mitochondrial membrane potential (MMP), cellular ATP levels, and cell cycle progression. The results showed that when arrested G0/G1 cells were stimulated in serum-containing medium, the amount of DNA and the expression levels of both mRNA and proteins in mitochondria started to increase at 2 h time point, whereas the MMP and ATP level elevated at 4 h. Furthermore, the cyclin D1 expression began to increase at 4 h after serum triggered cell cycle. ATP synthesis inhibitor-oligomycin-treatment suppressed the cyclin D1 and cyclin B1 expression levels and blocked cell cycle progression. Taken together, our results suggested that increased mitochondrial gene expression levels, oxidative phosphorylation activation, and cellular ATP content increase are important events for triggering cell cycle. Finally, we demonstrated that mitochondrial gene expression levels and cellular ATP content are tightly regulated and might play a central role in regulating cell proliferation.
Article: Increases in mitochondrial DNA content and 4977-bp deletion upon ATM/Chk2 checkpoint activation in HeLa cells.[show abstract] [hide abstract]
ABSTRACT: Activation of the Mec1/Rad53 damage checkpoint pathway influences mitochondrial DNA (mtDNA) content and point mutagenesis in Saccharomyces cerevisiae. The effects of this conserved checkpoint pathway on mitochondrial genomes in human cells remain largely unknown. Here, we report that knockdown of the human DNA helicase RRM3 enhances phosphorylation of the cell cycle arrest kinase Chk2, indicating activation of the checkpoint via the ATM/Chk2 pathway, and increases mtDNA content independently of TFAM, a regulator of mtDNA copy number. Cell-cycle arrest did not have a consistent effect on mtDNA level: knockdown of cell cycle regulators PLK1 (polo-like kinase), MCM2, or MCM3 gave rise, respectively, to decreased, increased, or almost unchanged mtDNA levels. Therefore, we concluded that the mtDNA content increase upon RRM3 knockdown is not a response to delay of cell cycle progression. Also, we observed that RRM3 knockdown increased the levels of reactive oxygen species (ROS); two ROS scavengers, N-acetyl cysteine and vitamin C, suppressed the mtDNA content increase. On the other hand, in RRM3 knockdown cells, we detected an increase in the frequency of the common 4977-bp mtDNA deletion, a major mtDNA deletion that can be induced by abnormal ROS generation, and is associated with a decline in mitochondrial genome integrity, aging, and various mtDNA-related disorders in humans. These results suggest that increase of the mitochondrial genome by TFAM-independent mtDNA replication is connected, via oxidative stress, with the ATM/Chk2 checkpoint activation in response to DNA damage, and is accompanied by generation of the common 4977-bp deletion.PLoS ONE 01/2012; 7(7):e40572. · 4.09 Impact Factor