Single-cell analysis of transcription kinetics across the cell cycle.
Samuel O SkinnerHeng XuSonal Nagarkar-JaiswalPablo R FreireThomas P ZwakaIdo GoldingPublished in: eLife (2016)
Transcription is a highly stochastic process. To infer transcription kinetics for a gene-of-interest, researchers commonly compare the distribution of mRNA copy-number to the prediction of a theoretical model. However, the reliability of this procedure is limited because the measured mRNA numbers represent integration over the mRNA lifetime, contribution from multiple gene copies, and mixing of cells from different cell-cycle phases. We address these limitations by simultaneously quantifying nascent and mature mRNA in individual cells, and incorporating cell-cycle effects in the analysis of mRNA statistics. We demonstrate our approach on Oct4 and Nanog in mouse embryonic stem cells. Both genes follow similar two-state kinetics. However, Nanog exhibits slower ON/OFF switching, resulting in increased cell-to-cell variability in mRNA levels. Early in the cell cycle, the two copies of each gene exhibit independent activity. After gene replication, the probability of each gene copy to be active diminishes, resulting in dosage compensation.
Keyphrases
- cell cycle
- copy number
- genome wide
- cell proliferation
- mitochondrial dna
- single cell
- genome wide identification
- induced apoptosis
- embryonic stem cells
- transcription factor
- binding protein
- dna methylation
- cell cycle arrest
- cell therapy
- genome wide analysis
- high throughput
- rna seq
- oxidative stress
- endoplasmic reticulum stress
- cancer stem cells