Dynamic Heterogeneity and DNA Methylation in Embryonic Stem Cells

Zakary S Singer,John Yong,Julia Tischler,Jamie A Hackett,Alphan Altinok,M Azim Surani,Long Cai,Michael B Elowitz

doi:10.1016/j.molcel.2014.06.029

Abstract

SummaryCell populations can be strikingly heterogeneous, composed of multiple cellular states, each exhibiting stochastic noise in its gene expression. A major challenge is to disentangle these two types of variability and to understand the dynamic processes and mechanisms that control them. Embryonic stem cells (ESCs) provide an ideal model system to address this issue because they exhibit heterogeneous and dynamic expression of functionally important regulatory factors. We analyzed gene expression in individual ESCs using single-molecule RNA-FISH and quantitative time-lapse movies. These data discriminated stochastic switching between two coherent (correlated) gene expression states and burst-like transcriptional noise. We further showed that the “2i” signaling pathway inhibitors modulate both types of variation. Finally, we found that DNA methylation plays a key role in maintaining these metastable states. Together, these results show how ESC gene expression states and dynamics arise from a combination of intrinsic noise, coherent cellular states, and epigenetic regulation.

Highlights

Many cell populations appear to consist of mixtures of cells in distinct cellular states
This heterogeneity could in principle arise from stochastic fluctuations, or ‘‘noise,’’ in gene expression (Eldar and Elowitz, 2010; Raj et al, 2008; Zenklusen et al, 2008). It could reflect the coexistence of multiple cellular states, each with a distinct gene expression pattern showing correlation between a set of genes (Guo et al, 2010; Gupta et al, 2011; Jaitin et al, 2014; Shalek et al, 2013). Disentangling these two sources of variation is important for interpreting the transcriptional states of individual cells and understanding stem cell dynamics
In order to accurately measure mRNA copy numbers in large Third, there were some genes whose mRNA distributions were numbers of individual Embryonic stem cells (ESCs), we developed an automated significantly better fit by a linear combination of two negative binomial (NB) distribuplatform for smFISH (Supplemental Information)

Summary

SUMMARY

Cell populations can be strikingly heterogeneous, composed of multiple cellular states, each exhibiting stochastic noise in its gene expression. Embryonic stem cells (ESCs) provide an ideal model system to address this issue because they exhibit heterogeneous and dynamic expression of functionally important regulatory factors. We analyzed gene expression in individual ESCs using single-molecule RNA-FISH and quantitative time-lapse movies. These data discriminated stochastic switching between two coherent (correlated) gene expression states and burst-like transcriptional noise. We found that DNA methylation plays a key role in maintaining these metastable states. Together, these results show how ESC gene expression states and dynamics arise from a combination of intrinsic noise, coherent cellular states, and epigenetic regulation

INTRODUCTION

Findings

DISCUSSION

EXPERIMENTAL PROCEDURES