A new method developed by researchers at the Institute in collaboration with researchers in the UK and Belgium makes it possible to study the epigenome and transcriptome of a single cell at the same time. The protocol, published in Nature Methods, helps scientists pinpoint the relationship between changes in DNA methylation and gene expression. The method is potentially transformative for epigenetics research, as it reveals unprecedented detail of the epigenetic control of genes.
The work at the Institute brought together two of the groups in the Institute’s Epigenetics research programme, led by group leaders Wolf Reik and Gavin Kelsey, and particularly utilised the Institute’s expertise in single-cell methods. Single-cell sequencing technology has progressed rapidly in recent years, and is widely used to study how gene expression profiles (‘transcriptomes’) vary between cells. Recent single-cell protocols also allow researchers to explore chemical modification of DNA (‘epigenetics’), for example DNA methylation, which is a driving force behind changes to gene expression. Until now, it has only been possible to study single-cell transcriptomes and epigenomes separately.
“This new experimental protocol lets you assay both DNA methylation and RNA of the same single cell in parallel,” says Oliver Stegle from the European Bioinformatics Institute (EMBL-EBI). “Our approach provides the first direct view on the relationship between heterogeneity in DNA methylation and variation of expression in specific genes across single cells.”
“The new advance allows us to examine the rules of gene expression in very rare or difficult to obtain cells, such as in the very early embryo,” Gavin Kelsey from the Babraham Institute goes on to explain.
To test the method, the group used mouse embryonic stem cells (ESCs) at a stage when they switch continuously between different gene-expression states. Just as in the cells of an early stage embryo, the identity of these cells is fluid rather than fixed. The researchers used two techniques in parallel: one that reveals detailed information about expression (how much variation there is, where that heterogeneity is coming from) and one to study DNA methylation in the same cells. For each cell, they obtained sufficient coverage to study epigenetic and transcriptome diversity of several thousand genes.
“The epigenetic state of ESCs is highly variable, and this variation is associated with changes in gene expression,” explains Wolf Reik of the Babraham Institute and Wellcome Trust Sanger Institute. “Much of the transcriptional variability we see is thought to be driven by modifications of DNA, but now we have a technique that allows us to look at a single cell and discover relationships between DNA methylation and gene expression that were previously unknown. To understand development, it is really important that we pin these relationships down, and get them right.”
“The technique has provided a big step forward in terms of being able to analyse what’s going on in a single cell, rather than making generalised observations from a population of diverse cells, “ says joint-first author Heather Lee from the Babraham Institute. “We’ve used this to uncover hundreds of associations between the epigenome and transcriptome which relate to the regulation of pluripotency in mouse embryonic stem cells,” continues joint-first author Stephen Clark, also of the Babraham Institute. “This information will help further our understanding of how cell differentiation is regulated.”
Going forward, the researchers expect the new protocol to offer new opportunities to study multiple different molecular layers simultaneously. This will go a long way towards understanding the connection between gene expression and DNA methylation in single cells, and identifying the factors that influence this relationship. Such research has implications for understanding normal development, and changes that occur with ageing and cancer.
The method was developed within the Sanger Institute/EMBL-EBI Single Cell Genomics Centre, a collaborative effort to develop single-cell technologies and apply them to new biological questions.
The new method allows integration of two types of data (methylation and gene expression) from a single cell. Image created by Spencer Phillips, EMBL-EBI.
Christof Angermueller (EMBL-EBI), Stephen Clark (Babraham Institute), Heather Lee (Babraham Institute) and Iain Macaulay (Wellcome Trust Sanger Institute) are co-first authors on this work.
Angermueller et al. (2016) Parallel single-cell sequencing links transcriptional and epigenetic heterogeneity. Nature Methods.
Stephen Clark, postdoc researcher (Kelsey and Reik labs)
Heather Lee, postdoc researcher (Reik lab)
Sebastien Smallwood, previous postdoc researcher (Kelsey lab)Gavin Kelsey, group leader, Epigenetics programmeWolf Reik, Head of Epigenetics programme and group leader at the Babraham Institute, and associated faculty at the Wellcome Trust Sanger Institute
As a publicly funded research institute, the Babraham Institute is committed to engagement and transparency in all aspects of its research. The research presented here used mice to obtain embryonic stem cells.
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11 January 2016