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Cardiac Differentiation of ESCs - A New Role for REST

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Review of “A role for REST in embryonic stem cells cardiac lineage specification” from Stem Cells by Stuart P. Atkinson

The repressor element 1 (RE-1)-silencing transcription factor (REST) is a jack of all trades; well-known for its role in regulating neurogenesis [1, 2], but also lends a hand in a range of other biological processes, including balancing embryonic stem cell (ESC) pluripotency and differentiation, and maintaining normal function in the adult heart. 

However, a new study from the laboratory of Lawrence W. Stanton (Genome Institute of Singapore) has suggested that REST may play a role in the heart at an earlier stage – during early cardiac differentiation from ESCs. Their study, published in Stem Cells [3], also identifies how REST functions: by transcriptionally activating Gata4, a major cardiac transcription factor, in addition to regulating Wnt/β-catenin signaling.

Initial gene expression analysis of REST knockout (KO) mouse ESCs uncovered no alteration in pluripotency gene expression but did find an endoderm-specific decrease in differentiation genes (Sox7, Sox17, Gata4, Gata6, and FoxA2). This analysis also suggested that REST regulated the formation of primitive endoderm (PrE), and not definitive endoderm (DE), which can give rise to cardiac mesoderm though the production of visceral endoderm. 

Following this line of investigation, the authors demonstrated that REST-KO ESCs undergoing directed cardiac differentiation produced less areas of beating tissue (indicative of cardiomyocyte production) than wild-type ESCs and expressed lower levels of cardiac marker genes, including early cardiac lineage markers (Gata4 and Mesp1). Further examination of wild-type differentiated cells showed that most differentiated cells, such as cTnT-(cardiomyocytes) or Flk1- (mesoderm) expressing cells, also expressed REST, suggesting a general importance of this transcription factor in fully differentiated cardiac cells.

The authors next sought to understand just how REST is important to the cardiac lineage and focused their search on the Wnt/β-catenin signaling pathway and Gata4. This found altered expression in 7 of the 23 Wnt family genes with regulatory regions bound by REST, while inhibition of Wnt signaling using the small molecule XAV-939 rescued some of the cardiac differentiation ability of REST-KO ESCs. Gata4 is another important cardiac regulator and its overexpression also overcame the cardiac differentiation impairment observed with REST-KO ESCs, with subsequent chromatin immunoprecipitation assays suggesting that REST may act as a co-activator to control the expression of Gata4 expression.

This study highlights an important role for REST in cardiac cells produced via the differentiation of ESCs - a novel function given the strong links between REST and differentiation towards a neural lineage. A better understanding of how REST regulates Wnt gene expression and signaling as well as Gata4 expression (See figure) may allow us to generate improved protocols for the production of cardiac tissues in humans if this regulatory mechanism proves to be conserved.

References

  1. Ballas N, Grunseich C, Lu DD, et al. REST and its corepressors mediate plasticity of neuronal gene chromatin throughout neurogenesis. Cell 2005;121:645-657.
  2. Chen ZF, Paquette AJ, and Anderson DJ NRSF/REST is required in vivo for repression of multiple neuronal target genes during embryogenesis. Nat Genet 1998;20:136-142.
  3. Aksoy I, Marcy G, Chen J, et al. A Role for RE-1-Silencing Transcription Factor in Embryonic Stem Cells Cardiac Lineage Specification. STEM CELLS 2016;34:860-872.