The transcription factor E2A drives neural differentiation in pluripotent cells.

Chandrika Rao,Sally Lowell,Mattias Malaguti,John O Mason

doi:10.1242/dev.184093

Chandrika Rao, Sally Lowell + Show 2 more

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https://doi.org/10.1242/dev.184093

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Abstract

ABSTRACTThe intrinsic mechanisms that link extracellular signalling to the onset of neural differentiation are not well understood. In pluripotent mouse cells, BMP blocks entry into the neural lineage via transcriptional upregulation of inhibitor of differentiation (Id) factors. We have previously identified the major binding partner of Id proteins in pluripotent cells as the basic helix-loop-helix (bHLH) transcription factor (TF) E2A. Id1 can prevent E2A from forming heterodimers with bHLH TFs or from forming homodimers. Here, we show that overexpression of a forced E2A homodimer is sufficient to drive robust neural commitment in pluripotent cells, even under non-permissive conditions. Conversely, we find that E2A null cells display a defect in their neural differentiation capacity. E2A acts as an upstream activator of neural lineage genes, including Sox1 and Foxd4, and as a repressor of Nodal signalling. Our results suggest a crucial role for E2A in establishing neural lineage commitment in pluripotent cells.

Highlights

Following the establishment of the primary germ layers during gastrulation, the embryonic neural plate is specified from the anterior ectoderm at approximately embryonic day (E)7.5 in a process known as neural induction (Tam and Zhou, 1996)
E2A is expressed heterogeneously in pluripotent embryonic stem cells (ESCs) and throughout neural differentiation To characterise the expression of E2A during early neural differentiation we first examined the temporal expression of E2A mRNA in ESCs, plated under standard neural monolayer conditions (Ying et al, 2003b), by qRT-PCR
As E2A is regulated by Id1 at the protein level, rather than the transcriptional level, we generated an endogenously tagged ESC line (Fig. 1B) using CRISPR/Cas9 targeting to follow the expression of E2A protein during differentiation

Summary

Introduction

Following the establishment of the primary germ layers during gastrulation, the embryonic neural plate is specified from the anterior ectoderm at approximately embryonic day (E)7.5 in a process known as neural induction (Tam and Zhou, 1996). Pluripotent embryonic stem cells (ESCs) recapitulate central features of this process when differentiated in culture (Ying et al, 2003b), providing a useful system in which to study the mechanisms guiding these initial cell fate decisions during development. The key extracellular signalling pathways that inhibit neural lineage commitment have long been established, less progress has been made in identifying the downstream effectors of these pathways. Inhibition of BMP signalling is crucial for the establishment of the neuroectoderm (Harland, 2000; Di-Gregorio et al, 2007). The finding that BMP signalling inhibits neural differentiation via transcriptional upregulation of Id1 (Ying et al, 2003a) provides compelling evidence that Id proteins block.

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