Abstract
The transcription factor brachyury (T, BRA) is one of the first markers of gastrulation and lineage specification in vertebrates. Despite its wide use and importance in stem cell and developmental biology, its functional genomic targets in human cells are largely unknown. Here, we use differentiating human embryonic stem cells to study the role of BRA in activin A-induced endoderm and BMP4-induced mesoderm progenitors. We show that BRA has distinct genome-wide binding landscapes in these two cell populations, and that BRA interacts and collaborates with SMAD1 or SMAD2/3 signalling to regulate the expression of its target genes in a cell-specific manner. Importantly, by manipulating the levels of BRA in cells exposed to different signalling environments, we demonstrate that BRA is essential for mesoderm but not for endoderm formation. Together, our data illuminate the function of BRA in the context of human embryonic development and show that the regulatory role of BRA is context dependent. Our study reinforces the importance of analysing the functions of a transcription factor in different cellular and signalling environments.
Highlights
The three primary germ layers arise from the pluripotent epiblast during gastrulation in the amniote embryo (Arnold and Robertson, 2009; Tam and Loebel, 2007); this can be modelled in vitro using pluripotent stem cells (Murry and Keller, 2008)
Brachyury is widely used as the earliest marker of mesodermal and endodermal differentiation in embryonic stem cell (ESC) studies and during gastrulation, because both these cell lineages derive from the primitive streak (Murry and Keller, 2008)
An in vitro differentiation system to study the role of BRA in human gastrulation We have previously optimised chemically defined conditions that cause Human embryonic stem cells (hESCs) to differentiate as progenitors of endoderm or of mesoderm (Bernardo et al, 2011)
Summary
The three primary germ layers (ectoderm, mesoderm and endoderm) arise from the pluripotent epiblast during gastrulation in the amniote embryo (Arnold and Robertson, 2009; Tam and Loebel, 2007); this can be modelled in vitro using pluripotent stem cells (Murry and Keller, 2008). Mice lacking one copy of the gene have a short tail, while homozygous embryos die around embryonic day (E) 9-10 (Chesley, 1935). The latter develop only the first seven somites and lack a proper notochord; in addition, they display defects in left-right asymmetry and in cell migration (Showell et al, 2004). Brachyury is widely used as the earliest marker of mesodermal and endodermal differentiation in embryonic stem cell (ESC) studies and during gastrulation, because both these cell lineages derive from the primitive streak (Murry and Keller, 2008). On a molecular level, the role of BRA in endoderm formation remains poorly understood
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