Please use this identifier to cite or link to this item: https://doi.org/10.1371/journal.pone.0004268
Title: Graded Smad2/3 activation is converted directly into levels of target gene expression in embryonic stem cells
Authors: Guzman-Ayala M.
Lee K.L. 
Mavrakis K.J.
Goggolidou P.
Norris D.P.
Episkopou V.
Keywords: 4 [4 (1,3 benzodioxol 5 yl) 5 (2 pyridinyl) 1h imidazol 2 yl]benzamide
Smad2 protein
Smad3 protein
Smad7 protein
1,3 dioxolane derivative
4 (5 benzo(1,3)dioxol 5 yl 4 pyridin 2 yl 1H imidazol 2 yl)benzamide
4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide
activin receptor 1
Acvr1b protein, mouse
Arkadia protein, mouse
benzamide derivative
Nodal protein, mouse
protein Nodal
Smad2 protein
Smad2 protein, mouse
Smad3 protein
Smad3 protein, mouse
transforming growth factor beta
ubiquitin
animal cell
animal tissue
article
controlled study
DNA microarray
embryo
embryo development
embryonic stem cell
feedback system
gene activation
gene expression profiling
gene identification
gene repression
genetic transcription
morphogenesis
mouse
nonhuman
nucleotide sequence
protein synthesis
signal transduction
transcription regulation
unindexed sequence
animal
biological model
flow cytometry
gene expression regulation
metabolism
Vertebrata
Activin Receptors, Type I
Animals
Benzamides
Dioxoles
Embryonic Stem Cells
Flow Cytometry
Gene Expression Regulation, Developmental
Mice
Models, Biological
Nodal Protein
Signal Transduction
Smad2 Protein
Smad3 Protein
Transcription, Genetic
Transforming Growth Factor beta
Ubiquitin
Issue Date: 2009
Citation: Guzman-Ayala M., Lee K.L., Mavrakis K.J., Goggolidou P., Norris D.P., Episkopou V. (2009). Graded Smad2/3 activation is converted directly into levels of target gene expression in embryonic stem cells. PLoS ONE 4 (1) : e4268. ScholarBank@NUS Repository. https://doi.org/10.1371/journal.pone.0004268
Abstract: The Transforming Growth Factor (TGF) ? signalling family includes morphogens, such as Nodal and Activin, with important functions in vertebrate development. The concentration of the morphogen is critical for fate decisions in the responding cells. Smad2 and Smad3 are effectors of the Nodal/Activin branch of TGF? signalling: they are activated by receptors, enter the nucleus and directly transcribe target genes. However, there have been no studies correlating levels of Smad2/3 activation with expression patterns of endogenous target genes in a developmental context over time. We used mouse Embryonic Stem (ES) cells to create a system whereby levels of activated Smad2/3 can be manipulated by an inducible constitutively active receptor (Alk4*) and an inhibitor (SB-431542) that blocks specifically Smad2/3 activation. The transcriptional responses were analysed by microarrays at different time points during activation and repression. We identified several genes that follow faithfully and reproducibly the Smad2/3 activation profile. Twenty-seven of these were novel and expressed in the early embryo downstream of Smad2/3 signalling. As they responded to Smad2/3 activation in the absence of protein synthesis, they were considered direct. These immediate responsive genes included negative intracellular feedback factors, like SnoN and I-Smad7, which inhibit the transcriptional activity of Smad2/3. However, their activation did not lead to subsequent repression of target genes over time, suggesting that this type of feedback is inefficient in ES cells or it is counteracted by mechanisms such as ubiquitin-mediated degradation by Arkadia. Here we present an ES cell system along with a database containing the expression profile of thousands of genes downstream of Smad2/3 activation patterns, in the presence or absence of protein synthesis. Furthermore, we identify primary target genes that follow proportionately and with high sensitivity changes in Smad2/3 levels over 15-30 hours. The above system and resource provide tools to study morphogen function in development. � 2009 Guzman-Ayala et al.
Source Title: PLoS ONE
URI: https://scholarbank.nus.edu.sg/handle/10635/161843
ISSN: 19326203
DOI: 10.1371/journal.pone.0004268
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