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| Status |
Public on Apr 16, 2013 |
| Title |
Spatial Transcriptional Profile of the Chick and Mouse Endocardial Cushions Identify Novel Regulators of Endocardial EMT in vitro |
| Organisms |
Gallus gallus; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Objective: We developed an unbiased strategy to identify genes important in endocardial epithelial-to-mesenchymal transformation (EMT) using a spatial transcriptional profile.
Methods and Results: Endocardial cells overlaying the cushions of the atrioventricular canal (AVC) and outflow tract (OFT) undergo an EMT to yield VICs. RNA sequencing (RNA-seq) analysis of gene expression between AVC, OFT, and ventricles (VEN) isolated from chick and mouse embryos at comparable stages of development (chick HH18; mouse E11.0) was performed. EMT occurs in the AVC and OFT cushions, but not VEN at this time. 210 genes in the chick (n=1) and 105 genes in the mouse (n=2) were enriched 2-fold in the cushions. Gene regulatory networks (GRN) generated from cushion-enriched gene lists confirmed TGFβ as a nodal point and identified NF-κB as a potential node. To reveal previously unrecognized regulators of EMT four candidate genes, Hapln1, Id1, Foxp2, and Meis2, and a candidate pathway, NF-κB, were selected. In vivo spatial expression of each gene was confirmed by in situ hybridization and a functional role for each in endocardial EMT was determined by siRNA knockdown in a collagen gel assay.
Conclusions: Our spatial-transcriptional profiling strategy yielded gene lists which reflected the known biology of the system. Further analysis accurately identified and validated previously unrecognized novel candidate genes and the NF-κB pathway as regulators of endocardial cell EMT in vitro.
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| Overall design |
3 separate regions of the developing heart tube were dissected and processed for RNA sequencing analysis in both HH18 chick and E11.0 ICR mouse (done in duplicate)
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| Contributor(s) |
DeLaughter DM, Christodoulou DC, Robinson JY, Seidman CE, Baldwin H, Seidman JG, Barnett JV |
| Citation(s) |
23557753 |
| Submission date |
Dec 28, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Daniel DeLaughter |
| Organization name |
Vanderbilt University Medical School
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| Department |
Cell and Developmental Biology
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| Lab |
Scott Baldwin Laboratory
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| Street address |
Preston Research Building Room 460
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| City |
Nashville |
| State/province |
TN |
| ZIP/Postal code |
37203 |
| Country |
USA |
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| Platforms (2) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
| GPL16133 |
Illumina HiSeq 2000 (Gallus gallus) |
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| Samples (9)
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| Relations |
| BioProject |
PRJNA184872 |
| SRA |
SRP017709 |
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