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RNAseq Analysis of Transcriptome expression profiles in E12.5 and E13.5 FACS sorted Osr2RFP/- (mutant)and Osr2RFP/+ (control) palatal mesenchymal cells

ABSTRACT: Purpose: The purpose of this study is to compare the transcriptome expression profiles of E12.5 and E13.5 Osr2RFP/- and Osr2RFP/+ palatal mesenchyme by using RNA-seq analysis. Methods: Osr2RFP/+ male mice were crossed with Osr2+/- female mice. The embryos were harvested at E12.5 and E13.5. The pair of palatal shelves were dissected from each Osr2-RFP positive embryo. The RFP+ palatal mesenchyme cells were isolated by using fluorescence-activated cell sorting (FACS). RNA-seq analysis was carried out using the FACS-isolated palatal mesenchyme from Osr2RFP/- and Osr2RFP/+ embryos, respectively.

ORGANISM(S): Mus musculus

PROVIDER: GSE95638 | GEO | 2018/02/13

REPOSITORIES: GEO

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Project description:We identify a role for two evolutionarily related, secreted metalloproteases of the ADAMTS family (A disintegrin-like and metalloprotease domain with thrombospondin type-1 motif), ADAMTS20 and ADAMTS9, in palatogenesis. Adamts20 mutations cause the mouse white spotting mutant belted (bt), whereas Adamts9 is essential for survival beyond 7.5 days of gestation (E7.5). Functional overlap of Adamts9 with Adamts20 was established in bt/bt:Adamts9+/- mice, which have increased white spotting relative to bt mice, as previously reported, and a fully penetrant cleft palate. Palatal closure was delayed, although eventually completed, in both bt/+;Adamts9+/- and bt/bt mice, demonstrating a cooperative role of these related genes. Adamts9 and Adamts20 are both expressed in palatal mesenchyme, with Adamts9 expressed exclusively in microvascular endothelial cells. Palatal shelves from bt/bt:Adamts9+/- mice fused in culture, suggesting an intact TGF signaling pathway in palatal epithelium, and indicating a temporally specific delay in palatal shelf elevation and growth toward the midline. Palatal shelf mesenchymal cells showed a statistically significant decrease of cell proliferation at E13.5 and E14.5, as well as decreased processing of versican, an ADAMTS substrate, at these stages. Vcan haploinsufficiency led to a greater penetrance of cleft palate in bt mice, and impaired proliferation was also seen in palatal mesenchymal cells of these mice, suggesting a role for ADAMTS-mediated versican proteolysis in palatal closure. In a parallel with recent work identifying a role for a bioactive ADAMTS-generated versican fragment in regulating apoptosis during interdigital web regression, we propose that versican proteolysis may influence palatal mesenchymal cell proliferation. Palatal shelves were dissected from four E13.75 Adamts9+/-:bt/bt embyos (correspond to the 4 samples: Palate_Adamts9+/-:bt/bt_Rep1, Palate_Adamts9+/-:bt/bt_Rep2, Palate_Adamts9+/-:bt/bt_Rep3 and Palate_Adamts9+/-:bt/bt_Rep4) and age-matched 3 wild-type C57Bl/6 embryos (correspond to the 3 samples: Palate_WT_Rep1, Palate_WT_Rep2, and Palate_WT_Rep3) that were used as the controls

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RNAseq Analysis of Transcriptome expression profiles in E12.5 and E13.5 FACS sorted Osr2RFP/- (mutant)and Osr2RFP/+ (control) palatal mesenchymal cells

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