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Published on: February 13, 2021
Transcriptional analysis of cleft palate in TGFβ3 mutant mice
J Liu1, S K Chanumolu2, K M White3
1Department of Oral Biology, College of Dentistry, University of Nebraska Medical Center, Lincoln, NE, 68583, USA.
Insights
Transforming growth factor-beta 3 (Tgf-β3) deficiency causes cleft palate (CP) in mice. This study identified specific genes and pathways involved in CP development by comparing gene expression in wild-type and Tgf-β3 null mutant embryos.
Area of Science:
- Developmental Biology
- Genetics
- Molecular Biology
Background:
- Cleft palate (CP) is a common craniofacial birth defect.
- Transforming growth factor-beta 3 (Tgf-β3) is essential for normal palate development in mice, with null mutants exhibiting 100% penetrance of CP.
Purpose of the Study:
- To compare global palatal transcriptomes of wild-type (WT) and Tgf-β3 -/- homozygous (HM) mouse embryos at key palatogenesis stages (E14.5 and E16.5).
- To identify genes uniquely up/downregulated in WT or HM embryos to uncover genes associated with CP.
- To analyze systems biology pathways related to cell behaviors and functions of WT and HM specific genes.
Main Methods:
- RNA sequencing (RNA-seq) was used to analyze palatal transcriptomes at E14.5 and E16.5.
- Differential gene expression analysis was performed to identify significantly altered genes (adjusted p < 0.05; |fold change| > 2.0).
- Systems biology analysis and qRT-PCR were employed to validate gene expression and identify functional pathways.
Main Results:
- 1,809 and 2,127 differentially expressed genes were found between E14.5 and E16.5 in WT and HM groups, respectively.
- Systems biology analysis revealed non-Smad pathways and a shift from epithelial-mesenchymal transition to apoptosis in HM embryos.
- 24 HM-specific and 11 WT-specific genes related to CP and/or Tgf-β3 signaling were identified, with 29 validated by qRT-PCR.
Conclusions:
- The study identified novel genes and pathways involved in Tgf-β3-regulated palate development and cleft palate formation.
- Findings suggest a functional shift towards apoptosis over epithelial-mesenchymal transition in Tgf-β3 deficient palates.
- This research enhances understanding of the genetic regulation underlying cleft palate, directly and indirectly influenced by TGF-β signaling.
Abstract:
Cleft palate (CP) is one of the most common craniofacial birth defects, impacting about 1 in 800 births in the USA. Tgf-β3 plays a critical role in regulating murine palate development, and Tgf-β3 null mutants develop cleft palate with 100% penetrance. In this study, we compared global palatal transcriptomes of wild type (WT) and Tgf-β3 -/- homozygous (HM) mouse embryos at the crucial palatogenesis stages of E14.5, and E16.5, using RNA-seq data. We found 1,809 and 2,127 differentially expressed genes at E16.5 vs. E14.5 in the WT and HM groups, respectively (adjusted p < 0.05; |fold change|> 2.0). We focused on the genes that were uniquely up/downregulated in WT or HM at E16.5 vs. E14.5 to identify genes associated with CP. Systems biology analysis relating to cell behaviors and function of WT and HM specific genes identified functional non-Smad pathways and preference of apoptosis to epithelial-mesenchymal transition. We identified 24 HM specific and 11 WT specific genes that are CP-related and/or involved in Tgf-β3 signaling. We validated the expression of 29 of the 35 genes using qRT-PCR and the trend of mRNA expression is similar to that of RNA-seq data . Our results enrich our understanding of genes associated with CP that are directly or indirectly regulated via TGF-β.
