The TET-BMP regulatory axis in pathogenesis of CFM [RNA-Seq]
Craniofacial microsomia (CFM) is a congenital defect that usually results from aberrant development of embryonic pharyngeal arches. However, the molecular basis of CFM pathogenesis is largely unknown. Here we employ zebrafish model to investigate the mechanism of CFM pathogenesis. In early embryos, tet2 and tet3 are highly expressed and are essential for pharyngeal cartilage development. Single-cell RNA sequencing and genetic analyses reveal that loss of Tet2/3 impaired chondrocyte differentiation largely due to insufficient BMP signaling. Mechanistically, Tet2/3-mediated 5-hydroxymethylcytosine modification allows the 5-hydroxymethylcytosine “reader”, Sall4, to specifically bind the bmp4 promoter, thereby promoting bmp4 expression and enabling efficient BMP signaling. These findings indicate the TET-BMP regulatory axis via 5-hydroxymethylcytosine to be critical for pharyngeal cartilage development. Whole-exome sequencing of CFM patient samples show that single nucleotide polymorphisms in TET and BMP pathway genes increase the risk of CFM. Collectively, our study provides novel insights into understanding craniofacial development and CFM pathogenesis. Overall design: Total RNAs were purified using TRIzol reagent (Life Technologies, 343702). Libraries were constructed using a VAHTS Universal V8 RNA-seq Library Prep Kit for MGI (Vazyme) according to the manufacturer's instructions and then sequenced on the MGISEQ-2000 platform.