Pioneering function of Yap and Taz in promoting osteogenesis of neural crest cells by preventing chondrogenesis

Neural crest cells (NCCs) are multipotent stem cells with a remarkable ability to differentiate into multiple cell lineages, including osteoblasts and chondrocytes. NCCs contribute to the majority of craniofacial skeleton, yet the molecular mechanisms regulating NCCs diversification into osteoblasts or chondrocytes remain poorly understood. We found that Yap and Taz function redundantly as key determinants of the osteogenesis versus chondrogenesis fate decision and differentiation in NCCs in vitro, ex vivo and in vivo, and Yap/Taz-deficiency in NCCs resulted in a switch from osteogenesis to chondrogenesis. Comprehensive analysis of unbiased datasets including CUT&RUN-seq and RNA-seq indicated that Yap/Taz directly regulate key genes that govern osteogenesis and chondrogenesis. During NCC-derived osteogenesis, Yap/Taz promote expression of osteogenic genes such as Runx2 and Sp7 but repress expression of chondrogenic genes such as Sox9 and Col2a1. Further, we found Yap/Taz directly interact with ß-catenin in NCCs to coordinately promote osteoblast differentiation and repress chondrogenesis. Together our data indicate that Yap/Taz promote osteogenesis in NCCs by preventing chondrogenesis, partly through interactions with the Wnt-ß-catenin pathway. Overall design: 1. We performed CUT&RUN in O9-1 neural crest cells using Yap, IGG and H3K27me3 antibodies, followed by sequencing. 2. O9-1 cells were cultured under conditioned basal medium. Then the cells were seperated into 2 group: one group used for Yap/Taz dKD and the other group used as control. For the siRNA-mediated KD of Yap and Taz, O9-1 cells were plated at 50% confluency in the conditioned basal media and transfected with siRNA SMART pools of Yap (Dharmacon, L-046247-01-0005), Taz (Dharmacon, L-041057-01), and nontargeting siRNA (Dharmacon, D-001810-04-05) for 48 h according to the guidelines of the RNAiMAX transfection procedure (Life Technologies, 13778075) and cultured to 80% confluency. Then the cells were harvested and used for CUT&Tag experiment.