Core microRNAs regulate neural crest delamination and condensation in the developing trigeminal ganglion

Neural crest cells (NCC) are crucial contributors to embryonic development, found in various regions like cranial, vagal, truncal, and sacral areas. Their ability to undergo an epithelial-mesenchymal transition (EMT) enables migration and formation of derivatives, such as the trigeminal ganglion (TG). Its formation involves the condensation and differentiation of cranial NCC, which occurs through a process similar to the mesenchymal-epithelial transition (MET). The regulation of these transitions is complex and finely controlled by regulatory networks. MicroRNAs (miRNAs) have been shown to play a key role in this process, particularly studied during EMT. However, there have been few studies related to the miRNA contribution during MET. Therefore, we aimed to identify and characterize the key miRNAs involved in delamination and condensation of the NCC during TG formation. We analyzed cranial NCCs during pre-migratory, migratory, and condensation stages using microRNA sequencing. Through comparative analysis between stages, we identified two groups of miRNAs that were upregulated during delamination (EMT-miRNAs: miR-140-3p, miR-140-5p, and miR-455-5p) and condensation (MET-miRNAs: miR-23b-3p, miR-187-3p and miR-363-5p). We then evaluated their functionality by in vivo loss-of-function experiments demonstrating their participation in delamination and ganglia formation. Additionally, we identified potential targets of microRNAs involved in neural crest EMT and MET by performing RNA sequencing on embryos treated with microRNA inhibitors. The expressing of MET-miRNAs in trunk NCCs from GFP-transgenic chick, followed by transplantation to the cranial region of a host embryo, enhanced the ability of trunk NCC to condensate and integrate into the trigeminal ganglia. From this, we can conclude that intrinsic miRNA abundance plays a key role in NCC delamination and condensation, shedding light on their regulatory mechanisms in trigeminal ganglion formation.