Notch signalling regulates epibranchial placode patterning and segregation

Abstract

Epibranchial placodes are the geniculate, petrosal and nodose placodes that generate parts of cranial nerves VII, IX and X, respectively. How the three spatially separated placodes are derived from the common posterior placodal area is poorly understood. Here, we reveal that the broad posterior placode area is first patterned into a Vgll2 + /Irx5 + rostral domain and a Sox2 + /Fgf3 + / Etv5 + caudal domain relative to the first pharyngeal cleft. This initial rostral and caudal patterning is then sequentially repeated along each pharyngeal cleft for each epibranchial placode. The caudal domains give rise to the neuronal and non-neuronal cells in the placode, whereas the rostral domains are previously unrecognized structures, serving as spacers between the final placodes. Notch signalling regulates the balance between the rostral and caudal domains: high levels of Notch signalling expand the caudal domain at the expense of the rostral domain, whereas loss of Notch signalling produces the converse phenotype. Collectively, these data unravel a new patterning principle for the early phases of epibranchial placode development and a role for Notch signalling in orchestrating epibranchial placode segregation and differentiation.

Hoxb3 Regulates Jag1 Expression in Pharyngeal Epithelium and Affects Interaction With Neural Crest Cells

“…We have previously demonstrated that in the developing PAs, the proximal pharyngeal epithelium was derived from the posterior placodal area and overlap with the epibranchial placodal cells ( Figure 1I , red dotted line; Zhang et al, 2017 ; Wang et al, 2020 ), while the distal PAs were enveloped by pharyngeal ectodermal epithelium. Therefore, the proximal and distal pharyngeal epithelium were composed of developmentally distinct epithelial cells.…”

“…As Notch signaling has been shown to be critical for the development of epibranchial placodal epithelium and proximal PAs ( Zhang et al, 2017 ; Wang et al, 2020 ), we examined Jag1 which was expressed in the proximal pharyngeal epithelium of developing PAs and restricted to the pharyngeal clefts and pouches as the segmental plates were formed at E9.5 ( Figures 2G,I,K ). Interestingly, we found that the expression of Jag1 was ectopically activated in the entire pharyngeal ectodermal region of PA2 and posterior PA1 in the Hoxb3 Tg/+ mutants at E9.5 ( Figures 2H,J ).…”

“…We have previously shown that over-expression of the Notch receptor protein N1-ICD in the epibranchial placodal epithelium would lead to misexpression of Fgf ligands and cell fate changes ( Wang et al, 2020 ). Therefore, we examined the expression of Fgf3 and Fgf8 , which marked placodal epithelial cells flanking the pharyngeal clefts ( Figures 2M,O ).…”

“…Defective pharyngeal segmentation as displayed in the Sox3 ( Rizzoti and Lovell-Badge, 2007 ) and Eya1 mutants ( Xu et al, 2002 ; Zhang et al, 2017 ) would lead to abnormal PA development. Around the pharyngeal clefts, the non-neuronal epibranchial placodal epithelial cells express various signaling factors including Notch and Fgfs that are required for PA morphogenesis ( Trokovic et al, 2005 ; Wang et al, 2020 ). The roles of the ectodermal pharyngeal epithelium in interacting with neural crest and other cell types in the developing PAs are not well understood.…”

“…In our previous studies, we showed that Notch1 intracellular domain (N1-ICD) was required in the epibranchial placodal epithelium in the proximal PAs to control cell fate commitment of neuronal and non-neuronal epibranchial placodal cells ( Zhang et al, 2017 ). Moreover, Notch receptor signaling controlled rostral-caudal patterning of the non-neuronal placodal epithelial cells around the pharyngeal clefts, indicating important roles of Notch signaling in epithelial cell fate commitment and differentiation ( Wang et al, 2020 ).…”