Characterization of a Novel Zebrafish Mutant Lacking all Craniofacial Cartilages

Cranial Neural Crest Cells (NCCs), a derivative of the neural crest, are responsible for several structures including most of the cartilage in the head and connective tissues.  The zebrafish mutant, ghost, lacks all craniofacial cartilages, which is not due to the loss of NCCs.  The microenvironment through which NCC’s migrate influence the fate of these multipotent cells.  Defects in cell layers would affect the signaling processes that influence cell specification, migration and differentiation, and ultimately, the anatomical structures of these precursors.  We hypothesize that the phenotype may be a result of defects in the midline signals and the microenvironment of the NCC’s migratory pathway. Objectives: The purpose of this study is to analyze the midline patterning in the ghost mutant and determine changes in cell signals that may result in loss of cartilage.  Methods: Whole mount in situ hybridization and immunohistochemistry were used to examine specific midline signals that participate in tissue patterning and examine the expression level of key signaling molecules through quantitative RT-PCR. Results: Defects were found in col2a1 and axial gene expression throughout several stages of development, indicating a decrease in mesenchymal cells and a mis-patterned pharyngeal endoderm. Decreased expression of hedgehog signals were also observed in the ghost mutants. Conclusions: These data suggest that the early patterning of the brain tissue is altered in the ghost mutant as a result of defective hedgehog signaling.  These findings suggest that the mutation lies in a gene involved in hedgehog signaling pathways. Supported by NIDCR grant R15-DE-019723-01 and UMD Healey Foundation Award to TLF.