Background: Transforming growth factor-β (TGF-β)/nodal signaling is
involved in early embryonic patterning in vertebrates. Nodal modulator (Nomo, also
called pM5) is a negative regulator of nodal signaling. Currently, the role of nomo gene
in cartilage development in vertebrates remains unknown.
Methods: Nomo mutants were generated in a knockout model of zebrafish by clustered
regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9
(CRISPR/Cas9) targeting of the fibronectin type III domain. The expression of related
genes, which are critical for chondrogenesis, was analyzed by whole-mount in situ
hybridization and qRT-PCR. Whole-mount alcian staining was performed to analyze the
Results: nomo is highly expressed in various tissues including the cartilage. We
successfully constructed a zebrafish nomo knockout model. nomo homozygous mutants
exhibited varying degrees of hypoplasia and dysmorphism on 4 and 5 dpf, which is
similar to chondrodysplasia in humans. The key genes of cartilage and skeletal
development, including sox9a, sox9b, dlx1a, dlx2a, osx, col10a1, and col11a2 were all
downregulated in nomo mutants compared with the wildtype.
Conclusion: The nomo gene positively regulates the expression of the master regulator
and other key development genes involved in bone formation and cartilage development
and it is essential for cartilage development in zebrafish.