The cellular and molecular etiology of the craniofacial defects in the avian ciliopathic mutant talpid2

Abstract

talpid2 is an avian autosomal recessive mutant with a myriad of congenital malformations, including polydactyly and facial clefting. Although phenotypically similar to talpid3, talpid2 has a distinct facial phenotype and an unknown cellular, molecular and genetic basis. We set out to determine the etiology of the craniofacial phenotype of this mutant. We confirmed that primary cilia were disrupted in talpid2 mutants. Molecularly, we found disruptions in Hedgehog signaling. Post-translational processing of GLI2 and GLI3 was aberrant in the developing facial prominences. Although both GLI2 and GLI3 processing were disrupted in talpid2 mutants, only GLI3 activator levels were significantly altered in the nucleus. Through additional fine mapping and whole-genome sequencing, we determined that the talpid2 phenotype was linked to a 1.4 Mb region on GGA1q that contained the gene encoding the ciliary protein C2CD3. We cloned the avian ortholog of C2CD3 and found its expression was ubiquitous, but most robust in the developing limbs and facial prominences. Furthermore, we found that C2CD3 is localized proximal to the ciliary axoneme and is important for docking the mother centriole to the ciliary vesicle and cell membrane. Finally, we identified a 19 bp deletion in talpid2 C2CD3 that produces a premature stop codon, and thus a truncated protein, as the likely causal allele for the phenotype. Together, these data provide insight into the cellular, molecular and genetic etiology of the talpid2 phenotype. Our data suggest that, although the talpid2 and talpid3 mutations affect a common ciliogenesis pathway, they are caused by mutations in different ciliary proteins that result in differences in craniofacial phenotype.

Primary Cilium-Mediated Crosstalk of Signaling Cascades in Ciliogenesis: Implications for Tumorigenesis and Senescence

“…Finally, further axoneme growth, docking, and fusing of the ciliary sheath to the plasma membrane, and subsequent axoneme outgrowth occur to complete ciliary assembly. 133 the extracellular pathway is characterized in madin-Darby canine kidney cells, the mouse inner medullar collecting duct 3 cell line, and other epithelial cells. in this pathway, axoneme growth begins after direct binding of the mother centriole to the plasma membrane.…”

The Ciliary Baton

“…In addition, two separate, naturally occurring ciliopathic models have been identified in the chicken. The causal genetic elements for both the talpid 2 and talpid 3 mutants have recently been identified (Chang et al, 2014;Yin et al, 2009). The accessibility of the chicken embryo, and its availability for culturing and transplanting tissues, provides promise for careful analysis of how primary cilia participate in each individual step of NCC development.…”

“…OFD has long been linked to mutations in OFD1 (Ferrante et al, 2001;Gurrieri, Franco, Toriello, & Neri, 2007). Recently however, a new -Robinet et al, 2014); the same gene recently found responsible for the talpid 2 avian mutant (Chang et al, 2014). The rapid identification of new ciliary genes and ciliopathic models suggests that many yet to be classified syndromes may in fact be ciliopathies.…”

“…The Ciliary Baton: Orchestrating Neural Crest Cell Development A curious example of a ciliopathy causing dental abnormalities is the avian ciliopathic mutant, talpid 2 , which develops crocodilian-like teeth (Brugmann et al, 2010;Chang et al, 2014;Harris, Hasso, Ferguson, & Fallon, 2006). Modern birds lack teeth.…”

“…Numerous animal ciliopathic models have glossal defects. The avian talpid 2 mutant exhibits hypoglossia/aglossia (Chang et al, 2014) and both the Kif3a f/f ;Wnt1-Cre and Ift88 f/f ;Wnt1-Cre mouse mutants display aglossia (Brugmann et al, 2010;Fig. 3).…”