Ciliopathies are genetic syndromes that link skeletal dysplasias to dysfunction of primary cilia. Primary cilia are sensory organelles synthesized by intraflagellar transport (IFT) - A and B complexes, which traffic protein cargo along a microtubular core. We have reported that deletion of IFT-A gene, Thm2, together with a null allele of its paralog, Thm1, causes a small skeleton with small mandible or micrognathia in juvenile mice. Using micro-computed tomography, here we quantify the craniofacial defects of Thm2-/-;Thm1aln/- triple allele mutant mice. At postnatal day 14, triple allele mutant mice exhibit micrognathia, maxillary hypoplasia, and a decreased facial angle due to shortened maxilla, premaxilla, and nasal bones, reflecting altered development of facial anterior-posterior elements. In contrast, other ciliopathy-related craniofacial defects, such as cleft lip and/or palate, hypo-/hypertelorism, broad nasal bridge, craniosynostosis, and facial asymmetry, were not observed, suggesting development of the facial transverse dimension is intact. Calvarial-derived osteoblasts of triple allele mutant mice showed reduced bone formation in vitro that was ameliorated by Hedgehog agonist, SAG. Together, these data indicate that Thm2 and Thm1 genetically interact to regulate bone formation and sculpting of the postnatal face. The triple allele mutant mice present as a novel model to study craniofacial bone development.

Orofacial clefts are among the most common congenital malformations, affecting approximately 1 in 700 births. Mutations in SPECC1L have been identified in patients with syndromic and nonsyndromic cleft lip and/or palate. Syndromic SPECC1Lmutations cluster in the second coiled coil domain (CCD2), which facilitates interaction of this cytoskeletal protein with microtubules. Our recent studies indicate that mice with an in‐frame deletion of SPECC1L‐CCD2 exhibit cleft palate and exencephaly in a dominant‐negative manner. Notably, orofacial clefts and exencephaly are common manifestations of ciliopathies – human disorders which arise from disruption of cilia – but an association between SPECC1L and cilia has not been reported. We investigated cilia in E13.5 Specc1l∆CCD2/∆CCD2 mutant palatal shelves. SPECC1L localizes to the cytoplasm and around the cilia base in wild‐type palatal mesenchyme, but this localization is diminished in Specc1l∆CCD2/∆CCD2 palatal mesenchyme. In addition, primary cilia lengths are decreased in Specc1l∆CCD2/∆CCD2 palatal mesenchyme and epithelium relative to wild‐type, on both the lingual and buccal sides of the palate. We hypothesize that an intracellular trafficking defect in the palatal mesenchyme results in shortened cilia and that the shortened cilia affects signaling in the palatal mesenchyme. Since the IFT‐A mouse mutant, Thm1aln/aln, shares overlapping phenotypes with Specc1l mutant mice, including cleft palate, exencephaly, and shortened cilia, we crossed the two mutants to look for a genetic interaction. Indeed, ~30% of double heterozygous Specc1l∆CCD2/+;Thm1aln/+ mice show cleft palate. Interestingly, none of the double heterozygous mice showed exencephaly, indicating a palate specificity of the genetic interaction. Previous studies have shown that conditional loss of Thm1 in neural crest cells (NCCs), which give rise to the palatal mesenchyme, results in cleft palate, and that Specc1l deficiency leads to NCC delamination defect. Our recent analysis of Specc1l deficient embryos demonstrates that cilia are shorter in mutant NCC even compared with surrounding non‐NCCs. While SPECC1L is broadly expressed during early craniofacial development, its expression is stronger in migrating NCCs. Thus, we propose that the genetic interaction between Specc1l and Thm1 initiates in the NCC. Overall, our studies indicate a novel role for SPECC1L in ciliogenesis and NCC function.