Mutations in interferon regulatory factor 6 (IRF6) account for ∼70% of cases of Van der Woude syndrome (VWS), the most common syndromic form of cleft lip and palate. In 8 of 45 VWS-affected families lacking a mutation in IRF6, we found coding mutations in grainyhead-like 3 (GRHL3). According to a zebrafish-based assay, the disease-associated GRHL3 mutations abrogated periderm development and were consistent with a dominant-negative effect, in contrast to haploinsufficiency seen in most VWS cases caused by IRF6 mutations. In mouse, all embryos lacking Grhl3 exhibited abnormal oral periderm and 17% developed a cleft palate. Analysis of the oral phenotype of double heterozygote (Irf6(+/-);Grhl3(+/-)) murine embryos failed to detect epistasis between the two genes, suggesting that they function in separate but convergent pathways during palatogenesis. Taken together, our data demonstrated that mutations in two genes, IRF6 and GRHL3, can lead to nearly identical phenotypes of orofacial cleft. They supported the hypotheses that both genes are essential for the presence of a functional oral periderm and that failure of this process contributes to VWS.

This retrospective study was carried out on consecutively collected dental casts, x-rays, and photos of 129 Swedish children who had been born with non-syndromic unilateral (U) cleft lip (CL), cleft lip and alveolus (CLA), or cleft lip and palate (CLP). The following dental characteristics were investigated in the primary and permanent dentitions: 1. the presence, eruption, position, and shape of the lateral incisor; 2. the prevalence of rotation and enamel hypoplasia of the permanent central incisor; 3. the occurrence of hypodontia outside the cleft region; and 4. the transition from the primary to the succeeding permanent lateral incisor in the cleft region. Patients with clefts involving the palate (UCLP) exhibited a high frequency of hypodontia. In patients with clefts, which did not include the palate, malformed lateral incisors were a common finding. In the primary and permanent dentition, the lateral incisor had erupted distal to the cleft in 31.8 and 24.8 per cent of the UCLA and UCLP patients, respectively. No significant pattern was seen regarding the transition from the primary to the succeeding permanent lateral incisor (P = 0.15). The central incisor was rotated in 55 per cent of the patients and this positional deviation was particularly frequent in cases where the lateral incisor was missing in the premaxilla (P < 0.05). Hypodontia outside the cleft region was recorded in 15.5 per cent of the patients. Patients with UCLP had more often crossbite than patients with a UCL or a UCLA phenotype (P < 0.001).

The aim of this study was to characterize Swedish families with non-syndromic cleft lip and/or palate (NSCL/P) for mutations or other sequence variants in the interferon regulatory factor 6 (IRF6) gene, as well as to describe their cleft phenotypes and hypodontia. Seventeen Swedish families with at least two family members with NSCL/P were identified and clinically evaluated. Extracted DNA from blood samples was used for IRF6 mutation screening. Exonic fragments of the IRF6 gene were sequenced and chromatograms were inspected. Statistical analysis was undertaken with marker- and haplotype association tests. No disease-associated IRF6 mutation could be determined in the families analyzed. One new and seven known single nucleotide polymorphisms (SNPs) were detected. The A allele of SNP rs861019 in exon 2 and the G allele of SNP rs7552506 in intron 3 showed association with cleft lip and palate (CLP; odds ratios of 3.1 and 5.45, respectively). Hypodontia was observed more commonly in individuals affected with CL/P as compared with family members without a cleft (P < 0.01). The hypodontia most often affected the cleft area, possibly representing a secondary effect. The distribution of cleft phenotypes in 15 of the 17 families with NSCL/P differed from the mixed cleft types seen in Van der Woude syndrome (VWS), in that CLP did not occur together with an isolated cleft palate within the same family. It was concluded that mutations of the IRF6 gene are not a common cause for cleft predisposition in Swedish NSCL/P families.