In our study we used the syndromic cleft palate susceptibility gene as a candidate gene for more common non-syndromic cleft palate. Efforts to identify the genetic factors have been most successful for monogenic, syndromic clefts, such as in Van der Woude syndrome (involving IRF6); Margarita Island syndrome (PVRL1); Kallmann syndrome (FGFR1); ectrodactyly, ectodermal dysplasia, and cleft lip and/or palate syndrome (p63); cleft lip and/or palate with hypodontia (MSX1); and X-linked cleft palate (TBX22) [3
]. The value of translating a mutation causing a rarely observed syndromic condition onto more common non-syndromic forms of the disorder has been elegantly demonstrated in the study by Sözen et al
] for the role of PVRL1 in cleft lip with or without cleft palate. Phenotypic penetrance within families is often variable, and has not been explained by the type or position of mutations within the genes [11
Considerable success has been achieved in identifying aetiological genes for syndromic forms of orofacial clefting with approximately 15-20% cloned to date. TBX22 is the important susceptibility gene currently found in X-linked cleft palate – ankyloglossia syndrome (CPX) and it is a major genetic determinant for familial cleft palate, particularly when combined with ankyloglossia. Linkage and/or haplotype analysis has been used to demonstrate that the cleft palate and ankyloglossia phenotype was X-linked and mapped to the approximate interval of CPX in Xq21 [4
]. As a transcription factor, TBX22 is expressed specifically in tissues related to palatogenesis and it is confirmed as a major genetic influence in normal palate development. TBX22 expression was reported to disappear just prior to palatal shelf fusion and was thought instead to be required for mesenchymal proliferation and shelf elevation. Identification of the downstream target genes remains a priority [3
TBX22 clearly makes a significant contribution to the prevalence of cleft palate and it will be important to study its biological role in the process of craniofacial development. To date, 13 different TBX22 mutations have been reported in CPX patients (http://www.hgmd.cf.ac.uk
), including missense, nonsense, splicing, regulatory, small deletions and small insertions. There were four mutations located in TBX22 exon 5: 641T→C(L214P) [4
], 671delC(221) [5
], 785C→T(T260M) [5
], 790A→T(A264T) [6
]. In our research the sequence alignment did not reveal any credible mutation. We have not examined all the exons of TBX22, so mutations may also be harboured in the other exons. Suphapeetiporn et al
] in their study performed mutation analysis that covered all the coding regions of the TBX22 gene in 53 unrelated Thai patients with non-syndromic cleft palate, and identified four potentially pathogenic mutations: 359G→A (R120Q), 452G→T (R151L), 1166C→A (P389Q) and 1252delG in four different patients. There were also no mutations in exon 5 in the Thai population with non-syndromic cleft palate.
A better understanding of the cellular biology will help us to afford appropriate advice, therapy and ultimately prevention. A missense variant (359G→A; R120Q) was detected in the Thai population with ankyloglossia alone, the same mutation being previously reported in Thai patients with non-syndromic cleft palate. This nucleotide substitution was next to the one that has been described in a Tunisian CPX family (358C→T; R120W). From the crystal structure of the Brachyury T domain, this position lies within the αβ barrel and is predicted to be involved in DNA binding. The amino acid of another missense variant in a CPX family (790A→T; A264T) is located in the T box domain at the start of the α helix 3, which is the position to recognise the target DNA in the minor groove and bridges the DNA backbone. Interfering with DNA binding is likely to be one of the major mechanisms underlying cleft palate with or without ankyloglossia as evidenced by a number of missense mutations found in the DNA-binding domain of TBX22 [6
Based on the family-based study which refers to the patients and their healthy parents, the parent-child trio becomes the research object. Transmission of a particular allele from heterozygous parents to an offspring is expected to be approximately 50%, when there is no association of the allele and the disease. If the allele is associated with an increased risk of the disease, excess transmission to the offspring is expected. This approach accounts for the non-independence of parental transmissions to affected offspring [17
]. The suspected correlation was subjected to validation using the transmission disequilibrium test. The allele is not transmitted to affected children. As a control with a pair of alleles for a 1 : 1 matched case-control study, TDT eliminates the case-control population stratification problem, largely avoiding the phenomenon of the occurrence of false positives and improving the accuracy of gene targeting [19
In conclusion, we found no mutations in TBX22 exon 5 in our patients with non-syndromic cleft palate, but the sample size is limited, and mainly concentrated in the selection of patients within Jiangzhe areas in China. As coverage is not wide enough, these restrictions may have some influences and induce negative results.