Candidate gene identification using whole exome sequencing method in familial dental transposition cases

Aim: Dental transposition is defined as the displacement of two adjacent teeth due to the disruption in the tooth eruption mechanism. Although different classifications have been made by different authors about the classification of dental transposition, it is generally accepted that dental transposition is divided into two sub-classes as completed and incomplete. While its prevalence in the world population is between 0.1-2.4%, in the Turkish population is between 0.1-0.6%. It is seen unilaterally in the permanent dentition and it is more common in women, differing between genders. There are several theories such as heredity, trauma, migration, etc. in etiology. Because of dental transposition is closely related to some dental anomalies such as congenital agenesis (hereditary tooth deficiency), shows familial clustering (familial agenesis) and occurs together with syndromes such as cleft lif-palate (CLP) (4-18%)- Down (3-15%) syndrome, the most widely accepted among of these theories has been the heredity. However, which of these theories is correct has not been clarified until today. In addition, no study has been found in the current literature to report genes or variations that may play a role in the pathway of dental transposition. In this thesis, in individuals with non-syndromic familial dental transposition; it was aimed to determine the genes and variations that cause dental transposition, also new variations in genes that cause anomalies and syndromes (agenesis, Down, CLP etc.) associated with dental transposition. Materials and methods: A total of 6 individuals (5F, 1M) from 3 families were included in this study. To obtain DNA from individuals, samples were taken from the buccal epithelium with a swab stick. Genomic DNA was isolated from this sample and the whole exome sequencing (WES) was performed with the NovaSeq Illumina 6000 device in accordance with the protocol of the kit. Candidate genes were determined by analyzing the obtained data with bioinformatics methods. These genes were filtered and interpreted using the following criteria: being from previously known genes, located in a pathway that plays a role in dental development, located in a pathway dental agenesis related to dental transposition, located in a pathway that causes CLP and Down syndrome, the Minor Allele Frequency value is consistent with population studies (MAF <0.6%). Results: Transposition appeared unilaterally and equally on the right and left sides, and it was more common in women, in accordance with the literature,. It was found that dental transposition was seen with impacted tooth, eruption delay, persistent deciduous tooth and congenital 3rd molar agenesis. However, no agenesis other than the 3rd Molar (especially in lateral teeth) or peg-shape lateral teeth was found. Also in all individuals participating in the study, it was determined that one of the affected teeth was maxillary canine. Candidate variants for dental transposition pathway were detected in DSPP, DSP, GLI3 and PRICKLE2 genes, which were previously reported to play a role in dental development. Variants in the DSPP gene cause structural abnormalities such as autosomal dominant (AD) dentinogenesis imperfecta and dentin dysplasia. Although DSPP variants of homozygous character were found in our individuals, such clinical diseases were not observed. For this reason, it seems unlikely that this gene plays a role in dental transposition. However, considering that there is also a heterozygous variant in the DSPP gene and that DSPP may play a role in the agenesis mechanism due to its relationship with the WNT pathway, it is thought that it may be effective in the dental transposition mechanism, albeit low. Verification studies with Sanger sequence analysis and segregation analysis are required. DSP and GLI3 genes play a role in dental development and are associated with syndromic and non-syndromic agenesis-oligodontics. Although variants in these genes were seen in two of the individuals in our study in homozygous character, hypodontics-agenesis was not observed in these individuals. For this reason, the probability of these two genes to play a role in the dental transposition pathway is unlikely. But it is known that DSP and GLI3 play a role in dental development, the GLI3 gene is associated with the SHH pathway, which plays an important role in dental development, and the GLI3 gene is thought to be effective in determining craniofacial skeletal morphology. So this situation should be verified by Sanger sequence analysis and segregation analysis for a group including unaffected family members. PRICKLE2 gene plays a role in the planar cell polarity (PCP) system, participates in cellular behavior, signaling and biological processes, and takes part in amelogenesis For this reason, it is thought that it can take part in the dental transposition mechanism. The fact that this variant is seen in heterozygous character in individuals suggests that it may be an AD inherited anomaly and dental transposition appears as a mild phenotype. In addition, dental impaction, eruption delay, 3rd molar agenesis, mandibular retrognathy, upper incisor retroclination in the index individual and 3rd molar agenesis in the parent together with dental transposition; it suggests that the PRICKLE2 gene may play a role in the cause of other dental anomalies. Also PRICKLE2 gene is located in the WNT pathway that causes agenesis, malformed teeth and root agenesis. In the light of these findings, PRICKLE2 gene seems to be our most likely candidate. However, in order to reach definitive results, it is planned to verify this gene with Sanger sequence analysis, segregation analysis to be carried out on the whole family, including family members who do not show dental transposition, and to be confirmed by further studies on healthy populations and to perform functional tests in animal model. Gli3 deficient mice are seen CLP, and the expression of GLI3 gene in the SHH pathway and PRICKLE2 gene in the WNT pathway changes in Down syndrome. Therefore, familial dental transposition is thought to be a subclinical phenotypic in these diseases. However, especially in CLP in accordance with the literature; it has been predicted that dental anomalies are more likely to occur due to environmental factors such as the displacement of alveolar crests and tooth germs in the cleft region, and maxillary growth failure due to the effect of scar tissue caused by surgical procedures. No significant variants were found in MSX1 and PAX9 genes, which are known to play a very important role in dental development and were previously thought to be an etiological factor in mandibular canine transposition. Conclusions: In this thesis, results were obtained in accordance with the general literature information such that dental transposition is mostly seen in maxillary canines, unilateral, equally on the right and left sides, mostly in women. It has been determined that dental transposition may also show AD inheritance due to variants in some genes involved in tooth development and dental agenesis mechanism, which has a hereditary character, and due to familial clustering. Because of the complexity of tooth development, it was thought that the occurrence of dental transposition was caused by multifactorial interactions rather than a single gene disorders. But it was determined that further family studies were needed. Surprisingly, since all the affected teeth are maxillary canines, it is thought that the migration theory should not be ignored. The WES method used this study was thought to be effective in determining candidate gene. The findings of this thesis obtained will lead to a better understanding of dental development. Thus, by bringing a different perspective to these types of anomalies and malocclusions, the most appropriate diagnosis and treatment opportunity will be provided in the early period.