Youn Jung Kim

Background/Objectives: The ectodysplasin A (EDA) gene, a member of the tumor necrosis factor ligand superfamily, is involved in the early epithelial–mesenchymal interaction that regulates ectoderm-derived appendage formation. Numerous studies have shown that mutations in the EDA gene can cause X-linked ectodermal dysplasia (ED) and non-syndromic oligodontia (NSO). Accordingly, this study aimed to identify the causative genetic mutations of the EDA gene. Methods: We investigated EDA gene mutations in two X-linked oligodontia families using candidate gene sequencing and whole-exome sequencing, with a single proband identified and studied for each family. The first family included a patient with NSO, while the second family had a patient exhibiting variable expression of ED. Results: Mutational analysis identified two missense mutations in the EDA gene (NM_001399.5): one novel mutation, c.787A>C p.(Lys263Gln), in family 2; and one previously reported mutation, c.457C>T p.(Arg153Cys), in family 1. All mutated residues are evolutionarily highly conserved amino acids. The p.(Arg153Cys) mutation would destroy the furin recognition site and affect the cleavage of EDA. The p.(Lys263Gln) mutation in a TNF homology domain would interfere with the binding of the EDA receptor. The p.(Lys263Gln) mutation was associated with NSO, while the other mutation demonstrated ED. Conclusions: This study helps to better understand the nature of EDA-related ED and NSO and their pathogenesis, and it expands the mutational spectrum of EDA mutations.

Background/Objectives: Oligodontia, the absence of six or more teeth excluding third molars, is a rare genetic condition, unlike hypodontia (missing one or more teeth), which is a relatively common human disease. Methods: To identify the genetic etiology of nonsyndromic oligodontia (NSO) families, we performed mutational analysis and investigated the functional effects of identified EDAR mutations. Whole-exome sequencing was conducted on recruited families with NSO. Bioinformatic analysis identified mutations in oligodontia-causing candidate genes, which were confirmed by Sanger sequencing and segregation within families. The impact of EDAR mutations on the EDA signaling pathway was assessed using luciferase activity analysis. Results: EDAR mutations were identified in three NSO families. A homozygous missense EDAR mutation (NM_022336.4: c.319A>G p.(Met107Val)) was found in the singleton proband of family 1. The proband of family 2 carried compound heterozygous EDAR mutations: a maternal missense mutation (c.319A>G p.(Met107Val)) and a paternal missense variant (c.1270G>A p.(Val424Met)). The proband of family 3 had heterozygous EDAR mutations: a maternal missense mutation (c.389T>A p.(Ile130Asn)) and paternal intronic variants in cis (c.[357-4G>A;440+50C>T]). Luciferase assays confirmed reduced transcriptional activity for all identified missense mutations, while splicing assays revealed altered splicing patterns. Conclusions: In conclusion, recessive EDAR mutations, including novel ones, were identified in NSO families, and their pathological mechanism was explored through transcriptional activity measurements.

Hypodontia, i.e., missing one or more teeth, is a relatively common human disease; however, oligodontia, i.e., missing six or more teeth, excluding the third molars, is a rare congenital disorder. Many genes have been shown to cause oligodontia in non-syndromic or syndromic conditions. In this study, we identified two novel PAX9 mutations in two non-syndromic oligodontia families. A mutational analysis identified a silent mutation (NM_006194.4: c.771G>A, p.(Gln257=)) in family 1 and a frameshift mutation caused by a single nucleotide duplication (c.637dup, p.(Asp213Glyfs*104)) in family 2. A minigene splicing assay revealed that the silent mutation resulted in aberrant pre-mRNA splicing instead of normal splicing. The altered splicing products are ones with an exon 4 deletion or using a cryptic 5’ splicing site in exon 4. Mutational effects were further investigated using protein expression, luciferase activity assay and immunolocalization. We believe this study will not only expand the mutational spectrum of PAX9 mutations in oligodontia but also strengthen the diagnostic power related to the identified silent mutation.

Hereditary conditions that affect tooth enamel in quantity and/or quality are called amelogenesis imperfecta (AI). AI can occur as an isolated condition or as a symptom of a syndrome. An OMIM search with the term “AI” yielded 79 result entries. Mutations in the same gene cause syndromic or non-syndromic AI, depending on the nature of the mutations. In this study, we recruited two AI families and performed mutational analysis using whole-exome sequencing. The proband of family 1, with hypoplastic pitted AI and mild localized atopic dermatitis, had compound heterozygous COL17A1 mutations (paternal NM_000494.4: c.3598G>T, p.Asp1200Tyr and maternal c.1700G>A, p.Gly567Glu). The proband of family 2, with hypoplastic pitted AI and Jervell and Lange-Nielsen syndrome, had a recurrent LAMB3 mutation (NM_000228.3: c.3463_3475del, p.(Glu1155Thrfs*51)) in addition to compound heterozygous mutations in the KCNQ1 gene.

Amelogenesis imperfecta (AI) is a heterogeneous collection of hereditary enamel defects. The affected enamel can be classified as hypoplastic, hypomaturation, or hypocalcified in form. A better understanding of normal amelogenesis and improvements in our ability to diagnose AI through genetic testing can be realized through more complete knowledge of the genes and disease-causing variants that cause AI. In this study, mutational analysis was performed with whole exome sequencing (WES) to identify genetic etiology underlying the hypomaturation AI condition in affected families. Mutational analyses identified biallelic WDR72 mutations in four hypomaturation AI families. Novel mutations include a homozygous deletion and insertion mutation (NM_182758.4: c.2680_2699delinsACTATAGTT, p.(Ser894Thrfs*15)), compound heterozygous mutations (paternal c.2332dupA, p.(Met778Asnfs*4)) and (maternal c.1287_1289del, p.(Ile430del)) and a homozygous 3694 bp deletion that includes exon 14 (NG_017034.2:g.96472_100165del). A homozygous recurrent mutation variant (c.1467_1468delAT, p.(Val491Aspfs*8)) was also identified. Current ideas on WDR72 structure and function are discussed. These cases expand the mutational spectrum of WDR72 mutations causing hypomaturation AI and improve the possibility of genetic testing to accurately diagnose AI caused by WDR72 defects.

The process of tooth formation is a series of reciprocal interactions between the ectoderm and mesoderm, and it is believed that many genetic factors are involved in this complex process. More than a dozen genes have been identified in non-syndromic tooth agenesis; however, the genetic etiology underlying tooth agenesis is not fully understood yet. In this study, we identified two novel LRP6 mutations in two non-syndromic oligodontia families. Both probands had 16 and 17 missing teeth in their permanent dentition. Mutational analysis identified a de novo frameshift mutation by a 1-bp insertion in exon 9 (NM_002336.2: c.1870dupA, p.(Met624Asnfs*29)) and a splicing donor site mutation in intron 8 (c.1762+2T>C). An in vitro splicing assay confirmed the deletion of exon 8, and the deletion would result in a frameshift. Due to the premature termination codons introduced by the frameshift, both mutant transcripts would be degraded by nonsense-mediated mRNA decay, resulting in haploinsufficiency.

Hereditary dentin defects are conventionally classified into three types of dentinogenesis imperfecta (DGI) and two types of dentin dysplasia (DD). Mutations in the dentin sialophosphoprotein (DSPP) gene have been identified to cause DGI type II and III and DD type II; therefore, these are not three different conditions, but rather allelic disorders. In this study, we recruited three families with varying clinical phenotypes from DGI-III to DD-II and performed mutational analysis by candidate gene analysis or whole-exome sequencing. Three novel mutations including a silent mutation (NM_014208.3: c.52-2del, c.135+1G>C, and c.135G>A; p.(Gln45=)) were identified, all of which affected pre-mRNA splicing. Comparison of the splicing assay results revealed that the expression level of the DSPP exon 3 deletion transcript correlated with the severity of the dentin defects. This study did not only expand the mutational spectrum of DSPP gene, but also advanced our understanding of the molecular pathogenesis impacting the severity of hereditary dentin defects.

Amelogenesis imperfecta (AI) is a group of rare genetic diseases affecting the tooth enamel. AI is characterized by an inadequate quantity and/or quality of tooth enamel and can be divided into three major categories: hypoplastic, hypocalcified and hypomaturation types. Even though there are some overlapping phenotypes, hypomaturation AI enamel typically has a yellow to brown discoloration with a dull appearance but a normal thickness indicating a less mineralized enamel matrix. In this study, we recruited four Turkish families with hypomaturation AI and performed mutational analysis using whole exome sequencing. These analyses revealed two novel homozygous mutations in the KLK4 gene: a nonsense mutation in exon 3 (NM_004917.4:c.170C>A, p.(Ser57*)) was found in families 1, 2 and 3 and a missense mutation in exon 6 (c.637T>C, p.(Cys213Arg)) in family 4. Functional analysis showed that the missense mutation transcript could not translate the mutant protein efficiently or generated an unstable protein that lacked functional activity. The two novel inactivating KLK4 mutations we identified caused a hypomaturation AI phenotype similar to those caused by the four previously described KLK4 nonsense and frameshift mutations. This study improves our understanding of the normal and pathologic mechanisms of enamel formation.

Hereditary dentin defects can be categorized as a syndromic form predominantly related to osteogenesis imperfecta (OI) or isolated forms without other non-oral phenotypes. Mutations in the gene encoding dentin sialophosphoprotein (DSPP) have been identified to cause dentinogenesis imperfecta (DGI) Types II and III and dentin dysplasia (DD) Type II. While DGI Type I is an OI-related syndromic phenotype caused mostly by monoallelic mutations in the genes encoding collagen type I alpha 1 chain (COL1A1) and collagen type I alpha 2 chain (COL1A2). In this study, we recruited families with non-syndromic dentin defects and performed candidate gene sequencing for DSPP exons and exon/intron boundaries. Three unrelated Korean families were further analyzed by whole-exome sequencing due to the lack of the DSPP mutation, and heterozygous COL1A2 mutations were identified: c.3233G>A, p.(Gly1078Asp) in Family 1 and c.1171G>A, p.(Gly391Ser) in Family 2 and 3. Haplotype analysis revealed different disease alleles in Families 2 and 3, suggesting a mutational hotspot. We suggest expanding the molecular genetic etiology to include COL1A2 for isolated dentin defects in addition to DSPP.

This study aimed to identify the molecular genetic etiology of an 8-year-old boy with amelogenesis imperfecta in permanent dentition. Bilateral cochlear implants were placed due to sensorineural hearing loss, and there was no other family member with a similar phenotype. Peripheral blood samples were collected with the understanding and written consent of the participating family members. A constitutional chromosome study was performed for the proband. Genomic DNA was isolated, and whole exome sequencing was performed. A series of bioinformatic analyses were performed with the obtained paired-end sequencing reads, and the variants were filtered and annotated with dbSNP147. There was no abnormality in the constitutional chromosome study. Whole exome sequencing analysis with trio samples identified a homozygous mutation (c.506T>C, p. (Leu169Pro)) in the PEX26 gene. We verified “temperature sensitivity (ts)” of patient-derived Pex26-L169P by expression in pex26 CHO mutant ZP167 cells to determine the effect of the L169P mutation on Pex26 function. The L169P mutation causes a mild ts-cellular phenotype representing the decreased peroxisomal import of catalase. This study supports the finding that the recessive mutations in PEX26 are associated with Heimler syndrome and demonstrates the importance of an early and correct diagnosis.