Paired Box 9 Research Articles

Measurement and analysis of the crown conical degree of maxillary incisors in patients with congenital tooth agenesis caused by different gene mutations

Objective: To measure the crown conical degree of the remaining maxillary incisors in patients with congenital tooth agenesis, and to analyze the influence of different gene mutations on the crown conical degree of patients. Methods: Whole exome sequencing was performed on 85 patients with congenital tooth agenesis (50 males, 35 females, median age 19 years old) who visited the Department of Prosthodontics, Peking University School and Hospital of Stomatology from January 2019 to January 2023. The pathogenic gene was identified. The width of the crowns of the maxillary central and lateral incisors at the incisal 1/3 and gingival 1/3 were measured on the panoramic radiograph, and the ratio was defined as the crown conical degree. The smaller the ratio is, the more likely is the crown to be peg shaped teeth. The control group was matched by age and gender with 85 other patients with intact maxillary permanent incisors who were treated in the Department of Prosthodontics, Peking University School and Hospital of Stomatology from January 2019 to January 2023. The conical degree of the tooth agenesis group was compared with the control group by t-test, and the differences in the crown conical degree in different gene mutation groups were compared using one-way analysis of variance, and the multiple comparisons among gene groups were carried out using the LSD method. Results: Among the 85 tooth agenesis patients, the numbers of patients in each gene mutation group were 20 in ectodysplasin A (EDA) group, 8 in ectodysplasin A receptor (EDAR) group, 15 in wingless-type MMTV integration site family, member 10A (WNT10A) group, 16 in paired box 9 (PAX9) group, 10 in Msh homeobox 1 (MSX1) group, 10 in low-density Lipoprotein receptor related protein 6 (LRP6) group, and 6 in bone morphogenetic protein4 (BMP4) group. The number of missing teeth were 1-27, median number 15 among the tooth agenesis patients. There was no significant difference in the conical degree between the left and right homonymous teeth in the congenital tooth agenesis group and the control group (P>0.05). The crown conical degree of maxillary central incisor and lateral incisor in the congenital missing teeth group (0.95±0.24, 0.90±0.22) was significantly smaller than that in the control group (1.12±0.09, 1.13±0.09) (t=-8.50, P<0.001; t=-11.47, P<0.001). In WNT10A mutants, the conical degree of lateral incisors (0.89±0.18) was less than that of central incisors (1.07±0.15)(t=3.68, P<0.001). The conical degree of central incisors and lateral incisors (0.70±0.23, 0.57±0.15) of EDA mutants was significantly lower than that in patients with other gene mutations (P>0.05). Conclusions: Compared with the normal control group, the remaining maxillary central and lateral incisors of the seven gene mutation groups of patients with congenital tooth agenesis all had different degrees of conical crown. Among them, the crown conical degree of maxillary central and lateral incisors of the EDA mutation was the most severe, and the WNT10A mutation affected the maxillary lateral incisors more specifically.

Expression of Paired box 9 defines an aggressive subset of lung adenocarcinoma preferentially occurring in smokers.

A distinct subset of lung adenocarcinomas (LADs), arising from a series of peripheral lung cells defined as the terminal respiratory unit (TRU), is characterized by thyroid transcription factor 1 (TTF-1) expression. The clinical relevance of transcription factors (TFs) other than TTF-1 remains unknown in LAD and was explored in the present study. Seventy-one LAD samples were subjected to high-throughput transcriptome screening of LAD using cap analysis gene expression (CAGE) sequencing data; CAGE provides genome-wide expression levels of the transcription start sites (TSSs). In total, 1083 invasive LAD samples were subjected to immunohistochemical examination for paired box 9 (PAX9) and TTF-1 expression levels. PAX9 is an endoderm development-associated TF that most strongly and inversely correlates with the expression of TTF-1 TSS subsets. Immunohistochemically, PAX9 expression was restricted to the nuclei of ciliated epithelial and basal cells in the bronchi and bronchioles and the nuclei of epithelial cells of the bronchial glands; moreover, PAX9 expression was observed in 304 LADs (28%). PAX9-positive LADs were significantly associated with heavy smoking, non-lepidic subtype, EGFR wild-type tumors, and PD-L1 expression (all p < 0.0001). All these characteristics were opposite to those of TRU-type LADs with TTF-1 expression. PAX9 expression was an independent prognostic factor for decreased overall survival (p = 0.022). Our results revealed that PAX9 expression defines an aggressive subset of LADs preferentially occurring in smokers that may arise from bronchial or bronchiolar cells.

Genes Linked with Orthodontic Issues and the Knowledge Gaps in This Association

Orthodontics is the study of malpositioned teeth and jaws. Some commonly found orthodontic issues are malocclusions, mandibular prognathism, retrognathism, and facial asymmetry. These issues are associated with several genetic factors. For instance, mandibular prognathism is related to gene variations, such as insulin-like growth factor 1 and the HoxC gene cluster. Facial asymmetry is related to NFATC1 and NBAS genes. Dental agenesis is related to several genes, including paired box 9 (PAX9), homeobox 1 (MSX1), and ectodysplasin A. Apart from these genes, several other genes, such as MYO1H gene, ACTN3 gene, ENPP1, ESR1 genes, PITX1 and PITX2 genes, IL1 gene, and PAX9 gene, have been studied in the orthodontic issues faced by people. Some of these genes, such as ENPP1 and ESR1 genes, are directly related to orthodontic issues, whereas some of the genes, such as ACTN3 genes, are associated with issues in the muscle tone that can eventually lead to orthodontic issues. This review deals with genes linked with orthodontics and the knowledge gaps in its association.

PAX9 in Cancer Development.

Paired box 9 (PAX9) is a transcription factor of the PAX family functioning as both a transcriptional activator and repressor. Its functional roles in the embryonic development of various tissues and organs have been well studied. However, its roles and molecular mechanisms in cancer development are largely unknown. Here, we review the current understanding of PAX9 expression, upstream regulation of PAX9, and PAX9 downstream events in cancer development. Promoter hypermethylation, promoter SNP, microRNA, and inhibition of upstream pathways (e.g., NOTCH) result in PAX9 silencing or downregulation, whereas gene amplification and an epigenetic axis upregulate PAX9 expression. PAX9 may contribute to carcinogenesis through dysregulation of its transcriptional targets and related molecular pathways. In summary, extensive studies on PAX9 in its cellular and tissue contexts are warranted in various cancers, in particular, HNSCC, ESCC, lung cancer, and cervical SCC.

Functional study of novel PAX9 variants: The paired domain and non-syndromic oligodontia.

To investigate pathogenic variants of the paired box 9 (PAX9) gene in patients with non-syndromic oligodontia, and the functional impact of these variants. Whole exome sequencing and Sanger sequencing were utilized to detect gene variants in a cohort of 80 patients diagnosed with non-syndromic oligodontia. Bioinformatic and conformational analyses, fluorescence microscopy and luciferase reporter assay were employed to explore the functional impact. We identified three novel variants in the PAX9, including two frameshift variants (c.211_212insA; p.I71Nfs*246 and c.236_237insAC; p.T80Lfs*6), and one missense variant (c.229C>G; p.R77G). Familial co-segregation verified an autosomal-dominant inheritance pattern. Conformational analyses revealed that the variants resided in the paired domain, and could cause corresponding structural impairment of the PAX9 protein. Fluorescence microscopy showed abnormal subcellular localizations of frameshift variants, and luciferase assay showed impaired downstream transactivation activities of the bone morphogenetic protein 4 (BMP4) gene in all variants. Our findings broaden the spectrum of PAX9 variants in patients with non-syndromic oligodontia and support that paired domain structural impairment and the dominant-negative effect are likely the underlying mechanisms of PAX9-related non-syndromic oligodontia. Our findings will facilitate genetic diagnosis and counselling, and help lay the foundation for precise oral health therapies.

Common variants of EDA are associated with non-syndromic hypodontia.

The aim of this case-control study was to investigate the association between non-syndromic hypodontia and nineteen common variants of candidate genes ectodysplasin A (EDA), paired box 9 (PAX9), msh homeobox 1 (MSX1) and axis inhibition protein 2 (AXIN2). Sixty-one hypodontia cases were frequency-matched to 253 controls with no missing teeth (excluding the third molars). Self-report data and DNA samples were collected from each participant. The sample had a mean age of 16.6years (SD=7.3), with most participants being female (59.6%), and of New Zealand European origin (75.4%). Using multiple logistic regression analysis, it was found that the T-allele of rs12853659 (EDA) and the G-allele of rs2428151 (EDA) were both associated with a higher risk of hypodontia (odds ratio, OR=2.79, 95% CI=1.11-7.01; and OR=2.87, 95% CI=1.04-7.94, respectively). The G-allele of rs2520378 (EDA) showed a protective effect with an OR of 0.61 (95% CI=0.38-0.99). The EDA SNP findings were consistent with previous reports included in a meta-analysis. No associations were found with the PAX9, AXIN2 and MSX1 genes, after adjusting for sex and ethnicity. Common variants of the EDA genes are associated with specific phenotypes of non-syndromic hypodontia, thus confirming their role in the regulatory pathways of normal tooth development. However, larger samples are needed to investigate the association further.

Paired Box 9 (PAX9), the RNA polymerase II transcription factor, regulates human ribosome biogenesis and craniofacial development.

Dysregulation of ribosome production can lead to a number of developmental disorders called ribosomopathies. Despite the ubiquitous requirement for these cellular machines used in protein synthesis, ribosomopathies manifest in a tissue-specific manner, with many affecting the development of the face. Here we reveal yet another connection between craniofacial development and making ribosomes through the protein Paired Box 9 (PAX9). PAX9 functions as an RNA Polymerase II transcription factor to regulate the expression of proteins required for craniofacial and tooth development in humans. We now expand this function of PAX9 by demonstrating that PAX9 acts outside of the cell nucleolus to regulate the levels of proteins critical for building the small subunit of the ribosome. This function of PAX9 is conserved to the organism Xenopus tropicalis, an established model for human ribosomopathies. Depletion of pax9 leads to craniofacial defects due to abnormalities in neural crest development, a result consistent with that found for depletion of other ribosome biogenesis factors. This work highlights an unexpected layer of how the making of ribosomes is regulated in human cells and during embryonic development.

WNT10A variants in relation to nonsyndromic hypodontia in eastern Slovak population

Nonsyndromic hypodontia is a congenital absence of less than six permanent teeth, with a most common subtype maxillary lateral incisor agenesis (MLIA). Mutations in several genes have been described in severe tooth agenesis. The aim of this study was to search for the variants in wingless-type MMTV-integration site family member (WNT10A), paired box 9 (PAX9) and axis inhibitor 2 (AXIN2) genes, and investigate their potential role in the pathogenesis of non-syndromic hypodontia. Clinical examination and panoramic radiograph were performed in the cohort of 60 unrelated Slovak patients of Caucasian origin with nonsyndromic hypodontia including 37 MLIA cases and 48 healthy controls. Genomic DNA was isolated from buccal swabs and Sanger sequencing of WNT10A, PAX9 and AXIN2 was performed. Altogether, we identified 23 single-nucleotide variants, of which five were novel. We have found three rare nonsynonymous variants in WNT10A (p.Gly165Arg; p.Gly213Ser and p.Phe228Ile) in eight (13.33%) of 60 patients. Analysis showed potentially damaged WNT10A variant p.Phe228Ile predominantly occurred only in MLIA patients, and with a dominant form of tooth agenesis (odds ratio (ORdom) = 9.841; P = 0.045; 95% confidence interval (CI) 0.492-196.701;ORrec = 0.773; P = 1.000; 95% CI 0.015-39.877). In addition, the WNT10A variant p.Phe228Ile showed a trend associated with familial nonsyndromic hypodontia (P = 0.024; OR= 1.20; 95% CI 0.97-1.48). After Bonferroni correction, these effects remained with borderline tendencies. Using a 3D WNT10A protein model, we demonstrated that the variant Phe228Ile changes the proteinsecondary structure. In PAX9 and AXIN2, common variants were detected. Our findings suggest that the identified WNT10A variant p.Phe228Ile could represent risk for the inherited nonsyndromic hypodontia underlying MLIA. However, further study in different populations is required.

Identification Of PAX9 Single Nucleotide Polymorphism In Class III Malocclusion Patients With Mandibular Prognatism

Introduction: PAX9 (Paired box 9) gene is one of the genes which play significant role during craniofacial development. Single nucleotide polymorphism (SNP) in PAX9 has been associated with Class II/Division 2 malocclusion (with or without hypodontia). However, the relationship between PAX9 SNP marker (rs8004560) with mandibular prognathism (MP) has not been analysed, at least in our local population. This study aimed to detect the presence of PAX9 (rs8004560) SNP in Class III malocclusion patients (with MP) in the local population. Materials and Methods: Genomic DNA were extracted from unstimulated saliva of 31 class I malocclusion (control samples) and 30 patients from Class III malocclusion (MP). Cephalometric measurements were performed prior to saliva samples collection. The DNA was amplified using the specific primers for the marker rs8004560 and the genotyping was done by sequencing. Chi-square test was used to determine the overrepresentation of marker allele (p&lt;0.05). Results: Presence of PAX9 SNP (rs8004560) was detected in local population analysed and the distribution of its genotype and allele could be observed. There were significant differences between allele (p=0.000) and genotype (p=0.000) frequency within control (Class I) and Class III malocclusion. Conclusion(s): The most common allele of a marker flanking PAX9 (rs8004560) was over-represented in the mandibular prognathism (MP) subjects indicating the genetic association of PAX9 (rs8004560) SNP in the incidence of MP. Further studies involving larger number of samples should be developed in order to understand the exact role and mechanism of PAX9 in different classes of malocclusions.

Genetic study of non-syndromic tooth agenesis through the screening of paired box 9, msh homeobox 1, axin 2, and Wnt family member 10A genes: a case-series.

Non-syndromic tooth agenesis (NSTA) is the most common developmental anomaly in humans. Several studies have been conducted on dental agenesis and numerous genes have been identified. However, the pathogenic mechanisms responsible for NSTA are not clearly understood. We studied a group of 28 patients with sporadic NSTA and nine patients with a family history of tooth agenesis. We focused on four genes - paired box 9 (PAX9), Wnt family member 10A (WNT10A), msh homeobox 1 (MSX1), and axin 2 (AXIN2) - using direct Sanger sequencing of the exons and intron-exon boundaries. The most prevalent variants identified in PAX9 and AXIN2 genes were analyzed using the chi-square test. The sequencing results revealed a number of variants in the AXIN2 gene, including one novel missense mutation in one patient with agenesis of a single second premolar. We also identified one variant in the AXIN2 gene as being a putative risk factor for tooth agenesis. Only one missense mutation was identified in the WNT10A gene and this mutation was found in two patients. Interestingly, WNT10A is reported as the most prevalent gene mutated in the European population with NSTA.

PAX9 gene mutations and tooth agenesis: A review.

Paired box 9 (PAX9) is one of the best-known transcription factors involved in the development of human dentition. Mutations in PAX9 gene could, therefore, seriously influence the number, position and morphology of the teeth in an affected individual. To date, over 50 mutations in the gene have been reported as associated with various types of dental agenesis (congenitally missing teeth) and other inherited dental defects or variations. The most common consequence of PAX9 gene mutation is the autosomal-dominant isolated (non-syndromic) oligodontia or hypodontia. In the present review, we are summarizing all known PAX9 mutations as well as their nature and precise loci in the DNA sequence of the gene. Where necessary, we have revised the loci of the mutations in line with the reference sequence of the PAX9 gene as it appears in the current DNA databases.

Prognostic value of PAX9 in patients with esophageal squamous cell carcinoma and its prediction value to radiation sensitivity.

Abnormal paired box 9 (PAX9) expression is associated with tumorigenesis, cancer development, invasion and metastasis. The present study investigated the prognostic significance of PAX9 in esophageal squamous cell carcinoma (ESCC) and its role in predicting radiation sensitivity. A total of 52.8% (121/229) ESCC tissues were positive for PAX9. The 1-, 3- and 5-year disease-free survival (DFS) rates were 72.2, 35.2 and 5.6%, respectively, and the overall survival (OS) rates were and 86.1, 44.4, and 23.1%, respectively, in PAX9-positive tumors. In PAX9-negative tumors, the one-, three- and five-year DFS rates were 76.9, 47.9 and 24.0%, and the OS rates were 90.9, 57.9 and 38.8%, respectively. Univariate analysis revealed that PAX9, differentiation, T stage, lymph node metastasis, and tumor-node-metastasis stage were associated with OS. Multivariate analysis of DFS and OS revealed that the hazard ratios for PAX9 were 0.624 (95% CI: 0.472–0.869, P=0.004) and 0.673 (95% CI: 0.491–0.922, P=0.014), respectively. Patients that received adjuvant therapy exhibited significant differences in the 5-year DFS (P<0.001) and OS (P<0.001). PAX9-positive ESCC patients who received post-surgery radiotherapy had a significantly greater 5-year DFS (P=0.011) and OS (P=0.009) than patients who received surgery only. Thus, PAX9 may be an independent prognostic factor for the surgical treatment of ESCC and a possible predictor of radiation sensitivity.

Mutation analysis by direct and whole exome sequencing in familial and sporadic tooth agenesis.

Dental agenesis is one of the most common congenital craniofacial abnormalities. Dental agenesis can be classified, relative to the number of missing teeth (excluding third molars), as hypodontia (1 to 5 missing teeth), oligodontia (6 or more missing teeth), or anodontia (lack of all teeth). Tooth agenesis may occur either in association with genetic syndromes, based on the presence of other inherited abnormalities, or as a non-syndromic trait, with both familiar and sporadic cases reported. In this study, we enrolled 16 individuals affected by tooth agenesis, prevalently hypodontia, and we carried out direct Sanger sequencing of paired box 9 (PAX9) and Msh homeobox 1 (MSX1) genes in 9 subjects. Since no mutations were identified, we performed whole exome sequencing (WES) in the members of 5 families to identify causative gene mutations either novel or previously described. Three individuals carried a known homozygous disease mutation in the Wnt family member 10A (WNT10A) gene (rs121908120). Interestingly, two of these individuals were siblings and also carried a heterozygous functional variant in EDAR-associated death domain (EDARADD) (rs114632254), another disease causing gene, generating a combination of genetic variants never described until now. The analysis of exome sequencing data in the members of other 3 families highlighted new candidate genes potentially involved in tooth agenesis and considered suitable for future studies. Overall, our study confirmed the major role played by WNT10A in tooth agenesis and the genetic heterogeneity of this disease. Moreover, as more genes are shown to be involved in tooth agenesis, WES analysis may be an effective approach to search for genetic variants in familiar or sporadic tooth agenesis, at least in more severe clinical manifestations.

Functional analysis of a novel missense mutation in AXIN2 associated with non-syndromic tooth agenesis.

Tooth agenesis is a congenital anomaly frequently seen in humans. Several genes have been associated with non-syndromic tooth agenesis, including msh homeobox 1 (MSX1), paired box 9 (PAX9), axis inhibition protein 2 (AXIN2), ectodysplasin A (EDA), and wingless-type MMTV integration site family member 10A (WNT10A). In this study, we investigated a Chinese family with non-syndromic tooth agenesis. A novel missense mutation (c.C1978T) in AXIN2 was identified in affected members. The mutation results in a His660Tyr substitution located between the Axin beta-catenin binding domain and the DIX domain of the axis inhibition protein 2 (AXIN2). We analysed this novel AXIN2 mutant, together with two reported AXIN2 mutants [c.1966C>T (p.Arg656Stop) and c.1994delG (p.Leu688Stop)] that cause colorectal cancer with and without oligodontia, to study the effect of the mutant p.His660Tyr on the Wnt/β-catenin signaling pathway and to compare the molecular pathogenesis of different AXIN2 mutants in tooth agenesis and carcinogenesis. Further in vitro experiments indicated that the mutant p.His660Tyr caused inhibition of the Wnt/β-catenin pathway, and the mutants p.Arg656Stop and p.Leu688Stop resulted in over-activation of the Wnt/β-catenin pathway. In line with previous AXIN2 mutation studies, we suggest that AXIN2 mutations with different levels of severity may have distinct effects on the Wnt pathway and the phenotype of disease. Our study provides functional evidence supporting the notion that both inhibition and over-activation of the Wnt pathway may lead to tooth agenesis.

A screen of a large Czech cohort of oligodontia patients implicates a novel mutation in the PAX9 gene.

Tooth agenesis is one of the most common developmental anomalies in humans. To date, many mutations involving paired box 9 (PAX9), msh homeobox 1 (MSX1), and axin 2 (AXIN2) genes have been identified. The aim of the present study was to perform screening for mutations and/or polymorphisms using the capillary sequencing method in the critical regions of PAX9 and MSX1 genes in a group of 270 individuals with tooth agenesis and in 30 healthy subjects of Czech origin. This screening revealed a previously unknown heterozygous g.9527G>T mutation in the PAX9 gene in monozygotic twins with oligodontia and three additional affected family members. The same variant was not found in healthy relatives. This mutation is located in intron 2, in the region recognized as the splice site between exon 2 and intron 2. We hypothesize that the error in pre-mRNA splicing may lead to lower expression of PAX9 protein and could have contributed to the development of tooth agenesis in the affected subjects.

WNT10A coding variants and maxillary lateral incisor agenesis with associated dental anomalies.

Congenital maxillary lateral incisor agenesis (MLIA) is one of the most common subtypes of dental agenesis. Because little is known with regard to the aetiology of this anomaly, the aim of the study was to determine the contribution of nucleotide variants in wingless-type MMTV integration site family, member 10A (WNT10A), msh homeobox 1 (MSX1), and paired box 9 (PAX9) to the risk of MLIA in a Polish population. Coding regions of the selected genes were analysed by direct sequencing in a group of 20 individuals with unilateral and bilateral MLIA, associated or not with other dental anomalies. The frequencies of the identified nucleotide variants were assessed in an additional cohort of patients with isolated dental agenesis (n = 147) and in 178 controls. Mutation screening showed four non-synonymous substitutions located in the highly conserved coding sequence of WNT10A in five (25%) of the 20 patients. Analysis of genotyping results revealed that three of these variants--p.Arg113Cys, p.Phe228Ile, and the newly identified p.Arg171Leu--may represent aetiological mutations underlying MLIA with associated dental anomalies. No mutations that were potentially aetiologic were identified in MSX1 and PAX9. In conclusion, this is the first report implicating coding variants in the WNT10A gene in the aetiology of MLIA. These results will require further confirmation using larger-scale studies.

PAX-9 polymorphism may be a risk factor for hypodontia: a meta-analysis.

To evaluate the association between paired box 9 (PAX9) gene polymorphisms and tooth agenesis in isolated humans, we performed a comprehensive meta-analysis. We examined 6 case-control studies, with a total of 855 hypodontia cases and 1201 healthy controls. The G allele and G carrier (AG + GG) of A1031G were positively associated with hypodontia susceptibility. Similarly, the T allele and T carrier (CT + TT) of C912T and rs12881240 in the PAX9 gene also indicated an increased risk of hypodontia. In addition, the C allele and C carrier (CG + CC) of 718C, IVS2-109, rs4904210, and rs7143727 showed no significant association with oligodontia. The G allele and G carrier (AG + GG) of IVS2-41 in the PAX9 gene were not related factors. Interestingly, the genotype (AG + GG) of IVS2-54 in the PAX9 gene may be a protective factor for oligodontia (odds ratio = 0.21, 95% confidence interval = 0.07-0.63, P = 0.005). However, no significant differences were found in the allele frequency of IVS2-54 in the PAX9 polymorphism between controls and subjects with sporadic tooth agenesis. In conclusion, our meta-analysis results revealed 4 genetic sites of the PAX9 gene involved in hypodontia cases, of which 3 sites may be risk factors and 1 may have a protective role.

Novel nonsense mutation in MSX1 in familial nonsyndromic oligodontia: subcellular localization and role of homeodomain/MH4

Nonsyndromic tooth agenesis is one of the most common anomalies in human development. Part of the malformation is inherited and is associated with paired box 9 (PAX9), msh homeobox 1 (MSX1), and axin 2 (AXIN2) mutations. To obtain a comprehensive understanding of the genetic and molecular mechanisms that underlie this genetic disease, we investigated six familial and seven sporadic Japanese cases of nonsyndromic tooth agenesis. Searches for mutations in these candidate genes detected a novel nonsense mutation (c.416G>A) in exon 1 of MSX1 from a family with oligodontia. This mutation co-segregated in the affected family members. Moreover, this mutation produced a termination codon in the first exon and therefore the gene product (W139X) was truncated at the C terminus, hence, the entire homeodomain/MH4, which has many functions, such as DNA binding, protein-protein interaction, and nuclear localization, was absent. We characterized the properties of this truncated MSX1 by investigating the subcellular localization of the mutant gene product in transfected cells. The wild-type MSX1 localized exclusively at the nuclear periphery of transfected cells, whereas the mutant MSX1 was stable but localized diffusely throughout the whole cell. These results indicate that W139X MSX1 is responsible for tooth agenesis.

Novel PAX9 mutation associated with syndromic tooth agenesis

Tooth agenesis is the most common anomaly of dental development. The purpose of the present study was to identify the causative mutation(s) in a family with a syndromic form of hypodontia. The male proband lacked 19 permanent teeth and showed defects of hair, but lacked ectodermal symptoms of skin and nails. Direct sequencing of the coding regions, including exon/intron boundaries of the msh homeobox 1 (MSX1), paired box 9 (PAX9), ectodysplasin A (EDA), and wingless-type MMTV integration site family, member 10 (WNT10A) genes, was carried out in affected family members. All identified nucleotide variations were tested in 200 healthy individuals using high-resolution melting (HRM) curve analysis to exclude the possibility that they represent rare polymorphisms. A novel heterozygous c.59delC mutation, segregating in the autosomal-dominant model, was identified in the PAX9 gene of the proband and the family members studied. This one-nucleotide deletion, located in a highly conserved paired box sequence, resulted in a frameshift (p.Pro20Argfs65) and in premature termination of translation, yielding a truncated protein 258 amino acids shorter than the wildtype protein. No pathogenic mutations were found in the MSX1, EDA, and WNT10A genes. In conclusion, the novel PAX9 deletion might be responsible for tooth agenesis and trichodysplasia in the investigated family. This c.59delC mutation potentially leads to PAX9 transcription factor haploinsufficiency.

TGF-β1 suppression of microRNA-450b-5p expression: a novel mechanism for blocking myogenic differentiation of rhabdomyosarcoma

Transforming growth factor beta 1 (TGF-β1) is the most potent inhibitor of myogenic differentiation (MyoD) of rhabdomyosarcoma (RMS); however, the underlying mechanisms of this inhibition remain unclear. In this study, we identified novel TGF-β1-related microRNAs (miRNAs); among these, miR-450b-5p is significantly regulated by TGF-β1. We provide evidence that TGF-β1 exerts it function by suppressing miR-450b-5p. Both in cultured cells and tumor implants, miR-450b-5p significantly arrested the growth of RMS and promoted its MyoD. Utilizing a bioinformatics approach, we identified miR-450b-5p target mRNAs. Among these candidates, only the expression of ecto-NOX disulfide-thiol exchanger 2 (ENOX2) and paired box 9 (PAX9) was augmented by miR-450b-5p knockdown examined by western blot; the engineered inhibition antagonized TGF-β1-mediated differentiation inhibition. Furthermore, we found that the Smad3 and Smad4 pathways, but not Smad2, are the principal mediator of TGF-β1 suppression of miR-450b-5p. Taken together, these results suggest that disrupting the TGF-β1 suppression of miR-450b-5p, or knockdown of ENOX2 and PAX9, are effective approaches in inducing RMS MyoD.