Animals ; Humans ; SOXE Transcription Factors ; genetics ; Waardenburg Syndrome ; etiology ; genetics

Objective: To screen and analyze the mutations of MITF gene in two children of type Ⅱ Waardenburg syndrome (WS2) from different families in Yunnan,China,and to explore the possible molecular pathogenesis. Methods: With informed consent, medical history collection, physical examinations, audiological evaluation, and high resolution computer tomography (HRCT) scan of temporal bone were performed on the two WS2 probands and their family members. Genomic DNA was extracted from peripheral blood of all individuals. The coding regions including all exons, part of introns and promoters of MITF, PAX3, SOX10, SNAI2, END3, ENDRB, and KITLG genes were sequenced by high-throughput sequencing. According to the results of high-throughput sequencing, pathogenic mutations detected in the probands and their parents were verified by Sanger sequencing. Results: The proband 1 carried c.641_643delGAA mutation in the 7th exon of MITF gene, which was a frame-shift mutation resulting in an amino acid change of p.214delR. It was a de novo mutation as the parents of proband 1 showed no variation on this site. The proband 2 carried heterozygous loss of the large fragment ranging from exon 1 to exon 9 of MITF gene, which defected the function of MITF protein. Conclusion: Genetic examinations provide important evidence for diagnosis of Waardenburg syndrome. Heterozygous mutation c.641_643delGAA and heterozygous loss of the large fragment ranging from exon 1 to exon 9 of MITF gene might be the molecular pathogenesis of the two WS2 probands in this study.
Asians/genetics* ; Child ; China ; Humans ; Mutation ; Pedigree ; SOXE Transcription Factors/genetics* ; Waardenburg Syndrome/genetics*

OBJECTIVETo explore the molecular mechanism of Waardenburg syndrome type II (WS2) resulting from SOX10 gene mutation E248fs through in vitro experiment.

METHODS293T cells were transiently transfected with wild type (WT) SOX10 and mutant type (MT) E248fs plasmids. The regulatory effect of WT/MT SOX10 on the transcriptional activity of MITF gene and influence of E248fs on WT SOX10 function were determined with a luciferase activity assay. The DNA binding capacity of the WT/MT SOX10 with the promoter of the MITF gene was determined with a biotinylated double-stranded oligonucleotide probe containing the SOX10 binding sequence cattgtc to precipitate MITF and E248fs, respectively. The stability of SOX10 and E248fs were also analyzed.

RESULTSAs a loss-of-function mutation, the E248fs mutant failed to transactivate the MITF promoter as compared with the WT SOX10 (P<0.01), which also showed a dominant-negative effect on WT SOX10. The WT SOX10 and E248fs mutant were also able to bind specifically to the cattgtc motif in the MITF promoter, whereas E248fs had degraded faster than WT SOX10.

CONCLUSIONDespite the fact that the E248fs has a dominant-negative effect on SOX10, its reduced stability may down-regulate the transcription of MITF and decrease the synthesis of melanin, which may result in haploinsufficiency of SOX10 protein and cause the milder WS2 phenotype.

Humans ; Microphthalmia-Associated Transcription Factor ; genetics ; Promoter Regions, Genetic ; SOXE Transcription Factors ; genetics ; Waardenburg Syndrome ; etiology ; genetics

The transcription factor SOX10, as a major actor in the development of the neural crest, plays a key role in the maintenance of progenitor cell multipotency, lineage specification, and cell differentiation. Abnormalities of neural crest development in humans lead to a number of genetic diseases known as neurocristopathies or neural crest disorders. The mutation of SOX10 can cause Kallmann syndrome (KS), which is a clinically and genetically heterogeneous condition and defined by the association between anosmia and hypogonadotropic hypogonadism due to incomplete migration of neuroendocrine gonadotropin-releasing hormone (GnRH) cells along the olfactory, vomeronasal, and terminal nerves. Since then, there have been a number of related reports that mutation of SOX10 will lead to KS with deafness. This review focuses on the SOX10 gene and the advances in the diagnosis and genetic studies of KS with deafness caused by the mutatuin of SOX10.
Cell Differentiation ; Deafness ; genetics ; Gonadotropin-Releasing Hormone ; Humans ; Hypogonadism ; Kallmann Syndrome ; genetics ; Mutation ; genetics ; SOXE Transcription Factors ; genetics

OBJECTIVE: To explore the genetic basis for a patient with syndromic hearing loss. METHODS: Genomic DNA of the patient was extracted, for which 127 deafness-related genes were enriched with a chip. Following next generation sequencing, pathogenic loci in exonic regions were analyzed through comparison against the databases. Genotype of her fetus for the suspected site was determined by testing the amniotic fluid sample. qPCR method was applied to verify the deletion of a large fragment. RESULTS: The proband was diagnosed with Waardenburg syndrome type 2, and had harbored a novel heterozygous deletion of the exons 3 and 4 of the SOX10 gene. Her fetus was found to carry the same deletion and presented with blue eyes and deafness after birth. CONCLUSION Waardenburg syndrome type 2 due to SOX10 gene deletion may feature autosomal dominant inheritance with incomplete penetrance. The deletion of exons 3 and 4 of the SOX10 gene probably underlies the disease in this family.
Eye Color ; Female ; Hearing Loss ; Humans ; Mutation ; Pedigree ; Pregnancy ; Prenatal Diagnosis ; SOXE Transcription Factors ; genetics ; Waardenburg Syndrome

OBJECTIVE: To explore the genetic basis for a Chinese pedigree featuring congenital profound syndromic deafness and chronic constipation, and provide prenatal diagnosis for a high-risk fetus. METHODS: Whole-exome sequencing was carried out to analyze the sequences of genes associated with hereditary deafness, and multiplex ligation-dependent probe amplification (MLPA) was used to verify the candidate variant in the proband's parents and the fetus. RESULTS: The proband was found to have harbored a heterozygous deletion of SOX10, a pathogenic gene associated with Waardenburg syndrome type 4C (WS4C). The same deletion was found in her mother (with profound syndromic deafness and chronic constipation) and the fetus, but not in her father with normal hearing. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG) and Association for Molecular Pathology (AMP), the SOX10 gene deletion was predicted to be a pathogenic variant (PVS1+PM2_Supporting+PP1+PP4). CONCLUSION The pedigree was diagnosed with WS4C, which has conformed to an autosomal dominant inheritance. Deletion of the entire SOX10 gene, as a loss-of-function variant, probably underlay its pathogenesis. Above finding has facilitated genetic counseling and prenatal diagnosis for this family.
Humans ; Female ; Pregnancy ; Pedigree ; Waardenburg Syndrome/genetics* ; East Asian People ; Genetic Testing ; Prenatal Diagnosis ; Hearing Loss, Sensorineural/genetics* ; Deafness/genetics* ; Mothers ; Constipation/genetics* ; Mutation ; SOXE Transcription Factors/genetics*

OBJECTIVETo perform genetic analysis for 7 patients with Waardenburg syndrome.

METHODSPotential mutation of MITF, PAX3, SOX10 and SNAI2 genes was screened by polymerase chain reaction and direct sequencing. Functions of non-synonymous polymorphisms were predicted with PolyPhen2 software.

RESULTSSeven mutations, including c.649-651delAGA (p.R217del), c.72delG (p.G24fs), c.185T>C (p.M62T), c.118C>T (p.Q40X), c.422T>C (p.L141P), c.640C>T (p.R214X) and c.28G>T(p.G43V), were detected in the patients. Among these, four mutations of the PAX3 gene (c.72delG, c.185T>C, c.118C>T and c.128G>T) and one SOX10 gene mutation (c.422T>C) were not reported previously. Three non-synonymous SNPs (c.185T>C, c.128G>T and c.422T>C) were predicted as harmful.

CONCLUSIONGenetic mutations have been detected in all patients with Waardenburg syndrome.

Adolescent ; Child ; Female ; Humans ; Male ; Microphthalmia-Associated Transcription Factor ; genetics ; Mutation ; PAX3 Transcription Factor ; Paired Box Transcription Factors ; genetics ; Polymorphism, Single Nucleotide ; SOXE Transcription Factors ; genetics ; Waardenburg Syndrome ; genetics

OBJECTIVETo study the exogenous expression and subcellular localization of wild type (WT) and mutant SOX10 proteins in vitro through generation of expression plasmids in order to reveal the pathogenesis of Waardenburg syndrome (WS).

METHODSThe plasmids pECE-SOX10 and pCMV-Flag were ligated after they were subjected to double enzyme digestion using molecular cloning technique to generate recombinant eukaryotic expression plasmid pCMV-SOX10-Flag, which was as a template to generate expression plasmids for novel mutations G37fs, G38fs and E248fs of the SOX10 gene. The constructs were verified by direct sequencing. NIH3T3 cells were transiently transfected with the expression plasmids of wide type SOX10, G37fs, G38fs and E248fs, respectively. The exogenous expression of WT SOX10 protein and mutant G37fs, G38fs and E248fs proteins were analyzed using Western blot assay, while their subcellular distribution were observed with an immunofluorescence assay.

RESULTSThe DNA sequences of expression plasmids for SOX10 and its mutant G37fs, G38fs and E248f were all correct. Both WT and mutant SOX10 proteins were detected at the expected site. WT SOX10 and E248fs proteins have only localized in the nucleus, whereas G37fs and G38fs proteins showed aberrant localization in both cytoplasm and nucleus.

CONCLUSIONRecombinant eukaryotic expression plasmids for the SOX10 gene and its mutants were successfully constructed. Preliminary analysis showed that the mutations have affected the subcellular distribution of WT SOX10 proteins, which has laid a basis for further study of the molecular mechanism of WS caused by SOX10 gene mutations.

Animals ; Base Sequence ; Humans ; Mice ; Molecular Sequence Data ; Mutation ; NIH 3T3 Cells ; Plasmids ; Recombination, Genetic ; SOXE Transcription Factors ; genetics ; Waardenburg Syndrome ; genetics

OBJECTIVETo explore the molecular etiology of two pedigrees affected with type II Waardenburg syndrome (WS2) and to provide genetic diagnosis and counseling.

METHODSBlood samples were collected from the proband and his family members. Following extraction of genomic DNA, the coding sequences of PAX3, MITF, SOX10 and SNAI2 genes were amplified with PCR and subjected to DNA sequencing to detect potential mutations.

RESULTSA heterozygous deletional mutation c.649_651delAGA in exon 7 of the MITF gene has been identified in all patients from the first family, while no mutation was found in the other WS2 related genes including PAX3, MITF, SOX10 and SNAI2.

CONCLUSIONThe heterozygous deletion mutation c.649_651delAGA in exon 7 of the MITF gene probably underlies the disease in the first family. It is expected that other genes may also underlie WS2.

Base Sequence ; DNA Mutational Analysis ; Exons ; genetics ; Family Health ; Female ; Genetic Predisposition to Disease ; genetics ; Heterozygote ; Humans ; Male ; Microphthalmia-Associated Transcription Factor ; genetics ; Molecular Sequence Data ; Mutation ; PAX3 Transcription Factor ; Paired Box Transcription Factors ; genetics ; Pedigree ; Polymerase Chain Reaction ; SOXE Transcription Factors ; genetics ; Sequence Deletion ; Snail Family Transcription Factors ; Transcription Factors ; genetics ; Waardenburg Syndrome ; classification ; diagnosis ; genetics