[Analysis of TGFBI gene mutation in a pedigree affected with corneal dystrophy].

OBJECTIVE: To detect potential mutation in a large Chinese pedigree affected with congenital corneal dystrophy. METHODS: Two patients from the pedigree were subjected to whole exome sequencing to determine the candidate gene. Suspected mutation was verified in 13 additional members by directional Sanger sequencing. Ccorrelation between genotype and phenotype was explored. RESULTS: A missense mutation, c.1877A>C (p.His626Pro), was detected in exon 14 of the TGFBI gene in 8 patients from the pedigree, but not in five unaffected members and 100 unrelated healthy controls. Respectively, the mutation was predicted as "affecting protein function", "probably damaging" and "disease causing" by SIFT, PolyPhen-2 and MutationTaster. CONCLUSION: The c.1877A>C mutation of the TGFBI gene probably underlies the disease in this pedigree.

[Analysis of chromosome regions 8q11.1-q13.3, 1q32-q34.3 and 14q31.1-q13.3 in a Chinese family with congenital preauricular fistula].

OBJECTIVE: To identify the candidate chromosomal region for congenital preauricular fistula (CPF) through analysis of an affected Chinese family. METHODS: Conventional linkage analysis using short tandem repeats (STR) markers was performed to investigate three chromosomal regions 8q11.1-q13.3, 1q32-q34.3 and 14q31.1-q31.3. RESULTS: None of 16 STRs could attain a LOD score of more than -2.0 (theta=0). Therefore, the three regions were all excluded as the candidate region for the disease. CONCLUSION: CPF features high genetic heterogeneity. The family may have a causative gene elsewhere. Whole-genome-based study is needed to identify its genetic etiology.

DNA methylation age acceleration is associated with age of onset in Chinese spinocerebellar ataxia type 3 patients.

Spinocerebellar ataxia type 3 (SCA3), also known as Machado Joseph disease (MJD), is a common dominantly inherited ataxia, and has heterogeneous clinical features and variable age of onset, ranging from 10 to 78 years. Repeats variability of ATXN3, HTT, ATN1 and ATXN2 can explain partially but not fully SCA3 age of onset heterogeneity. Aging is a reported modifier of SCA3 severity and closely linked to DNA methylation (DNAm). DNAm age acceleration was associated with disease risk and/or variable disease phenotypes in several repeat associated neurodegenerative diseases (such as Huntington's disease and Amyotrophic lateral sclerosis). To understand if DNAm age acceleration is associated with SCA3 age of onset, we performed a genome-wide DNAm study of a Chinese SCA3 family with variable age of onset and clinical presentations. All patients showed unsteady gait, deterioration of extremities coordination, speech (dysarthria) and swallowing problems (dysphagia, choking on eating and/or drinking) and oculomotor abnormalities, with variable age of onset ranging from 27 to 52 years. We found that DNAm age acceleration is associated with age of onset (p-value = 0.0023, B = -1.26), suggesting that every 5 year increase in DNAm-age acceleration is corresponding to a 6.3 year earlier disease onset. This association remains significant after the adjustment to ATXN3 CAG repeats (adjusted p-value = 0.037, adjusted B = -1.0). In an independent SCA3 cohort (n = 40), we also observed the association between DNAm age acceleration and age of onset (adjusted p-value = 0.007, adjusted B = -0.69). Of note, we found no significant association between DNAm of single-CpG locus and/or CpG-SNPs and SCA3 age of onset in the current family or the SCA3 cohort. Our findings suggested that DNAm age acceleration might be a SCA3 age of onset modifier, and encourage further investigations in extended SCA3 cohorts to clarify the role of epigenetic aging in modifying disease onset.

[Genetic analysis of a Chinese pedigree with split hand and foot malformation].

OBJECTIVE: To analyze the clinical manefestation and genetic basis of split hand and foot malformation (SHFM) in a Chinese pedigree. METHODS: The affected people in the family were checked by X-rays. Eighteen patients provided their peripheral blood, and the genomic DNA of the samples was extracted. The linkage and haplotype analysis were carried out using the microsatellite markers, and the limb malformation related gene Dactylin (DAC) including the coding region, exon-intron boundaries and part of promoter region was sequenced. RESULTS: Most members of the family with the disease phenotype showed absence or hypoplasia of the index finger, and absence or 3-4 syndactyly of the middle finger. The degree of abnormality in feet was severer than that in hands. All phenotypes of the patients display the basic characters of SHFM. Since the maximum two point LOD score of the D10S192 was 3.50 (theta=0.00), the SHFM in this pedigree can be categorized to the SHFM3. The haplotype analysis of recombination events revealed the candidate locus to a 21cM region between D10S185 and D10S1693. No mutation was found by the sequencing result of DAC gene. CONCLUSION: Through the analysis of phenotype of the patients, the typical SHFM disease can be confirmed. The linkage and haplotype analysis demonstrated that the 21cM region in 10q23-q26 locus was the major cause to the disease in this pedigree. The mutation of DAC gene can be excluded from cause of SHFM3 phenotype.

Systematic screening for polymorphisms in the CYP3A4 gene in the Chinese population.

OBJECTIVES: Cytochrome P450 3A4 (CYP3A4) is a major CYP enzyme in the liver and intestine. It is involved in the metabolism of over 50% of all drugs currently in use. The present study was designed to determine the genetic basis of CYP3A4 variability. METHODS: Single nucleotide polymorphism (SNP) analysis of the CYP3A4 gene was performed on 60 healthy Chinese subjects consisting of 20 Han, 30 She and ten Dong subjects, using direct sequencing. Linkage disequilibrium, haplotype inference and Hardy-Weinberg equilibrium were also determined for these samples. RESULTS: A total of 20 SNPs were found in the CYP3A4 gene, including 11 known SNPs and nine novel SNPs. The known SNPs detected in our study were CYP3A4*1B, CYP3A4*1G, CYP3A4*10, CYP3A4*13, CYP3A4*14, CYP3A4*15, CYP3A4*17, CYP3A4*18, rs3091339, rs3091430 and rs28371761, and the novel SNPs were -658 A-->C, G27A (E10K), T48A, G14284A (G167D), A15623G (N191D), C15635A (L196I), T15656C (F203L), G14199A (intron 5) and C15566T (intron 6). The allelic frequencies found in our sample varied from 1-37%. The novel SNPs detected in the CYP3A4 gene suggest that the Chinese population has different patterns of allele frequency compared with other populations. CONCLUSION: Several SNPs were detected in the CYP3A4 gene. The study of genetic variants in CYP3A4 may have an important significance for the understanding of genotype and phenotype relationships.

A novel mutation of PAX3 in a Chinese family with Waardenburg syndrome.

PURPOSE: The molecular characterization of 34 members of a Chinese family, with 22 members in four generations, affected with Waardenburg syndrome (WS1). METHODS: A detailed family history and clinical data were collected. A genome-wide scan by two-point linkage analysis using more than 400 microsatellite markers in combination with haplotype analysis was performed. Mutation screening was carried out in the candidate gene by sequencing of amplified products. RESULTS: A maximum two-point lod score of 6.53 at theta = 0.00 was obtained with marker D2S2248. Haplotype analysis placed the WS1 locus to a 45.74 cM region between D2S117 and D2S206, in close proximity to the PAX3 gene on chromosome 2q35. Mutation screening in PAX3 identified a 701T > C mutation which converted a highly conserved Leu to Pro. This nucleotide alteration was neither seen in unaffected members of the family nor found in 50 unrelated control subjects. CONCLUSIONS: The present study identified a novel 701T > C mutation in PAX3. The mutation observed in this family highlights the phenotypic heterogeneity of the disorder.

[Mutation analysis of 12S rRNA and tRNA-Ser(UCN) genes in a large Chinese family with maternally inherited nonsyndromic hearing loss by intermarriage].

OBJECTIVE: To identify the possible mutation at possible sites in different mitochondrial genes that leads to hearing loss in a large Chinese pedigree. METHODS: Blood samples from a Hunan pedigree were obtained with informed consent. Genomic DNA was extracted from peripheral blood leukocytes using kit. The target fragments were amplified and detected by polymerase chain reaction (PCR) and directly sequencing respectively. RESULTS: The result of direct sequencing revealed the A1555G mutation in 12S rRNA gene was inherited in this pedigree and no one has A7445G mutation or other mutations in its neighborhood region. CONCLUSION: Sequence analysis confirmed that the pedigree carries the A1555G mutation. With some members ever exposure of aminoglycoside antibiotics, mutation of A1555G may play a pivotal role in the pathogenesis of hearing loss in the large pedigree.

[Mutation detection of COL1A1 gene in a pedigree with osteogenesis imperfecta].

Osteogenesis imperfecta (OI) is heritable bone fragility,which is inherited as an autosomal dominant trait clinical presentation. Clinical symptom, in general, is dominantly inherited OI with blue sclerae, hearing loss and mild-moderate skeletal deformity. Genetic loci of OI have been mapped to17q21.31-q22 and 7q22.1, in which COL1A1 and COL1A2 are known to be the causal genes. In this work,we performed linkage analysis in a kindred with autosomal dominant hereditary OI. A tight linkage to the markers on chromosome 17q21.31-q22 (maximum two-point lod score: 9.31 at theta = .00) was observed. Sequence analysis of COL1A1 revealed a single-base mutation that converted the consensus sequence at the 5' end of intron 26 from GT to AT to form an abnormal splicing site leading to OI.

[A family with nonsyndromic hearing impairment caused by intermarry].

Deafness is the most prevalent sensory system impairment in human, about 70 % of genetic deafness belongs to nonsyndromic hearing impairment. It was estimated that the total number of genes involved in nonsyndromic hereditary deafness was over 100. So far, approximate 80 loci have been mapped to human chromosome, and 23 genes have been identified. In this paper, a family with nonsyndromic hearing impairment caused by intermarry was reported. There were 13 sufferers in two generations. Deduced from genetic analysis, neither autosomal dominant nor autosomal recessive inheritance was identified in this family, which suggested that hearing impairment in the family was probably caused by mitochondrial mutations.

A novel insertion mutation in the FOXL2 gene is detected in a big Chinese family with blepharophimosis-ptosis-epicanthus inversus.

Blepharophimosis-ptosis-epicanthus inversus syndrome (BPES), an autosomal dominant syndrome in which an eyelid malformation is associated (type I) or not (type II) with premature ovarian failure (POF), has recently been ascribed to mutations in the forkhead transcription factor 2 (FOXL2) gene. In this work, we reveal a novel insertion mutation in the 3'UTR of the FOXL2 gene in a big Chinese family which is to our knowledge the first BPES (type II) family reported in China. It is the first time that a 3'UTR mutation in the FOXL2 gene has ever been found to demonstrate a close correlation between genotype and BPES. Our result gains a greater insight into the function of 3'UTR in the FOXL2 gene.

[Genetic analysis of a Chinese pedigree with congenital synpolydactyly].

Syndactyly is a limb malformation that shows a characteristic manifestation in both hands and feet. This condition is inherited as an autosomal dominant trait with reduced penetrance. Clinical presentation, in general, is complete or partial webbing between 3rd and 4th fingers. Syndactyly type I, II and III were mapped to 2q34-36, 2q31-q32 and 6q21-23.2 respectively. Syndactyly type II is named as synpolydactyly (SPD). Expansion of a polyalanine tract in the HOXD13 gene is known to cause synpolydactyly. HOXD13 gene locates in the HoxD complex. Nine homologous genes (HOXD1, -D3, -D4, -D8, -D9, -D10, -D11, -D12, -D13) of HoxD complex locate on chromosome 2 in the order of HOXD1 to HOXD13, among which HOXD13 is closest to the centromere. Deletions and duplications in HoxD complex or its upstream regulator factors have been identified to cause hand heteroplasia and consequently lead to abnormity of finger number or abnormity of configuration. We performed linkage analysis in a kindred with autosomal dominant hereditary syndactyly. Tight linkage to markers on chromosome 2q31-q32 (maximum two-point lod score: 6.78 at recombination fraction theta = 0.00) was observed. Multipoint linkage analysis produced a maximum LOD score of 7.02. Haplotype construction and analysis of recombination events narrowed this locus to a 20.61 cM region between markers D2S2302 and D2S315. No mutation was found in the coding region, the intro-exon boundaries, or part of the promoter region of HOXD13. Our result demonstrates that synpolydactyly locus in the Chinese Han Population is in the region of chromosome 2q31-q32 but a different causal gene can be involved.

Systematic screening for polymorphisms within the UGT1A6 gene in three Chinese populations and function prediction through structural modeling.

AIMS: To date, there have been relatively few studies on the UGT1A6 gene in the Chinese population. The present study was designed to determine the allele frequencies and haplotypes of this gene in the population and predict the candidate functional mutations. MATERIALS & METHODS: We carried out the first systematic screening of polymorphisms of the gene in an SNP analysis involving 1074 Chinese subjects from three ethnic groups, namely Han, Dong and She, using direct sequencing. We identified the putative substrate binding pocket using a homology-modeled structure and produced a practical model for predicting the function of polymorphisms in UGT1A6. RESULTS: A total of six SNPs and 10 mutations were detected including nine known and seven novel ones. The novel mutations were 73G>A (V25I), 89T>G (L30R), 222A>C, 657C>A, 773A>T (D258V), 1040A>G (N347S) and 1467C>T. In addition, we detected, for the first time in the Chinese population, SNPs 105C>T, 627G>T as well as mutations 308C>A (S103X), IVS2+15T>C and 1088C>T (P363L). Strong linkage disequilibrium was observed among 19T>G, 315A>G, 541A>G and 552A>C. There were seven haplotypes whose frequencies were more than 0.01 in one or more of the three ethnic groups. P363L in the C-terminal domain might weaken the binding of cofactor UDPGA to the domain and induce a poor metabolism genotype of UGT1A6. CONCLUSION: Our study suggests that genetic polymorphisms in UGT1A6 may contribute to interindividual and intra-ethnic differences. The results should prove helpful in the development of pharmacogenomics in China.

Screening for SNPs and haplotypes in the CYP3A7 gene in Chinese populations.

OBJECTIVE: To determine the frequencies and haplotypes of the cytochrome P450 (CYP)3A7 gene in three Chinese ethnic groups, namely, Han, She and Dong. METHODS: SNP analyses of the CYP3A7 gene were carried out on three groups of healthy Chinese subjects consisting of 539 Han, 264 She and 273 Dong subjects, using direct sequencing. Linkage disequilibrium, haplotype inference and Hardy-Weinberg equilibrium were also determined for these samples. RESULTS: Significant differences were observed in the distribution of SNP rs2257401 (CYP3A7*2), SNP 1227 T>C (Novel), SNP rs4646468 and SNP rs10211 between the Han, She and Dong groups. CONCLUSION: Some allele and haplotype frequencies show variation among groups, highlighting the need to analyze clinically relevant SNPs and haplotypes in a variety of different racial groups within the Chinese population as well as in other ethnic groups. These results suggest that genetic polymorphisms in the CYP3A7 gene in the Han, She and Dong populations may contribute to interindividual as well as intra-ethnic differences in response to the clearance of CYP3A7 substrates.