A novel mutation of exon 7 in growth hormone receptor mRNA in a patient with growth hormone insensitivity syndrome and neurofibromatosis type I

Growth hormone insensitivity syndrome (GHIS), a genetic disease characterized by growth retardation combined with high serum concentration of growth hormone (GH) and low insulin-like growth factor 1 (IGF-1) levels, can be caused by mutations in the GH receptor (GHR) gene. We investigated the molecular defects in the GHR gene in a patient with neurofibromatosis type 1 (NF-1). The patient, a 2-year-old boy with NF-1, was assessed on his short stature by auxological, biochemical and molecular studies. Height of the patient and his family members were measured and compared to normal control. Serum concentrations of GH, IGF-1 and IGF-binding protein 3 (IGFBP3) in the patient were measured during a GH stimulation test. We examined the GHR gene in the patient and his parents. Genomic DNA and mRNA of the GHR gene were extracted from peripheral lymphocytes. All the exons and the flanking regions of the GHR gene were amplified by PCR, and directly sequenced. The patient's height was 75 cm (-2.89 SDS) with gradually reducing growth velocity, while the heights of the other family members were within the normal range. The GH stimulation test revealed that serum GH concentrations in the patient were much higher than those in the control group, and serum IGF-1 and IGFBP3 levels were extremely low. There was no germline mutation in the exons or the flanking regions of the patient's GHR gene. Interestingly, a deletion of 166 bases of exon 7 in the GHR mRNA was found, and it was suggested that the novel mutation resulted in premature termination (M207 fs. X8). This mutation decreases GH binding affinity to the GHR, and, thus, would be responsible for growth retardation.     

完全型雄激素不敏感综合征雄激素受体基因突变分析

目的 对完全型雄激素不敏感综合征一家系雄激素受体(androgen receptor,AR)基因进行突变检测;并对发现突变的基因进行分析.方法 应用PCR扩增、DNA序列测定等技术分析所有AR基因外显子及其邻近DNA序列片段;应用核苷酸内切酶诊断方法观察其是否存在于正常人群;应用跨物种比对方法探讨突变所在位置的保守性.结果 3例患者AR基因第4外显子均发生E681D(GAG→GAT)错义突变,患者母亲为此突变杂合子携带者;患者父亲未见异常;正常人群未发现AR基因E681D突变;681位谷氨酸在不同物种间高度保守.结论 AR基因E681D(GAG→GAT)突变可能是导致完全型雄激素不敏感综合征新的突变方式.     

Novel androgen receptor gene mutations in Australian patients with complete androgen insensitivity syndrome

We have identified androgen receptor (AR) gene mutations in eight Australian subjects with complete androgen insensitivity syndrome (AIS). Four individuals, from three families, have novel mutations that introduce premature termination codons. Two siblings have the nonsense mutation Glu681X, and another subject has the nonsense mutation p.Ser884X. The other subject has a CA insertion at codon 829 (c.2847_2848insCA), causing a frameshift mutation that introduces four nonsense amino acids prior to a Stop codon. All the termination codons occur in the ligand binding domain, and cause reduced androgen binding in patient genital skin fibroblasts. Four further patients have missense mutations. One subject has two different mutations, p.Ala645Asp in the hinge region of the receptor, and p.Arg752Gln in the ligand binding domain. Both these mutations have previously been reported in patients with AIS, but the combination of these two mutations in one subject is unique. Another subject has a novel c.2533G>C transversion at the first nucleotide in exon 5, introducing the amino acid change p.Gly724Ala at a highly conserved residue in the ligand binding domain. Androgen binding is normal in fibroblasts from this subject, although other point mutations at this amino acid totally abolish binding. Two other subjects have mutations previously described as causing AIS, namely p.Arg779Trp and p.Val889Met substitutions in the ligand binding domain of the receptor. The p.Arg779Trp mutation is associated with the detection of a truncated AR protein in this patient's fibroblasts, suggesting the mutation renders the receptor susceptible to proteolysis.     

The growth hormone receptor gene mutation of a japanese patient with laron syndrome

Summary Deletions and point mutations of the growth hormone (GH) receptor gene (GHR) have been identified in patients with Laron syndrome. We report the first detection of theGHR mutation among Japanese patients with Laron syndrome. Using the Japanese female patient’s genomic DNA as a template, all exons and flanking portions of introns ofGHR were amplified by polymerase chain reaction (PCR). Sequencing of the PCR products showed that the patient was homozygous for a G to A substitution at the first position of intron 4. This substitution was same as that detected in a Spanish patient and a north European patient. The base change occurred at the 5′ splice consensus sequence of intron 4, resulting in the abolition of aBanI restriction site. Since this substitution was not detected by aBanI restriction analysis in 85 control individuals, it is more likely a disease-related splice mutation than a polymorphism. The mutation in our patient was predicted to destroy the original 5′ splice site of intron 4 ofGHR and to produce a new cryptic splice site, leading to abnormal mRNA processing and a lack of GH binding activity of GH-binding protein (GHBP).     

Five novel androgen receptor gene mutations associated with complete androgen insensitivity syndrome

Mutations in the androgen receptor (AR) gene result in androgen insensitivity syndrome (AIS). We have identified five novel mutations that result in a complete loss in AR function and are associated with complete AIS. The mutations span all three AR major functional domains. In two cases, the loss of AR function could be explained on the basis of the current knowledge of AR molecular structure and function. N-terminal mutation c.256C>T (p.Gln86X) leads to an early stop codon and abolishes all DNA and ligand binding. The DNA-binding domain mutation c.1685G>A (p.Cys562Tyr) is located in the N-terminal part of the first zinc finger; a mutation in this position is likely to impair the association of the mutated AR with the androgen response element of target genes. The splice site mutation at intron 2/exon 3 junction (c.1766-1G>A) is shown to lead to c.1765_1766 ins69 (p.[Gly589_Lys590ins23;Gly589Glu]). The two novel ligand-binding domain mutations identified were recreated by site-directed mutagenesis. Both mutations c.2171G>T (p.Gly724Val) and c.2435T>C (p.Leu812Pro) abolished AR ligand binding and severely impaired AR mediated transactivation. Residue p.Gly724 is located in the ligand binding domain, between helices 3 and 4. This region is known to be involved not only in ligand binding but also in AR N/C-terminal interactions. The mutation p.Leu812Pro is located in the C-terminal end of helix 8. This domain is highly conserved and critical for ligand binding. This study extends current understanding of AR mutations associated with CAIS.     

Biallelic deletions of the Waardenburg II syndrome gene,SOX10, cause a recognizable arthrogryposis syndrome

Random mating in the general population tends to limit the occurrence of homozygous and compound heterozygous forms of dominant hereditary disorders. Certain phenotypes, the most recognized being skeletal dysplasias associated with short stature, lead to cultural interaction and assortative mating. To this well‐known example, may be added deafness which brings together individuals with a variety of deafness genotypes, some being dominant. Waardenburg syndrome is one such autosomal dominant disorder in which affected individuals may interact culturally because of deafness. Biallelic genetic alterations for two Waardenburg genes, PAX3 and MITF have been previously recognized. Herein, we report biallelic deletions in SOX10, a gene associated with Waardenburg syndromes type II and IV. The affected fetuses have a severe phenotype with a lack of fetal movement resulting in four‐limb arthrogryposis and absence of palmar and plantar creases, white hair, dystopia canthorum, and in one case cleft palate and in the other a cardiac malformation.     

Androgen receptor gene mutations identified by SSCP in fourteen subjects with androgen insensitivity syndrome.

The androgen insensitivity syndrome (AIS) is a disorder of male sexual development resulting in a wide range of clinical phenotypes. AIS is classified into two phenotypic forms: complete (CAIS) and partial (PAIS). To determine the molecular basis of the phenotypic diversity in AIS, we have studied 27 subjects (13 CAIS, 14 PAIS), spanning the full range of AIS phenotypes. We report the results of a mutation screen of the androgen receptor gene. The coding regions of the gene were amplified by the polymerase chain reaction and screened for single strand conformation polymorphisms to identify mutations. This was followed by DNA sequencing of putative mutant segments. Androgen receptor gene mutations were identified in nine CAIS and five PAIS subjects. Two of the CAIS mutations in exon A resulted in frameshifts. A third CAIS mutation resulted in the deletion of a single amino acid from the ligand binding domain of the receptor. All other mutations caused single amino acid substitutions in the ligand binding domain. These results suggest that mutations affecting the ligand binding domain of the androgen receptor are the most frequent cause of AIS, although some cases of PAIS may be the result of other, as yet undefined, genetic lesions.     

Noonan-like syndrome with loose anagen hair associated with growth hormone insensitivity and atypical neurological manifestations

Noonan-like syndrome with loose anagen hair (NS/LAH; OMIM 607721), recently related to the invariant c.4A>G missense change in SHOC2, is characterized by features reminiscent of Noonan syndrome. Ectodermal involvement, short stature associated with growth hormone (GH) deficiency (GHD), and cognitive deficits are common features. We report on a patient with molecularly confirmed NS/LAH exhibiting severe short stature associated with GH insensitivity (GHI), and chronic complex tics, a neurological feature never described before in this syndrome. IGF1 generation test revealed only a blunted increase in IGF1 after exogenous GH treatment, revealing mild GH insensitivity associated with proper STAT5 activation. Most common causes of secondary tics in childhood were excluded.     

Gross deletions in TCOF1 are a cause of Treacher-Collins-Franceschetti syndrome

Treacher-Collins-Franceschetti syndrome (TCS) is an autosomal dominant craniofacial disorder characterised by midface hypoplasia, micrognathia, downslanting palpebral fissures, eyelid colobomata, and ear deformities that often lead to conductive deafness. A total of 182 patients with signs consistent with a diagnosis of TCS were screened by DNA sequence and dosage analysis of the TCOF1 gene. In all, 92 cases were found to have a pathogenic mutation by sequencing and 5 to have a partial gene deletion. A further case had a novel in-frame deletion in the alternatively spliced exon 6A of uncertain pathogenicity. The majority of the pathogenic sequence changes were found to predict premature protein termination, however, four novel missense changes in the LIS1 homology motif at the 5' end of the gene were identified. The partial gene deletions of different sizes represent ~5.2% of all the pathogenic TCOF1 mutations identified, indicating that gene rearrangements account for a significant proportion of TCS cases. This is the first report of gene rearrangements resulting in TCS. These findings expand the TCOF1 mutation spectrum indicating that dosage analysis should be performed together with sequence analysis, a strategy that is predicted to have a sensitivity of 71% for patients in whom TCS is strongly suspected.     

Multi-exon deletions of the FBN1 gene in Marfan syndrome

Background  

Mutations in the fibrillin -1 gene (FBN1) cause Marfan syndrome (MFS), an autosomal dominant multi-system connective tissue disorder. The 200 different mutations reported in the 235 kb, 65 exon-containing gene include only one family with a genomic multi-exon deletion.     

Somatic mosaic deletions involving SCN1A cause Dravet syndrome

Somatic mosaicism in single nucleotide variants of SCN1A is known to occur in a subset of parents of children with Dravet syndrome (DS). Here, we report recurrent somatic mosaic microdeletions involving SCN1A in children diagnosed with DS. Through the evaluation of 237 affected individuals with DS who did not show SCN1A or PCHD19 mutations in prior sequencing analyzes, we identified two children with mosaic microdeletions covering the entire SCN1A region. The allele frequency of the mosaic deletions estimated by multiplex ligation‐dependent probe amplification and array comparative genomic hybridization was 25–40%, which was comparable to the mosaic ratio in lymphocytes and buccal mucosa cells observed by fluorescence in situ hybridization analysis. The minimal prevalence of SCN1A mosaic deletion is estimated to be 0.9% (95% confidence level: 0.11–3.11%) of DS with negative for SCN1A and PCDH19 mutations. This study reinforces the importance of somatic mosaicism caused by copy number variations in disease‐causing genes, and provides an alternative spectrum of SCN1A mutations causative of DS. Somatic deletions in SCN1A should be considered in cases with DS when standard screenings for SCN1A mutations are apparently negative for mutations.

     

Spectrum of growth hormone receptor mutations and associated haplotypes in Laron syndrome

Laron syndrome is a rare autosomal recessive disorder characterizedby resistance to growth hormone (GH). In 10 patients of differentethnic origins, we have analyzed all the GH receptor (GHR)-codingexons along with their splice junctions and 6 intragenic polymorphicsites defining several GHR gene haplotypes. This allowed usto identify the mutations in the 20 chromosomes studied andto describe a new GHR haplotype. Eleven different mutationsassociated with various GHR haplotypes were observed; they included3 nonsense mutations, 3 splice defects and 5 missense mutations.Of the 11 mutations, 8 were novel. All the mutations involvedthe exoplasmic domain of the receptor and all the missense mutationswere clustered in a short polypeptide segment. Most of the missensemutations affected residues conserved among GHRs from differentspecies and the related molecules that belong to the cytokinereceptor superfamily. Adding to the 5 mutations so far described,these findings illustrate the allelic heterogeneity of thissyndrome and document the independent origin of the moleculardefects, all features of clinical relevance for genetic counselling.