Androgen receptor genotyping in a large Australasian cohort with androgen insensitivity syndrome; identification of four novel mutations.

We genotyped the androgen receptor (AR) gene in 31 Australasian patients with androgen insensitivity syndrome (AIS). The entire coding region of AR was examined including analysis of polymorphic CAG and GGN repeats in all patients. AR defects were found in 66.7% (6/9) of patients with complete AIS (CAIS) and 13.6% (3/22) of patients with partial AIS (PAIS). A novel deletion (N858delG) leading to a premature stop codon was found in CAIS patient P1. CAIS patient P2 has a novel deletion (N2676delGAGT) resulting in a stop at codon 787. These mutations would result in inactivation of AR protein. A novel insertion of a cysteine residue in the first zinc finger of the AR DNA-binding domain (N2045_2047dupCTG) was found in CAIS patient P3. PAIS patient P4 has a novel amino acid substitution (Arg760Ser) in the AR ligand binding domain, which may impair ligand binding. Five patients were found to have previously reported AR mutations and no mutations were identified in the remaining patients.

Deletions involving long-range conserved nongenic sequences upstream and downstream of FOXL2 as a novel disease-causing mechanism in blepharophimosis syndrome.

The expression of a gene requires not only a normal coding sequence but also intact regulatory regions, which can be located at large distances from the target genes, as demonstrated for an increasing number of developmental genes. In previous mutation studies of the role of FOXL2 in blepharophimosis syndrome (BPES), we identified intragenic mutations in 70% of our patients. Three translocation breakpoints upstream of FOXL2 in patients with BPES suggested a position effect. Here, we identified novel microdeletions outside of FOXL2 in cases of sporadic and familial BPES. Specifically, four rearrangements, with an overlap of 126 kb, are located 230 kb upstream of FOXL2, telomeric to the reported translocation breakpoints. Moreover, the shortest region of deletion overlap (SRO) contains several conserved nongenic sequences (CNGs) harboring putative transcription-factor binding sites and representing potential long-range cis-regulatory elements. Interestingly, the human region orthologous to the 12-kb sequence deleted in the polled intersex syndrome in goat, which is an animal model for BPES, is contained in this SRO, providing evidence of human-goat conservation of FOXL2 expression and of the mutational mechanism. Surprisingly, in a fifth family with BPES, one rearrangement was found downstream of FOXL2. In addition, we report nine novel rearrangements encompassing FOXL2 that range from partial gene deletions to submicroscopic deletions. Overall, genomic rearrangements encompassing or outside of FOXL2 account for 16% of all molecular defects found in our families with BPES. In summary, this is the first report of extragenic deletions in BPES, providing further evidence of potential long-range cis-regulatory elements regulating FOXL2 expression. It contributes to the enlarging group of developmental diseases caused by defective distant regulation of gene expression. Finally, we demonstrate that CNGs are candidate regions for genomic rearrangements in developmental genes.

Principal mutation hotspot for central core disease and related myopathies in the C-terminal transmembrane region of the RYR1 gene.

The congenital myopathies are a group of disorders characterised by the predominance of specific histological features observed in biopsied muscle. Central core disease and nemaline myopathy are examples of congenital myopathies that have specific histological characteristics but significantly overlapping clinical pictures. Central core disease is an autosomal dominant disorder with variable penetrance which has been linked principally to the gene for the skeletal muscle calcium release channel (RYR1). Two recent reports have identified the 3' transmembrane domain of this gene as a common site for mutations. Two other studies have reported single families that have features of both central core disease and nemaline myopathy (core/rod disease) caused by mutations in RYR1. Screening of the 3' region (exons 93-105) of the RYR1 gene for mutations in 27 apparently unrelated patients with either central core disease or core/rod disease by single strand conformation polymorphism analysis and DNA sequencing identified three described and nine novel mutations in 15 patients.

Ectrodactyly and glaucoma associated with a 7q21.2-q31.2 interstitial deletion.

An infant with ectrodactyly, glaucoma, cleft palate, congenital heart defect and genital anomalies associated with a 7(q21.2q31.2) deletion is presented. Glaucoma and ectrodactyly in association with a 7q deletion has not been previously reported. We recommend that early ophthalmological assessment is required in infants with such deletions.

Fine mapping of the neurally expressed gene SOX14 to human 3q23, relative to three congenital diseases.

Members of the Sox gene family encode transcription factors that have diverse and important functions during development. We have recently described the cloning of chick and mouse Sox14 and the expression of these genes in a population of ventral interneurons in the embryonic spinal cord. We report here the cloning and sequencing of the human orthologue of Sox14. Human SOX14 shows remarkable sequence conservation compared with orthologues from other vertebrate species and probably mirrors the expression of these genes in the developing brain and spinal cord. Using radiation hybrid mapping and fluorescence in situ hybridisation, we have localised SOX14 close to the sequence tagged site D3S1576 on human chromosome 3q23. Three congenital disorders have been localised to this region: blepharophimosis-ptosis-epicanthus inversus syndrome (BPES), Charcot-Marie-Tooth neuropathy type IIB (CMT2B) and Mobius syndrome type 2 (MBS2). We have found that SOX14 is unlikely to be involved in any of these disorders because of the position of SOX14 proximal to a BPES breakpoint and the lack of SOX14 coding region alterations in BPES, CMT2B and MBS2 patients.

Investigation of a cryptic interstitial duplication involving the Prader-Willi/Angelman syndrome critical region.

A 3-year-old female referred with developmental delay, hypotonia and seizures was found to have a cryptic interstitial duplication of the Prader-Willi/Angelman critical region (PWACR). Her clinical features form part of a common phenotype characteristic of PWACR duplications including developmental delay, behavioural problems and speech difficulties. Microsatellite analysis showed that the duplication had arisen de novo, was maternal in origin and involved the entire 4-Mb PWACR between the common deletion breakpoints. The existence of cryptic rearrangements emphasises the need for molecular tests alongside conventional cytogenetics when investigating abnormalities involving this imprinted region.

A FISH approach to defining the extent and possible clinical significance of deletions at the WAGR locus.

Nineteen patients were analysed by fluorescence in situ hybridisation (FISH) with selected 11p13 markers. They were examined because they had either isolated sporadic or familial aniridia, or aniridia with one or more of the WAGR (Wilms' tumour, aniridia, genital anomalies, and mental retardation) syndrome anomalies. The FISH markers from distal 11p13 were cosmids FO2121, PAX6 (aniridia), D11S324, and WT1 (Wilms' tumour predisposition). Two of the patients with isolated aniridia were abnormal, one with an apparently balanced reciprocal 7;11 translocation and an 11p13 breakpoint, which by FISH was shown to be approximately 30 kb distal to the aniridia (PAX6) gene, and the other had a submicroscopic deletion involving part of PAX6 that extended distally for approximately 245 kb. Two patients with aniridia together with other WAGR malformations had deletions involving all four cosmids. One case with aniridia associated with developmental and growth delay had a deletion including FO2121 and PAX6 but not D11S324 and WT1, while in a further case the deletion included all four test cosmids. These studies show that a combined conventional and molecular cytogenetic approach to patients presenting with aniridia is a useful method for differentiating between those with deletions extending into and including WT1 and therefore between those with high and low risks of developing Wilms' tumour.

FISH studies in a patient with sporadic aniridia and t(7;11) (q31.2;p13).

A 2 year old female presenting with bilateral sporadic aniridia was found to have an apparently balanced reciprocal translocation with a chromosome 11 breakpoint within band p13. Fluorescence in situ hybridisation (FISH) studies with distal 11p13 specific cosmids showed that the chromosome 11 breakpoint lay between the aniridia (PAX6) locus and a region approximately 100 kb distal to PAX6 defined by the cosmid FO2121. Although this patient did not have a detectable deletion within PAX6, her aniridia may have resulted from a disruption of the distal chromatin domain containing either enhancers or regulators for PAX6. This case may therefore be another example of aniridia caused by a position effect as recently described in two familial aniridia patients in which the phenotype cosegregated with chromosome abnormalities with 11p13 breakpoints.

Crossover analysis in a British family suggests that Coffin-Lowry syndrome maps to a 3.4-cM interval in Xp22.

Coffin-Lowry syndrome (CLS; MIM 303600) in an uncommon X-linked disorder causing mental retardation and skeletal abnormalities. Most recently it was mapped to a 5.6-centimorgan (cM) region of Xp22, flanked distally by AFM291wf5 and proximally by DXS1052 [Biancalana et al., 1994: Genomics 22:617-625]. We present information which supports this localization and further narrows the region to approximately 3.4 cM. A recombination in a carrier from a British family mean that DXS365 is the closest proximal flanking marker identified to date for the region thought to contain the CLS gene. This information reduces the region of interest by approximately 2.2 cM, a significant decrease in terms of the scale of effort which will be required to isolate and analyze candidate genes.

Brachydactyly and mental retardation: an Albright hereditary osteodystrophy-like syndrome localized to 2q37.

We report five patients with a combination of brachymetaphalangia and mental retardation, similar to that observed in Albright hereditary osteodystrophy (AHO). Four patients had cytogenetically visible de novo deletions of chromosome 2q37. The fifth patient was cytogenetically normal and had normal bioactivity of the alpha subunit of Gs (Gs alpha), the protein that is defective in AHO. In this patient, we have used a combination of highly polymorphic molecular markers and FISH to demonstrate a microdeletion at 2q37. The common region of deletion overlap involves the most telomeric 2q marker, D2S125, and extends proximally for a maximum distance of 17.6 cM. We suggest this represents a consistent phenotype associated with some deletions at 2q37 and that genes important for skeletal and neurodevelopment lie within this region. Screening for deletions at this locus should be considered in individuals with brachymetaphalangia and mental retardation. Furthermore, 2q37 represents a candidate region for type E brachydactyly.

Nevoid basal cell carcinoma syndrome: review of 118 affected individuals.

One hundred eighteen cases of nevoid basal cell carcinoma syndrome (NBCCS, Gorlin's syndrome or basal cell nevus syndrome) are presented in this study. In aiming to ascertain all the affected families in Australia, we have examined the largest series to date. Relative frequencies of associated complications are presented and compared with those of the recent English survey by Evans et al. [J Med Genet 30:460-464, 1993]. The frequencies of most manifestations are similar. However, one major difference is that the multiple basal cell carcinomas are manifest from an earlier age in the Australian population, which probably reflects greater exposure to ultraviolet radiation. Of the 64 families ascertained, 37 represented simplex cases, and, accordingly, the apparent new mutation rate is surprisingly high (14-81%) given the lack of impact of NBCCS on reproductive capabilities. There is some evidence to suggest that this may be attributable to anticipation.

Clinical identification of a human equivalent to the short ear (se) murine phenotype.

Mutations in the BMP-5 gene at the mouse short-ear locus alter size, shape, and number of many different skeletal elements, and greatly reduce the size of the external ear. The alterations in short-ear mice are confined to particular skeletal features and a human equivalent is not known. We report on 5 patients whose features fit into the clinical criteria of the EPS (ear, patella, short stature) syndrome characterized by very short external ears, small jaw, growth retardation, and different skeletal abnormalities including absent patellae. We postulate on clinical data and phenotype comparisons that some EPS cases might be a human equivalent to the short ear murine disorder.

Femoral hypoplasia unusual facies syndrome with preaxial polydactyly.

Preaxial polydactyly of the foot is an unusual feature in femoral hypoplasia unusual facies syndrome, having been recorded with certainty in only two previous reports. We now add a further two instances of this rare association and emphasize that this finding should not preclude the underlying syndromic diagnosis.

Further localization of the gene for nevoid basal cell carcinoma syndrome (NBCCS) in 15 Australasian families: linkage and loss of heterozygosity.

Nevoid basal cell carcinoma syndrome (NBCCS; basal cell nevus syndrome or Gorlin syndrome) is a cancer-predisposition syndrome characterized by multiple basal cell carcinomas (BCCs) and diverse developmental defects. The gene for NBCCS has been mapped to 9q23.1-q31 in North American and European families. In addition, loss of heterozygosity (LOH) for genetic markers in this region has been detected in sporadic BCCs, indicating that the NBCCS gene is probably a tumor-suppressor gene. In this study we have determined that the NBCCS gene is also linked to this region in Australasian pedigrees and that there is no significant evidence of heterogeneity. We have defined the localization of the gene by multipoint and haplotype analysis of 15 families, using four microsatellite markers. LOH at these loci was detected in 50% of sporadic BCCs, a rate that is significantly higher than that in other skin lesions used as controls.

Evidence for locus heterogeneity in autosomal dominant torsion dystonia.

Idiopathic torsion dystonia (ITD) is most commonly an autosomal dominant disorder with reduced penetrance and variable expression. A locus on the distal long arm of chromosome 9 has been identified in one large non-Jewish and several Jewish families in the United States. Linkage analysis in a large Australian kindred with ITD, also containing two patients with Wilson's disease, excludes a locus for ITD in chromosome 9q34 or the region of chromosome 13 containing the Wilson disease gene. This study provides evidence for locus heterogeneity in autosomal dominant ITD and also gives additional information on gene order in chromosome 9q.

A YAC contig in Xp21 containing the adrenal hypoplasia congenita and glycerol kinase deficiency genes.

The gene loci for adrenal hypoplasia congenita (AHC) and glycerol kinase deficiency (GK) map in Xp21 distal to Duchenne muscular dystrophy (DMD), and proximal to DXS28 (C7), by analysis of patient deletions. We have constructed a yeast artificial chromosome (YAC) contig encompassing a 1.2 Mb region extending distally from DMD, and containing DXS708 (JC-1), the distal junction clone of a patient with GK and DMD. A pulsed-field gel electrophoresis map of the YAC contig identified 3 potential CpG islands. Whole YAC hybridization identified cosmids both for construction of cosmid contigs, and isolation of single copy probes. Thirteen new single copy probes and DXS28 and DXS708 were hybridized on a panel of patients; the deletion mapping indicates that the YAC contig contains both GK and at least part of AHC, and together with the physical map defines a GK critical region of 50-250 kb. In one AHC patient with a cytogenetically detectable deletion we used the new probes to characterize a complex double deletion. Non-overlapping deletions observed in other unrelated AHC patients indicate that the AHC gene is large, extending over at least 200-500 kb. This mapping provides the basis for the identification of the AHC and GK genes.