Homologous nonallelic recombinations between the iduronate-sulfatase gene and pseudogene cause various intragenic deletions and inversions in patients with mucopolysaccharidosis type II.

About 20% of patients with mucopolysaccharidosis type II (MPS II) have gross structural rearrangements involving the iduronate-sulfatase (IDS) gene in Xq27.3-q28. A nearby IDS pseudogene (IDS-2) promotes nonallelic recombination between highly homologous sequences. Here we describe major rearrangements due to gene/pseudogene recombination. In two unrelated patients, partial IDS gene deletions were found joining introns 3 and 7 of the IDS gene together with gene to pseudogene conversion in the area of breakpoints. In a third patient, a junction between intron 3 of IDS-2 and intron 7 of IDS was seen that was due to a deletion and inversion of the 5' part of the gene. Characterisation of breakpoints in six patients with large inversions revealed that all recombinations of this type occurred in the same area of homology between IDS and IDS-2; they were molecularly balanced, and accompanied by gene conversions in most cases. Apart from diagnostic implications, such naturally occurring recombination 'hot spots' may allow some insight into general features of crossover events in mammals.

Mutations in RPE65 cause autosomal recessive childhood-onset severe retinal dystrophy.

Autosomal recessive childhood-onset severe retinal dystrophy (arCSRD) designates a heterogeneous group of disorders affecting rod and cone photoreceptors simultaneously. The most severe cases are termed Leber congenital amaurosis (LCA), while the less aggressive forms are usually considered juvenile retinitis pigmentosa. Recently, mutations in the retinal-specific guanylate cyclase gene were found in patients with LCA. Disease genes implicated in other forms of arCSRD are expected to encode proteins present in the neuroretina or in the retinal pigment epithelium (RPE). The RPE, a monolayer of cells separating the vascular-rich choroid and the neuroretina, is in intimate contact with the outer segments of rods and cones via the microvilli surrounding the photoreceptors. The RPE expresses a tissue-specific and evolutionarily highly conserved 61 kD protein (RPE65) present at high levels in vivo. Although the function of RPE65 is not yet known, an important role in the RPE/photoreceptor vitamin-A cycle is suggested by the fact that RPE65 associates both with serum retinol-binding protein and with the RPE-specific 11-cis retinol dehydrogenase, an enzyme active in the synthesis of the visual pigment chromophore 11-cis retinal. Here we report that the analysis of RPE65 in a collection of about 100 unselected retinal-dystrophy patients of different ethnic origin revealed five that are likely to be pathogenic mutations, including a missense mutation (Pro363Thr), two point mutations affecting splicing (912 + 1G-->T and 65 + 5G-->A) and two small re-arrangements (ins144T and 831del8) on a total of nine alleles of five patients with arCSRD. In contrast to other genes whose defects have been implicated in degenerative retinopathies, RPE65 is the first disease gene in this group of inherited disorders that is expressed exclusively in the RPE, and may play a role in vitamin-A metabolism of the retina.

Mucopolysaccharidosis type II (Hunter syndrome): mutation "hot spots" in the iduronate-2-sulfatase gene.

Mucopolysaccharidosis type II (MPS II, Hunter syndrome) is an X-chromosomal storage disorder due to deficiency of the lysosomal enzyme iduronate-2-sulfatase (IDS). We have identified IDS mutations in a total of 31 families/patients with MPS II, of which 20 are novel and unique and a further 1 is novel but has been found in 3 unrelated patients. One of the mutations detected is of special interest as an AG-->G substitution in an intron, far apart from the coding region, is deleterious by creating a new 5'-splice-donor site that results in the inclusion of a 78-bp intronic sequence. While the distribution of gene rearrangements (deletions, insertions, and duplications) of <20 bp seems to be random over the IDS gene, the analysis of a total of 101 point mutations lying within the coding region shows that they tend to be more frequent in exons III, VIII, and IX. Forty-seven percent of the point mutations are at CpG dinucleotides, of which G:C-to-A:T transitions constitute nearly 80%. Almost all recurrent point mutations involve CpG sites. Analysis of a collective of 50 families studied in our laboratory, to date, revealed that mutations occur more frequently in male meioses (estimated male-to-female ratio between 3.76 and 6.3).

[Microwave volumetry of the extremities: a technical measuring alternative to the noninvasive venous occlusion test]. / Mikrowellen-Volumetrie der Extremitäten: eine messtechnische Alternative für den rückwirkungs-freien Venenverschlusstest.

Venous occlusion plethysmography is a well-established noninvasive procedure for the investigation of venous outflow disorders. Various different sensor systems are available for the purpose, e.g. photoplethysmography, strain-gauge plethysmography and impedance plethysmography. The technique of microwave volumetry described in this paper is based on the interaction of the electromagnetic fields within the loss medium, tissue, and uses an air-filled resonant cavity as a sensor. As in the case of a microwave oven, this is excited by a high-frequency generator, but involves the use of far less power. As the volume of the measured objects (foot, arm) increases, the losses in the resonator also increase and can be analysed in a computer. The results can be represented in the form of a classical venous occlusion curve.

Evidence for an iduronate-sulfatase pseudogene near the functional Hunter syndrome gene in Xq27.3-q28.

We are currently characterizing mutations of the iduronate-2-sulfatase (IDS) gene in patients with Hunter syndrome (mucopolysaccharidosis type II). Surprisingly, all 17 patients with a mutation in exon III of the IDS gene identified by us were found to carry both the mutant and wild-type sequences in polymerase chain reaction (PCR) products amplified from genomic DNA. Similarly, two unaffected male controls showed a heterozygous pattern for two different point mutations in exon III. Collectively, the data suggest that at least intron 2, exon III, and the 3'-half of exon II of the functional IDS gene are present in the human genome as (part of) a non-expressed IDS gene. Deletion mapping further suggests that the pseudogene is in distal Xq in physical proximity to the functional IDS gene. The high degree of sequence homology observed between the functional IDS gene and pseudogene results in permanent co-amplification in PCR-based screening methods and makes mutation analysis at the genomic DNA level difficult.