Steffen Lenzner

Positional cloning of the gene for X-linked retinitis pigmentosa 2

X-linked retinitis pigmentosa (XLRP) results from mutations in at least two different loci, designated RP2 and RP3, located at Xp11.3 and Xp21.1, respectively. The RP3 gene was recently isolated by positional cloning, whereas the RP2 locus was mapped genetically to a 5-cM interval. We have screened this region for genomic rearrangements by the YAC representation hybridization (YRH) technique and detected a LINE1 (L1) insertion in one XLRP patient. The L1 retrotransposition occurred in an intron of a novel gene that consisted of five exons and encoded a polypeptide of 350 amino acids. Subsequently, nonsense, missense and frameshift mutations, as well as two small deletions, were identified in six additional patients. The predicted gene product shows homology with human cofactor C, a protein involved in the ultimate step of ß-tubulin folding. Our data provide evidence that mutations in this gene, designated RP2, are responsible for progressive retinal degeneration.

Mutations in PHF8 are associated with X linked mental retardation and cleft lip/cleft palate

Truncating mutations were found in the PHF8 gene (encoding the PHD finger protein 8) in two unrelated families with X linked mental retardation (XLMR) associated with cleft lip/palate (MIM 300263). Expression studies showed that this gene is ubiquitously transcribed, with strong expression of the mouse orthologue Phf8 in embryonic and adult brain structures. The coded PHF8 protein harbours two functional domains, a PHD finger and a JmjC (Jumonji-like C terminus) domain, implicating it in transcriptional regulation and chromatin remodelling. The association of XLMR and cleft lip/palate in these patients with mutations in PHF8 suggests an important function of PHF8 in midline formation and in the development of cognitive abilities, and links this gene to XLMR associated with cleft lip/palate. Further studies will explore the specific mechanisms whereby PHF8 alterations lead to mental retardation and midline defects.

A novel X-linked recessive mental retardation syndrome comprising macrocephaly and ciliary dysfunction is allelic to oral–facial–digital type I syndrome

We report on a large family in which a novel X-linked recessive mental retardation (XLMR) syndrome comprising macrocephaly and ciliary dysfunction co-segregates with a frameshift mutation in the OFD1 gene. Mutations of OFD1 have been associated with oral–facial–digital type 1 syndrome (OFD1S) that is characterized by X-chromosomal dominant inheritance and lethality in males. In contrast, the carrier females of our family were clinically inconspicuous, and the affected males suffered from severe mental retardation, recurrent respiratory tract infections and macrocephaly. All but one of the affected males died from respiratory problems in infancy; and impaired ciliary motility was confirmed in the index patient by high-speed video microscopy examination of nasal epithelium. This family broadens the phenotypic spectrum of OFD1 mutations in an unexpected way and sheds light on the complexity of the underlying disease mechanisms.

Mutations in the polyglutamine binding protein 1 gene cause X-linked mental retardation.

We found mutations in the gene PQBP1 in 5 of 29 families with nonsyndromic (MRX) and syndromic (MRXS) forms of X-linked mental retardation (XLMR). Clinical features in affected males include mental retardation, microcephaly, short stature, spastic paraplegia and midline defects. PQBP1 has previously been implicated in the pathogenesis of polyglutamine expansion diseases. Our findings link this gene to XLMR and shed more light on the pathogenesis of this common disorder.

: CGHPRO – a comprehensive data analysis tool for array CGH

BackgroundArray CGH (Comparative Genomic Hybridisation) is a molecular cytogenetic technique for the genome wide detection of chromosomal imbalances. It is based on the co-hybridisation of differentially labelled test and reference DNA onto arrays of genomic BAC clones, cDNAs or oligonucleotides, and after correction for various intervening variables, loss or gain in the test DNA can be indicated from spots showing aberrant signal intensity ratios.Now that this technique is no longer confined to highly specialized laboratories and is entering the realm of clinical application, there is a need for a user-friendly software package that facilitates estimates of DNA dosage from raw signal intensities obtained by array CGH experiments, and which does not depend on a sophisticated computational environment.ResultsWe have developed a user-friendly and versatile tool for the normalization, visualization, breakpoint detection and comparative analysis of array-CGH data. CGHPRO is a stand-alone JAVA application that guides the user through the whole process of data analysis. The import option for image analysis data covers several data formats, but users can also customize their own data formats. Several graphical representation tools assist in the selection of the appropriate normalization method. Intensity ratios of each clone can be plotted in a size-dependent manner along the chromosome ideograms. The interactive graphical interface offers the chance to explore the characteristics of each clone, such as the involvement of the clones sequence in segmental duplications. Circular Binary Segmentation and unsupervised Hidden Markov Model algorithms facilitate objective detection of chromosomal breakpoints. The storage of all essential data in a back-end database allows the simultaneously comparative analysis of different cases. The various display options facilitate also the definition of shortest regions of overlap and simplify the identification of odd clones.ConclusionCGHPRO is a comprehensive and easy-to-use data analysis tool for array CGH. Since all of its features are available offline, CGHPRO may be especially suitable in situations where protection of sensitive patient data is an issue. It is distributed under GNU GPL licence and runs on Linux and Windows.

Homozygosity mapping in consanguineous families reveals extreme heterogeneity of non-syndromic autosomal recessive mental retardation and identifies 8 novel gene loci

Autosomal recessive gene defects are arguably the most important, but least studied genetic causes of severe cognitive dysfunction. Homozygosity mapping in 78 consanguineous Iranian families with nonsyndromic autosomal recessive mental retardation (NS-ARMR) has enabled us to determine the chromosomal localization of at least 8 novel gene loci for this condition. Our data suggest that in the Iranian population NS-ARMR is very heterogeneous, and they argue against the existence of frequent gene defects that account for more than a few percent of the cases.

Mutations in the FTSJ1 gene coding for a novel S-adenosylmethionine-binding protein cause nonsyndromic X-linked mental retardation

Nonsyndromic X-linked mental retardation (NSXLMR) is a very heterogeneous condition, and most of the underlying gene defects are still unknown. Recently, we have shown that approximately 30% of these genes cluster on the proximal Xp, which prompted us to perform systematic mutation screening in brain-expressed genes from this region. Here, we report on a novel NSXLMR gene, FTSJ1, which harbors mutations in three unrelated families--one with a splicing defect, one with a nonsense mutation, and one with a deletion of one nucleotide. In two families, subsequent expression studies showed complete absence or significant reduction of mutant FTSJ1 transcripts. FTSJ1 protein is a homolog of Escherichia coli RNA methyltransferase FtsJ/RrmJ and may play a role in the regulation of translation. Further studies aim to elucidate the function of human FTSJ1 and its role during brain development.

MCT8 mutation analysis and identification of the first female with Allan-Herndon-Dudley syndrome due to loss of MCT8 expression

Mutations in the thyroid monocarboxylate transporter 8 gene (MCT8/SLC16A2) have been reported to result in X-linked mental retardation (XLMR) in patients with clinical features of the Allan–Herndon–Dudley syndrome (AHDS). We performed MCT8 mutation analysis including 13 XLMR families with LOD scores >2.0, 401 male MR sibships and 47 sporadic male patients with AHDS-like clinical features. One nonsense mutation (c.629insA) and two missense changes (c.1A>T and c.1673G>A) were identified. Consistent with previous reports on MCT8 missense changes, the patient with c.1673G>A showed elevated serum T3 level. The c.1A>T change in another patient affects a putative translation start codon, but the same change was present in his healthy brother. In addition normal serum T3 levels were present, suggesting that the c.1A>T (NM_006517) variation is not responsible for the MR phenotype but indicates that MCT8 translation likely starts with a methionine at position p.75. Moreover, we characterized a de novo translocation t(X;9)(q13.2;p24) in a female patient with full blown AHDS clinical features including elevated serum T3 levels. The MCT8 gene was disrupted at the X-breakpoint. A complete loss of MCT8 expression was observed in a fibroblast cell-line derived from this patient because of unfavorable nonrandom X-inactivation. Taken together, these data indicate that MCT8 mutations are not common in non-AHDS MR patients yet they support that elevated serum T3 levels can be indicative for AHDS and that AHDS clinical features can be present in female MCT8 mutation carriers whenever there is unfavorable nonrandom X-inactivation.

CCDC22 deficiency in humans blunts activation of proinflammatory NF-kappa B signaling

NF-κB is a master regulator of inflammation and has been implicated in the pathogenesis of immune disorders and cancer. Its regulation involves a variety of steps, including the controlled degradation of inhibitory IκB proteins. In addition, the inactivation of DNA-bound NF-κB is essential for its regulation. This step requires a factor known as copper metabolism Murr1 domain-containing 1 (COMMD1), the prototype member of a conserved gene family. While COMMD proteins have been linked to the ubiquitination pathway, little else is known about other family members. Here we demonstrate that all COMMD proteins bind to CCDC22, a factor recently implicated in X-linked intellectual disability (XLID). We showed that an XLID-associated CCDC22 mutation decreased CCDC22 protein expression and impaired its binding to COMMD proteins. Moreover, some affected individuals displayed ectodermal dysplasia, a congenital condition that can result from developmental NF-κB blockade. Indeed, patient-derived cells demonstrated impaired NF-κB activation due to decreased IκB ubiquitination and degradation. In addition, we found that COMMD8 acted in conjunction with CCDC22 to direct the degradation of IκB proteins. Taken together, our results indicate that CCDC22 participates in NF-κB activation and that its deficiency leads to decreased IκB turnover in humans, highlighting an important regulatory component of this pathway.

Duplication of the MID1 first exon in a patient with Opitz G/BBB syndrome.

Opitz G/BBB syndrome is a malformation syndrome of the ventral midline mainly characterized by hypertelorism, swallowing difficulties, hypospadias and developmental delay. SSCP analysis and genomic sequencing of the MID1 open reading frame have identified mutations in 80% of the families with X-linked inheritance. However, in many patients the underlying genetic defect remains undetected by these techniques. Using RNA diagnostics we have now identified a duplication of the MID1 first exon in a patient with X-linked Opitz G/BBB syndrome. This duplication introduces a premature termination codon. In addition, we could significantly lower the threshold for mutation detection on the DNA level by combining SSCP analysis with DHPLC technology.