Kirsten Hoffmann

Mutations in the X-Linked Cyclin-Dependent Kinase–Like 5 (CDKL5/STK9) Gene Are Associated with Severe Neurodevelopmental Retardation

Recently, we showed that truncation of the X-linked cyclin-dependent kinase-like 5 (CDKL5/STK9) gene caused mental retardation and severe neurological symptoms in two female patients. Here, we report that de novo missense mutations in CDKL5 are associated with a severe phenotype of early-onset infantile spasms and clinical features that overlap those of other neurodevelopmental disorders, such as Rett syndrome and Angelman syndrome. The mutations are located within the protein kinase domain and affect highly conserved amino acids; this strongly suggests that impaired CDKL5 catalytic activity plays an important role in the pathogenesis of this neurodevelopmental disorder. In view of the overlapping phenotypic spectrum of CDKL5 and MECP2 mutations, it is tempting to speculate that these two genes play a role in a common pathogenic process.

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.

A balanced chromosomal translocation disrupting ARHGEF9 is associated with epilepsy, anxiety, aggression, and mental retardation

Clustering of inhibitory γ‐aminobutyric acidA (GABAA) and glycine receptors at synapses is thought to involve key interactions between the receptors, a “scaffolding” protein known as gephyrin and the RhoGEF collybistin. We report the identification of a balanced chromosomal translocation in a female patient presenting with a disturbed sleep‐wake cycle, late‐onset epileptic seizures, increased anxiety, aggressive behavior, and mental retardation, but not hyperekplexia. Fine mapping of the breakpoint indicates disruption of the collybistin gene (ARHGEF9) on chromosome Xq11, while the other breakpoint lies in a region of 18q11 that lacks any known or predicted genes. We show that defective collybistin transcripts are synthesized and exons 7–10 are replaced by cryptic exons from chromosomes X and 18. These mRNAs no longer encode the pleckstrin homology (PH) domain of collybistin, which we now show binds phosphatidylinositol‐3‐phosphate (PI3P/PtdIns‐3‐P), a phosphoinositide with an emerging role in membrane trafficking and signal transduction, rather than phosphatidylinositol 3,4,5‐trisphosphate (PIP3/PtdIns‐3,4,5‐P) as previously suggested in the “membrane activation model” of gephyrin clustering. Consistent with this finding, expression of truncated collybistin proteins in cultured neurons interferes with synaptic localization of endogenous gephyrin and GABAA receptors. These results suggest that collybistin has a key role in membrane trafficking of gephyrin and selected GABAA receptor subtypes involved in epilepsy, anxiety, aggression, insomnia, and learning and memory. Hum Mutat 0,1–9, 2008.

Mutations in the ZNF41 gene are associated with cognitive deficits: identification of a new candidate for X-linked mental retardation

Nonsyndromic X-linked mental retardation (MRX) is defined by an X-linked inheritance pattern of low IQ, problems with adaptive behavior, and the absence of additional specific clinical features. The 13 MRX genes identified to date account for less than one-fifth of all MRX, suggesting that numerous gene defects cause the disorder in other families. In a female patient with severe nonsyndromic mental retardation and a de novo balanced translocation t(X;7)(p11.3;q11.21), we have cloned the DNA fragment that contains the X-chromosomal and the autosomal breakpoint. In silico sequence analysis provided no indication of a causative role for the chromosome 7 breakpoint in mental retardation (MR), whereas, on the X chromosome, a zinc-finger gene, ZNF41, was found to be disrupted. Expression studies indicated that ZNF41 transcripts are absent in the patient cell line, suggesting that the mental disorder in this patient results from loss of functional ZNF41. Moreover, screening of a panel of patients with MRX led to the identification of two other ZNF41 mutations that were not found in healthy control individuals. A proline-to-leucine amino acid exchange is present in affected members of one family with MRX. A second family carries an intronic splice-site mutation that results in loss of specific ZNF41 splice variants. Wild-type ZNF41 contains a highly conserved transcriptional repressor domain that is linked to mechanisms of chromatin remodeling, a process that is defective in various other forms of MR. Our results suggest that ZNF41 is critical for cognitive development; further studies aim to elucidate the specific mechanisms by which ZNF41 alterations lead to MR.

Disruption of Netrin G1 by a balanced chromosome translocation in a girl with Rett syndrome

We have identified a girl with characteristic features of Rett syndrome (RTT) who carries a de novo balanced translocation involving chromosomes 1 and 7. Both breakpoints were mapped by fluorescence in situ hybridization with selected genomic clones from the regions of interest. Southern blot hybridisations, utilizing probes derived from breakpoint spanning BACs, detected several aberrant fragments specific for the patient. Sequence analysis of the cloned junction fragment indicated that on chromosome 1 the predominantly brain-expressed Netrin G1 (NTNG1) gene is disrupted, whereas on chromosome 7 there was no indication for a truncated gene. The chromosome 1 breakpoint lies within the 3′ part of NTNG1 and affects alternatively spliced transcripts, suggesting that the phenotype in this patient is the result of disturbed NTNG1 expression. In silico translation of the NTNG1 splice variants predicted protein isoforms with different C-termini: one membrane bound through a glycosylphosphatidylinositol anchor and the other soluble. The membrane-bound protein isoform would be affected by the breakpoint, whereas the soluble form would remain intact. Our results suggest that the central nervous system is sensitive to NTNG1 expression levels and that NTNG1 is a novel candidate disease gene for RTT.

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.

Heterotaxy and cardiac defect in a girl with chromosome translocation t(X;1)(q26;p13.1) and involvement of ZIC3.

We report on a 2-year-old girl with situs ambiguus comprising right-sided stomach and spleen, left-sided liver and complex cardiac defect. Psychomotor development of this patient was normal, and no other major abnormalities were present. Chromosome analysis revealed a de novo balanced chromosome translocation t(X;1)(q26;p13.1). Molecular cytogenetic investigations identified a breakpoint spanning BAC clone on the X-chromosome containing the ZIC3 gene. Mutations in ZIC3 are associated with situs ambiguus and cardiac defects predominantly in males. This is the first report of a live born girl with an X-autosome translocation involving the ZIC3 region.

Molecular characterization of a balanced chromosome translocation in psoriasis vulgaris

To the Editor: Psoriasis vulgaris is a chronic inflammatory skin disease which affects approximately 2–5% of the Caucasian population (1). The etiology of this disorder is largely unknown (2). Familial clustering (3) and a high concordance rate in monozygotic compared to dizygotic twins (4) indicates a strong contribution of genetic susceptibility factors. Genome-wide linkage analyzes have identified 19 susceptibility loci to date (5). The PSORS1 locus on 6p21.3 has been consistently reproduced in several independent studies (6–11). Linkage to 4q31 (PSORS9) was found in a large cohort of Chinese Han families (10). A meta-analysis combining the results of six genome-wide scans and a recent study in the Icelandic population confirmed significant linkage to PSORS9 on 4q31 (5, 11). However, as is the case for other complex disorders, positional cloning of psoriasis disease genes faces substantial difficulties due to genetic heterogeneity, the multitude of genetic risk factors, and the fact that there are different risk factors in different populations. No psoriasis susceptibility gene in 4q31 has been identified to date. Breakpoint analyzes in patients with balanced chromosome rearrangements have led to the identification of numerous genes involved in monogenic disorders (12) and facilitated the detection of candidate genes for complex and late-onset diseases, e.g., schizophrenia (13) and developmental dyslexia (14). Here, we describe a 43-year-old male patient with a balanced translocation 46,XY,t(2;4) (p25;q31.1) (Fig. 1a) and familial psoriasis vulgaris. The proband has suffered from psoriasis vulgaris since the age of 20. The father of our proband, who died of a heart attack at the age of 62, also suffered from psoriasis, and, unlike his son, he had additional arthritic problems. No chromosome analysis was performed; however, we speculate that he was also a carrier of this translocation because two female partners had multiple spontaneous abortions. No other relatives were available for chromosome analysis, and no other family members were known to have suffered from psoriasis. Molecular cytogenetic breakpoint analysis by FISH experiments (15) using YAC and BAC clones listed in Table 1 revealed breakpoint-spanning BAC clones on chromosome 2 (RP11-796O1, which contains no genes) and on chromosome 4 [RP11-1079H4 (Fig. 1b) and RP11-90M18]. The presence of several genes in this region – SET7, MGST2 and MAML3 – prompted us to determine the breakpoint at the molecular level. For Southern blot analysis, patient and control genomic DNA samples were digested with several restriction enzymes and hybridized with a[P]dCTP-labeled DNA probes derived from BAC RP11-1079H4. Hybridization with probe PSORPZ (amplified with the following primers: 50-AGAGTGAGGCCTTCTCAGCA-30 and 50-GCCATGTTTCCAGGTTACCA-30) revealed aberrant fragments exclusively in patient DNA digested with restriction enzymes SspI, MboI, and PstI (Fig. 1c). Polymerase chain reaction (PCR) on adaptorligated genomic DNA was performed as described previously (16) for breakpoint cloning. Patient genomic DNA was digested with MboI. Amplification was performed using adaptorspecific primers and chromosome 4-specific nested primers. Sequencing of the cloned PCR product revealed a fusion fragment that aligned to chromosomes 4 and 2 (Fig. 1d), which indicated the breakpoint location between position chr4:140,938,199 and chr4:140,938,200 on the UCSC Genome Browser May 2004 assembly. The breakpoint lies within intron 3 of EST CR742434, 6581 bp upstream of the start codon of MGST2 (Fig. 2). Both semiquantitative RT-PCR and Northern blot investigations of MGST2 failed to detect Clin Genet 2006: 69: 189–193 Copyright # Blackwell Munksgaard 2005 Printed in Singapore. All rights reserved CLINICALGENETICS doi: 10.1111/j.1399-0004.2005.00558.x