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Purpose: To identify the mutation for Volkmann cataract (CTRCT8) at 1p36.33. Methods: The genes in the candidate region 1p36.33 were Sanger and parallel deep sequenced, and informative single nucleotide polymorphisms (SNP') were identified for linkage analysis. Expression analysis with reverse transcription polymerase chain reaction (RT-PCR) of the candidate gene was performed using RNA from different human tissues. Quantitative transcription polymerase chain reaction (qRT-PCR) analysis of the GNB1 gene was performed in affected and healthy individuals. Bioinformatic analysis of the linkage regions including the candidate gene was performed. Results: Linkage analysis of the 1p36.33 CCV locus applying new marker systems obtained with Sanger and deep sequencing reduced the candidate locus from 2.1 Mb to 0.389 Mb flanked by the markers STS-22AC and rs549772338 and resulted in an logarithm of the odds (LOD) score of Z = 21.67. The identified mutation, rs763295804, affects the donor splice site in the long non-coding RNA gene RP1-140A9.1 (ENSG00000231050). The gene including splice-site junctions is conserved in primates but not in other mammalian genomes, and two alternative transcripts were shown with RT-PCR. One of these transcripts represented a lens cell-specific transcript. Meta-analysis of the Cross-LinkingImmuno-Precipitation sequencing (CLIP-Seq) data suggested the RNA binding protein (RBP) eIF4AIII is an active counterpart for RP1-140A9.1, and several miRNA and transcription factors binding sites were predicted in the proximity of the mutation. ENCODE DNase I hypersensitivity and histone methylation and acetylation data suggest the genomic region may have regulatory functions. Conclusions: The mutation in RP1-140A9.1 suggests the long non-coding RNA as the candidate cataract gene associated with the autosomal dominant inherited congenital cataract from CCV. The mutation has the potential to destroy exon/ intron splicing of both transcripts of RP1-140A9.1. Sanger and massive deep resequencing of the linkage region failed to identify alternative candidates suggesting the mutation in RP1-140A9.1 is causative for the CCV phenotype.

OBJECTIVE: SCN8A encodes the sodium channel voltage-gated α8-subunit (Nav1.6). SCN8A mutations have recently been associated with epilepsy and neurodevelopmental disorders. We aimed to delineate the phenotype associated with SCN8A mutations.METHODS: We used high-throughput sequence analysis of the SCN8A gene in 683 patients with a range of epileptic encephalopathies. In addition, we ascertained cases with SCN8A mutations from other centers. A detailed clinical history was obtained together with a review of EEG and imaging data.RESULTS: Seventeen patients with de novo heterozygous mutations of SCN8A were studied. Seizure onset occurred at a mean age of 5 months (range: 1 day to 18 months); in general, seizures were not triggered by fever. Fifteen of 17 patients had multiple seizure types including focal, tonic, clonic, myoclonic and absence seizures, and epileptic spasms; seizures were refractory to antiepileptic therapy. Development was normal in 12 patients and slowed after seizure onset, often with regression; 5 patients had delayed development from birth. All patients developed intellectual disability, ranging from mild to severe. Motor manifestations were prominent including hypotonia, dystonia, hyperreflexia, and ataxia. EEG findings comprised moderate to severe background slowing with focal or multifocal epileptiform discharges.CONCLUSION: SCN8A encephalopathy presents in infancy with multiple seizure types including focal seizures and spasms in some cases. Outcome is often poor and includes hypotonia and movement disorders. The majority of mutations arise de novo, although we observed a single case of somatic mosaicism in an unaffected parent.

OBJECTIVE: Benign familial infantile seizures (BFIS), paroxysmal kinesigenic dyskinesia (PKD), and their combination - known as infantile convulsions and paroxysmal choreoathetosis (ICCA) - are related autosomal dominant diseases. PRRT2 (proline-rich transmembrane protein 2 gene) has been identified as the major gene in all three conditions, found to be mutated in 80-90% of familial and 30-35% of sporadic cases.METHODS: We searched for the genetic defect in PRRT2-negative, unrelated families with BFIS or ICCA using whole exome or targeted gene panel sequencing, and performed a detailed clinico-neurophysiological workup.RESULTS: In three families with a total of 16 affected members, we identified the same, co-segregating heterozygous missense mutation (c.4447G>A; p.E1483K) in SCN8A, encoding a voltage-gated sodium channel. A founder effect was excluded by linkage analysis. All individuals but one had normal cognitive and motor milestones, neuroimaging and interictal neurological status. Fifteen affected members presented with afebrile focal or generalized tonic-clonic seizures during the first-second year of life; 5 of them experienced single unprovoked seizures later on. One patient had seizures only at school age. All patients stayed otherwise seizure-free, most without medication. Interictal EEG was normal in all cases but two. Five/16 patients developed additional brief paroxysmal episodes in puberty, either dystonic/dyskinetic or "shivering" attacks, triggered by stretching, motor initiation or by emotional stimuli. In one case, we recorded typical PKD spells by video-EEG-polygraphy, documenting a cortical involvement.INTERPRETATION: Our study establishes SCN8A as a novel gene in which a recurrent mutation causes BFIS/ICCA, expanding the clinical-genetic spectrum of combined epileptic and dyskinetic syndromes. This article is protected by copyright. All rights reserved.

Primers used in ChIP-QPCR and mRNA expression analyses. This file contains primer sequences used for ChIP-QPCR verification of ChIPseq peaks located in the promoter regions of five selected genes. (PDF 21 kb)

Integrative analysis of ChIP-seq and gene-expression data. In order to investigate the gene-regulatory potential of BRD1-S and BRD1-L, ChIPseq data were integrated with expression data. A window of +/– 200 kb from BRD1 binding was constructed, fragmented in 9 kb segments, and determined the number of DEGs having TSSs in these segments. As opposed to the 10 kb segment used in the main manuscript the 9 kb segments used here span across 0 on the x-axis. A, B The analysis was performed for DEGs identified after upregulating BRD1-S or BRD1-L or (C, D) downregulating endogenous BRD1. Significantly more up- or downregulated genes were identified in segments where BRD1-S or BRD1-L binds in close proximity to TSSs compared to all segments across the window (P

Spatiotemporal co-expression landscapes of BRD1-S and BRD1-L PPIs and PTGs in the human brain. The fraction of co-expressed genes with BRD1 across 32 spatiotemporal intervals. A The fraction of genes co-expressed with BRD1 across all genes (gray) and genes in the BRD1-S and BRD1-L PPI networks (red and orange). None of these were significant at P

Chromatin binding profiles for BRD1-S and BRD1-L in relation to identified epigenetic marks in HEK293 cell lines. ChIP-seq data (fastq files), from HEK293 cell lines and cell line derivatives, were obtained from the ENCODE project and mapped to hg19 (for further details see Methods). An in-house histone H3K9ac ChIP-seq dataset was also included in the analysis. The minimum distance from (A) BRD1-S or (B) BRD1-L ChIP-seq peak to transcription factor binding site or histone mark was identified for all 2205 and 1722 peaks, respectively. The results were summarized as histograms with a +/â 30Â kb window from BRD1 binding (distance from BRD1 peaks). The number of peaks within each of the summed bins (columns) can be read on the y-axis (number of peaks). Random ChIP-seq peak regions (Random), with the same composition of chromosomes and peak region sizes, were generated to evaluate if the results could be explained be chance. (PDF 635 kb)

Verification of five selected ChIP-seq promoter targets by ChIP-QPCR. From the 251 common promotor target genes (PTGs) of BRD1-S and BRD1-L, we selected five for validation with ChIP-QPCR. A The promoter loci at the WD repeat domain 7 (WDR7), digestive organ expansion factor homolog (C1ORF107 or DIEXF), proteasome subunit β type, 2 (PSMB2), ZC3H15, and zinc finger protein 226 (ZNF226) showed BRD1-S and BRD1-L binding, while the promoter loci of the lymphocyte expressed gene granzyme M (GZMM) showed no significant binding of the BRD1 isoforms and was subsequently selected as a negative control. The BRD1-S or BRD1-L binding is shown in black whereas the control ChIP-seq (IP with anti-HA antibody) is shown in pink. B Primers were designed to amplify a 80–150 bp sequence in the promoter region or the intron of all six genes (for primer sequences see Additional file 7). ChIP was performed with either anti-V5 antibody (V5) conjugated beads or anti-HA (HA) conjugated beads using extracts from BRD1-S, BRD1-L, or HEK293T cells (for further details see “Methods”). ChIP-QPCR showed higher enrichment of BRD1-S and BRD1-L at the promoter loci of WDR7, DIEXF, PSMB2, ZC3H15, and ZNF226 compared to controls. Also ChIP-QPCR showed less BRD1-S and BRD1-L binding at introns of WDR7, DIEXF, PSMB2, ZC3H15, and ZNF226 while ChIP-QPCR showed lower and approximately equal BRD1-S and BRD1-L binding at the GZMM promoter and intron. (PDF 433 kb)

Expression arrays from siRNA knockdown and over-expression. After RNA purification from BRD1-S, BRD1-L and HEK293T cells, and cells treated with siRNA directed against BRD1 and cells treated with scrambled siRNA, expression microarray analyses were performed on Affymentrix U219 chips. Individual probe emission intensities were normalized to the mean intensity of the entire array and differential values were calculated as log2(sample) – log2(control) and a cutoff value of +/– 1.5-fold determined if genes were considered upregulated or downregulated. The figure shows MA-plots, where M = log2(sample) – log2(control) and A = ½ × (log2(sample) – log2(control). The two horizontal lines represent 1.5 in expression fold change compared to controls. Probes regulated more than 1.5-fold are indicated by red dots. Probes that are not regulated more than 1.5-fold are colored blue. MA-plots from the top: (A) BRD1-S-V5 stable cells vs. control, BRD1-L-V5 stable cells vs. control and (B) cells treated with siRNA directed against BRD1 vs. cells treated with scrambled siRNA (2 array for method 1, M1.1, and M1.2, and 1 for method 2, M2), and control vs. control (ctrl_v_ctrl, from HEK293T cells treated with scrambled siRNA). The MA-plot of two independent siRNA knockdown control experiments (ctrl_v_ctrl) showed 0.5 % of the probes below or above a 1.5-fold threshold from a total of 49,386 probes. On average, the analysis showed 4.4 % of the probes to be within threshold limits. (PDF 225 kb)

Immunoprecipitation of PBRM1 and identification of BRD1-S and BRD1-L by western blotting. Co-immunoprecipitations of endogenous PBRM1 were performed on BRD1-S, BRD1-L, and untransfected HEK293T cells, by either including antibody against PBRM1 (AB +) or by not including antibody (AB –). BRD1-S-V5 and BRD1-L-V5 were visualized on the western blot using anti-V5 antibody (Invitrogen) and HRP-conjugated anti-mouse IgG antibody. Crude cell extracts (E) from BRD1-S-V5 and BRD1-L-V5 were included as migration markers of the epitope tagged proteins, at approximately 130 kDa and 140 kDa, respectively. (PDF 122 kb)

Chromatin binding profiles for BRD1-S and BRD1-L in relation to identified epigenetic marks in HEK293 cell lines. ChIP-seq data (fastq files), from HEK293 cell lines and cell line derivatives, were obtained from the ENCODE project and mapped to hg19 (for further details see Methods). An in-house histone H3K9ac ChIP-seq dataset was also included in the analysis. The minimum distance from (A) BRD1-S or (B) BRD1-L ChIP-seq peak to transcription factor binding site or histone mark was identified for all 2205 and 1722 peaks, respectively. The results were summarized as histograms with a +/â 30Â kb window from BRD1 binding (distance from BRD1 peaks). The number of peaks within each of the summed bins (columns) can be read on the y-axis (number of peaks). Random ChIP-seq peak regions (Random), with the same composition of chromosomes and peak region sizes, were generated to evaluate if the results could be explained be chance. (PDF 635 kb)

Verification of five selected ChIP-seq promoter targets by ChIP-QPCR. From the 251 common promotor target genes (PTGs) of BRD1-S and BRD1-L, we selected five for validation with ChIP-QPCR. A The promoter loci at the WD repeat domain 7 (WDR7), digestive organ expansion factor homolog (C1ORF107 or DIEXF), proteasome subunit β type, 2 (PSMB2), ZC3H15, and zinc finger protein 226 (ZNF226) showed BRD1-S and BRD1-L binding, while the promoter loci of the lymphocyte expressed gene granzyme M (GZMM) showed no significant binding of the BRD1 isoforms and was subsequently selected as a negative control. The BRD1-S or BRD1-L binding is shown in black whereas the control ChIP-seq (IP with anti-HA antibody) is shown in pink. B Primers were designed to amplify a 80–150 bp sequence in the promoter region or the intron of all six genes (for primer sequences see Additional file 7). ChIP was performed with either anti-V5 antibody (V5) conjugated beads or anti-HA (HA) conjugated beads using extracts from BRD1-S, BRD1-L, or HEK293T cells (for further details see “Methods”). ChIP-QPCR showed higher enrichment of BRD1-S and BRD1-L at the promoter loci of WDR7, DIEXF, PSMB2, ZC3H15, and ZNF226 compared to controls. Also ChIP-QPCR showed less BRD1-S and BRD1-L binding at introns of WDR7, DIEXF, PSMB2, ZC3H15, and ZNF226 while ChIP-QPCR showed lower and approximately equal BRD1-S and BRD1-L binding at the GZMM promoter and intron. (PDF 433 kb)

Spatiotemporal mRNA expression of BRD1 in human brain. A RNA-seq data (obtained from Brainspan; http://www.brainspan.org ) showing the temporal expression of BRD1 across 26 brain regions; primary auditory cortex, core (A1C), amygdaloid complex (AMY), cerebellar cortex (CBC), cerebellum (CB), caudal ganglionic eminence (CGE), dorsolateral prefrontal cortex (DFC), dorsal thalamus (DTH), hippocampus (hippocampal formation) (HIP), posteroventral (inferior) parietal cortex (IPC), inferolateral temporal cortex (area TEv) (ITC), lateral ganglionic eminence (LGE), primary motor cortex (area M1) (M1C), primary motor-sensory cortex (samples) (M1C-S1C), mediodorsal nucleus of thalamus (MD), anterior (rostral) cingulate (medial prefrontal) cortex (MFC), medial ganglionic eminence (MGE), occipital neocortex (Ocx), orbital frontal cortex (OFC), parietal neocortex (PCx), primary somatosensory cortex (area S1) (S1C), posterior (caudal) superior temporal cortex (area 22c) (STC), striatum (STR), temporal neocortex (TCx), upper (rostral) rhombic lip (URL), primary visual cortex (striate cortex) (V1C), ventrolateral prefrontal cortex (VFC). Age on the x-axis is shown as log10(days) (log10(age)) and the gene expression is shown as the average reads per kilobase per million (avg_rpkm). B Expression microarray data (obtained from the Human Brain Transcriptome, HBT; http://hbatlas.org ) showing the temporal expression of BRD1 across six brain regions: neocortex (NCX), hippocampus (HIP), amygdala (AMY), striatum (STR), mediodorsal nucleus of thalamus (MD), cerebellar cortex (CBC). Both datasets show high expression of BRD1 in early fetal stages (until approximately day 180) and both datasets show high expression of BRD1 in the cerebellar cortex throughout the timeline of the datasets. (PDF 208 kb)

Sample IDs, BMIs, ages and case/control classifications for the samples that were included in this study that passed quality control (nâ =â 284). (DOCX 41 kb)

All associating haplotypes (P value 0.05. (DOCX 24 kb)

All associating haplotypes (P value 0.05. (DOCX 24 kb)

Introduction: Early onset absence epilepsy (EOAE) constitutes an idiopathic generalized epilepsy syndrome with typical absences starting before the age of four years. Mutations in SLC2A1, encoding the glucose transporter of the blood-brain barrier (GLUT-1), account for approximately 10% of EOAE cases. The role of SLC2A1 mutations in absence epilepsies with a later onset has not been assessed so far. Therefore, we aimed to compare the role of SLC2A1 mutations in EOAE and Childhood and Juvenile Absence Epilepsy (CAE, JAE). Method: 26 cases with EOAE and 40 probands with CAE or JAE were screened for SCL2A1 mutations by sequence analysis. Extensive phenotyping was performed in patients and family members. Results: Mutations in SLC2A1 were detected in 2/26 EOAE patients and 0/40 patients with familial absence epilepsy. One EOAE patient with a mild phenotype had a variant in exon 8 (c.1008G>C) leading to an amino acid exchange (336Leu>Val), the family history was unremarkable. The other EOAE patient with a very early onset of a severe epilepsy phenotype and movement disorder had a base exchange at position c.1189C>T causing a stop codon (p.Q397X) in exon 9. Familial GTCS were reported in his brother and the paternal grandmother.Conclusion: Our study confirmed the role of SLC2A1 mutation carriers in EOAE and demonstrated that SLC2A1 do not seem to play a major role in CAE and JAE. Since ketogenic diet is a well known treatment option in GLUT-1-deficiency, pediatricians as well as neurologists may revisit the age of onset in patients diagnosed with absence epilepsies.

Epilepsy with myoclonic absences (EMA) is a rare form of generalized epilepsy occurring in childhood and is often difficult to treat. The underlying etiology of EMA is unknown in the majority of patients. Herein, we describe a patient with EMA and intellectual disability who carries a de novo balanced translocation: t(6;22)(p21.32;q11.21). We mapped the translocation breakpoints by fluorescence in situ hybridization (FISH), and the breakpoint at 6p21.32 was found to truncate the N-methyl-d-aspartate (NMDA)-receptor associated gene SYNGAP1. The breakpoint at 22q11.21 was within a highly variable region without known protein-coding genes. Mutations of SYNGAP1 are associated with nonsyndromal intellectual disability (NSID). Two-thirds of the patients described so far also have generalized epilepsy. This finding, together with our report, suggests that dysfunction of SYNGAP1 contributes to the development of generalized epilepsy, including EMA.

Rab11a is a member of the rab-branch of the ras-like small GTP-binding protein superfamily that is associated with both constitutive and regulated secretory pathways. Using a direct procedure for cDNA cloning of small ras-related GTPases, that is based on the screening of eukaryotic cDNA expression libraries using [α-32P]GTP as a probe, we have isolated two cDNA clones encoding rab11a. Both clones share identical coding sequences, but differ in the length and sequence of their 3′ untranslated regions (3′-UTR). Northern blot hybridisation analysis of various human tissues revealed indeed two mRNA species with lengths of 1.0 and 2.3 kb, respectively. Sequence analysis of the cDNAs identified two different putative polyadenylation signals (AATAAA) at positions 927 and 2302 of the larger transcript. In addition, the 3′-UTR of the larger transcript exhibited several AU-rich elements (ARE) that are believed to control gene expression by regulating the rate of mRNA degradation. Southern blots of human DNA digested with several rare restriction enzymes, and separated by pulse-field gel electrophoresis, yielded the same macro-restriction fragment pattern when hybridised with probes that discriminate between the two transcripts. Taken together, these findings imply that the two mRNA species originate from a single gene, which we have mapped to 15q21.3-q22.31, by the use of diferent polyadenylation sites. As expected, both rab11a-cDNAs yielded the same protein product when transiently expressed in COS-1 cells, and surprisingly, upregulated the proteome expression profile (de novo synthesis or posttranslational modification of preexisting proteins) of a few other, yet unknown GTP-binding proteins.

Upregulation of BRD1-S and BRD1-L. A QPCR measurement of BRD1 mRNA in BRD1-S and BRD1-L cells showed a ~24-fold higher BRD1 mRNA levels compared to HEK293T controls (*P

Immunofluorescence staining of HEK293T cells stably expressing V5-epitope tagged BRD1-S and BRD1-L. Stable cell lines (BRD1-S-V5 and BRD1-L-V5) and the untransfected HEK293T cell line (HEK293T) were cultured for 48 h in 2 mL slide flasks followed by fixation in 2 % freshly prepared paraformaldehyde solution. Slides were incubated with anti-V5 antibody (Invitrogen), washed, and incubated with anti-mouse immunoglobulins/FITC F(ab’)2. Cell nuclei were stained by Hoechst staining. From the left: immunoflourescence staining of cell nuclei (Hoechst, blue), imunnoflourescence staining of V5-tagged BRD1-S and BRD1-L (Anti-V5, green), and the two images merged (Merge, purple). (PDF 94 kb)

Immunofluorescence staining of HEK293T cells stably expressing V5-epitope tagged BRD1-S and BRD1-L. Stable cell lines (BRD1-S-V5 and BRD1-L-V5) and the untransfected HEK293T cell line (HEK293T) were cultured for 48 h in 2 mL slide flasks followed by fixation in 2 % freshly prepared paraformaldehyde solution. Slides were incubated with anti-V5 antibody (Invitrogen), washed, and incubated with anti-mouse immunoglobulins/FITC F(ab’)2. Cell nuclei were stained by Hoechst staining. From the left: immunoflourescence staining of cell nuclei (Hoechst, blue), imunnoflourescence staining of V5-tagged BRD1-S and BRD1-L (Anti-V5, green), and the two images merged (Merge, purple). (PDF 94 kb)

Supporting information for ChIP-seq analysis. A Fragmented DNA for ChIP-seq, with an average size of 100â 150Â bp from HEK293T cells and stable BRD1-S-V5 and BRD1-L-V5 expressing HEK293T cells. B IP-western blot of IPed BRD1-S-V5 and BRD1-L-V5 using V5-ChIP-protocol. V5-tagged BRD1-S and BRD1-L was IPed from stable expressing HEK293T cells using anti-V5 antibody conjugated agarose beads (IP-V5) or anti-HA antibody conjugated beads (IP-HA) as control IP. V5-tagged BRD1-S and BRD1-L were visualized on a western blot using an anti-V5 antibody (Invitrogen) and a HRP-conjugated anti-mouse IgG. (PDF 78 kb)

Supporting information for ChIP-seq analysis. A Fragmented DNA for ChIP-seq, with an average size of 100â 150Â bp from HEK293T cells and stable BRD1-S-V5 and BRD1-L-V5 expressing HEK293T cells. B IP-western blot of IPed BRD1-S-V5 and BRD1-L-V5 using V5-ChIP-protocol. V5-tagged BRD1-S and BRD1-L was IPed from stable expressing HEK293T cells using anti-V5 antibody conjugated agarose beads (IP-V5) or anti-HA antibody conjugated beads (IP-HA) as control IP. V5-tagged BRD1-S and BRD1-L were visualized on a western blot using an anti-V5 antibody (Invitrogen) and a HRP-conjugated anti-mouse IgG. (PDF 78 kb)

Upregulation of BRD1-S and BRD1-L. A QPCR measurement of BRD1 mRNA in BRD1-S and BRD1-L cells showed a ~24-fold higher BRD1 mRNA levels compared to HEK293T controls (*P

Polycomb Repressive Complex (PRC) 1 and PRC2 regulate genes involved in differentiation and development. However, the mechanism for how PRC1 and PRC2 are recruited to genes in mammalian cells is unclear. Here we present evidence for an interaction between the transcription factor REST, PRC1, and PRC2 and show that RNF2 and REST co-regulate a number of neuronal genes in human teratocarcinoma cells (NT2-D1). Using NT2-D1 cells as a model of neuronal differentiation, we furthermore showed that retinoic-acid stimulation led to displacement of PRC1 at REST binding sites, reduced H3K27Me3, and increased gene expression. Genome-wide analysis of Polycomb binding in Rest−/− and Eed−/− mouse embryonic stem (mES) cells showed that Rest was required for PRC1 recruitment to a subset of Polycomb regulated neuronal genes. Furthermore, we found that PRC1 can be recruited to Rest binding sites independently of CpG islands and the H3K27Me3 mark. Surprisingly, PRC2 was frequently increased around Rest binding sites located in CpG-rich regions in the Rest−/− mES cells, indicating a more complex interplay where Rest also can limit PRC2 recruitment. Therefore, we propose that Rest has context-dependent functions for PRC1- and PRC2- recruitment, which allows this transcription factor to act both as a recruiter of Polycomb as well as a limiting factor for PRC2 recruitment at CpG islands., Author Summary Multicellular organisms are composed of a large number of specialized cell types that all originate from the Embryonic Stem cell (ES cell). It is crucial for the maintenance of naïve ES cells that developmental genes are kept in an off-state until appropriate differentiation stimuli are received. Polycomb Repressive Complexes, PRC1 and PRC2, are bound at and repress the activity of a large number of key developmental genes in ES cells and at different stages of differentiation. While in Drosophila the PRC complexes are recruited to DNA elements called Polycomb Response Elements (PREs), through the interaction with transcription factors; examples of such factors remain poorly characterized in mammals. We here demonstrate that the transcription factor Rest interacts with and is required for recruitment of PRC1 and PRC2 to a subset of Rest target genes in mouse embryonic stem (mES) cells. In line with REST being a repressor of neuronal genes, we found that PRC1 and PRC2 co-localized with REST at genes involved in neuronal development and got displaced during neuronal differentiation. Based on our data we propose that the PRC1 and PRC2 complexes function as co-repressors for Rest to control the timed expression of developmental genes in the process of cellular differentiation.

Early prenatal diagnosis of the fragile X was attempted in 44 pregnancies, including one twin pregnancy at risk of Martin-Bell (MB) syndrome. The sex ratio was 24M:21F. The fragile site was reproducibly demonstrated in cultured chorionic villus (CV) cells in eight male and five female fetuses. Six of the male and three of the female fetuses were terminated. Simultaneous RFLP analysis provided confirmative data with flanking DNA markers in 3 of 13 analysed cases. Recombination and/or non-informativeness at available distal and/or proximal loci were found in nine cases. In one male fetus, discordance between the haplotype and cytogenetics (fragile-X-negative) suggested the presence of a normal male transmitter, a double meiotic cross-over within the region, or a false-negative cytogenetic diagnosis. However, discordance between prenatal and post-termination/postnatal cytogenetic findings was not observed in this series. The use of excess thymidine for induction of the fragile X in cultured CV cells provided in the majority of cases a safe and rapid method for cytogenetic diagnosis, with options for early induced termination in fragile-X-positive pregnancies, for simultaneous RFLP analysis, and for subsequent second-trimester analysis of fetal blood in complicated cases.

Made available in DSpace on 2019-09-12T16:53:26Z (GMT). No. of bitstreams: 0 Previous issue date: 2006 Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD) The extensive heterogeneity underlying the genetic component of mental retardation (MR) is the main cause for our limited understanding of the aetiology of this highly prevalent condition. Hence we set out to identify genes involved in MR. We investigated the breakpoints of two balanced X;autosome translocations in two unrelated female patients with mild/moderate MR and found that the Xp11.2 breakpoints disrupt the novel human KIAA1202 (hKIAA1202) gene in both cases. We also identified a missense exchange in this gene, segregating with the Stocco dos Santos XLMR syndrome in a large four-generation pedigree but absent in > 1,000 control X-chromosomes. Among other phenotypic characteristics, the affected males in this family present with severe MR, delayed or no speech, seizures and hyperactivity. Molecular studies of hKIAA1202 determined its genomic organisation, its expression throughout the brain and the regulation of expression of its mouse homologue during development. Transient expression of the wild-type KIAA1202 protein in HeLa cells showed partial colocalisation with the F-actin based cytoskeleton. On the basis of its domain structure, we argue that hKIAA1202 is a new member of the APX/Shroom protein family. Members of this family contain a PDZ and two ASD domains of unknown function and have been shown to localise at the cytoskeleton, and play a role in neurulation, cellular architecture, actin remodelling and ion channel function. Our results suggest that hKIAA1202 may be important in cognitive function and/or development. Max Planck Inst Mol Genet, Dept Human Mol Genet, D-14195 Berlin, Germany; ULP, INSERM, CNRS, Inst Genet & Biol Mol & Cellularie,Dept Mol Patho, Illkirch Graffenstaden, France; Greenwood Genet Ctr, JC Self Res Inst, Greenwood, SC 29646 USA; Univ Ulm, Dept Human Genet, Ulm, Germany; Wilhelm Johannsen Ctr Funct Genome Res, Dept Med Biochem & Genet, Copenhagen, Denmark; INSERM, Serv Genet, Tours, France; Katholieke Univ Leuven Hosp, Ctr Human Genet, B-3000 Louvain, Belgium; CNRS, INSERM, Inst Cochin Genet Mol, Paris, France; Univ Nijmegen St Radboud Hosp, Med Ctr, Dept Human Genet, Nijmegen, Netherlands; Univ Adelaide, Dept Med Genet, Womens & Childrens Hosp, Adelaide, SA, Australia; Univ Adelaide, Dept Pediat, Adelaide, SA, Australia; Hop Univ Strasbourg, Serv Genet Med, Strasbourg, France; Universidade de Taubaté (Unitau), Inst Butantan, Genet Lab, Sao Paulo, Brazil; Erasmus MC, Dept Genet, Rotterdam, Netherlands