A functional tetranucleotide (AAAT) polymorphism in an Alu element in the NF1 gene is associated with mental retardation.

Mental retardation (MR) is frequent in neurofibromatosis type 1 (NF1). Allele 5 of a tetranucleotide polymorphism in an Alu element (GXAlu) localized in intron 27b of the NF1 gene has previously been associated with autism. We considered that the microsatellite GXAlu could also represent a risk factor in MR without autism. We developed a rapid method for genotyping by non-denaturing HPLC and assayed the allelic variation of GXAlu marker on in vitro gene expression in Cos-7 cells. A French population of 157 individuals (68 non syndromic non familial MR (NS-MR) patients diagnosed in the University Hospital of Tours; 89 controls) was tested in a case-control assay. We observed a significant association (χ(2)=7.96; p=0.005) between alu4 carriers (7 AAAT repeats) and MR (OR: 7.86; 95% C.I.: 2.13-28.9). The relative in vitro expression of a reporter gene encoding chloramphenicol acetyl transferase (CAT) was higher for alu4 and alu5, suggesting a regulation effect for these alleles on gene expression in vivo. Our results showed an association with a polymorphism regulating the NF1 gene or other genes during brain development.

Study of the serotonin transporter (SLC6A4) and BDNF genes in French patients with non syndromic mental deficiency.

BACKGROUND: Mental deficiency has been linked to abnormalities in cortical neuronal network connectivity and plasticity. These mechanisms are in part under the control of two interacting signalling pathways, the serotonergic and the brain-derived neurotrophic (BDNF) pathways. The aim of the current paper is to determine whether particular alleles or genotypes of two crucial genes of these systems, the serotonin transporter gene (SLC6A4) and the brain-derived neurotrophic factor gene (BDNF), are associated with mental deficiency (MD). METHODS: We analyzed four functional polymorphisms (rs25531, 5-HTTLPR, VNTR, rs3813034) of the SLC6A4 gene and one functional polymorphism (Val66 Met) of the BDNF gene in 98 patients with non-syndromic mental deficiency (NS-MD) and in an ethnically matched control population of 251 individuals. RESULTS: We found no significant differences in allele and genotype frequencies in the five polymorphisms studied in the SLC6A4 and BDNF genes of NS-MD patients versus control patients. While the comparison of the patterns of linkage disequilibrium (D') in the control and NS-MD populations revealed a degree of variability it did not, however, reach significance. No significant differences in frequencies of haplotypes and genotypes for VNTR/rs3813034 and rs25531/5-HTTLPR were observed. CONCLUSION: Altogether, results from the present study do not support a role for any of the five functional polymorphisms of SLC6A4 and BDNF genes in the aetiology of NS-RM. Moreover, they suggest no epistatic interaction in NS-MD between polymorphisms in BDNF and SLC6A4. However, we suggest that further studies on these two pathways in NS-MD remain necessary.

Association study of the ubiquitin conjugating enzyme gene UBE2H in sporadic ALS.

Ubiquitin inclusions represent a cytopathological hallmark of ALS. The ubiquitin-dependent protein degradation pathway may also be involved in the pathophysiology of SOD1 mutated ALS cases as demonstrated in transgenic animals. UBE2H is an ubiquitin conjugating enzyme known to act on histones and cytoskeletal proteins, both involved in the degenerative pathway of the motor neuron. We screened the whole coding sequence of the UBE2H gene in 24 sporadic ALS (SALS) patients using single strand conformation polymorphism (SSCP). All variants detected by SSCP were analysed by genomic DNA sequencing. We found one known polymorphism (rs12539800) and two new synonymous single nucleotide polymorphisms (SNP) (nG78A and nG501A). The allele distribution of the rs12539800 (A336G) SNP were tested for association in 252 SALS patients and 357 controls. The allele and genotype distributions were identical in the two groups. The UBE2H gene is not implicated in SALS; however, the ubiquitin pathway is worthy of further investigation in ALS.

Autism and nonsyndromic mental retardation associated with a de novo mutation in the NLGN4X gene promoter causing an increased expression level.

BACKGROUND: Pathogenic mutations in the X-linked Neuroligin 4 gene (NLGN4X) in autism spectrum disorders (ASDs) and/or mental retardation (MR) are rare. However, nothing is known regarding a possible altered expression level of NLGN4X that would be caused by mutations in regulatory sequences. We investigated this issue by analyzing these regions in patients with ASDs and no mutation in the NLGN4X coding sequence. METHODS: We studied 96 patients who met all DSM-IV criteria for autism. The entire coding sequence and the regulatory sequences of the NLGN4X gene were analyzed by polymerase chain reaction and direct sequencing. RESULTS: We identified a de novo 1 base pair (-335G>A) substitution located in the promoter region in a patient with autism and nonsyndromic profound MR. Interestingly, this variation is associated with an increased level of the NLGN4X transcript in the patient compared with male control subjects as well as his father. Further in vitro luciferase reporter and electrophoretic mobility shift assays confirmed, respectively, that this mutation increases gene expression and is probably caused by altered binding of transcription factors in the mutated promoter sequence. CONCLUSIONS: This result brings further insight about the phenotypic spectrum of NLGN4X mutations and suggests that the analysis of the expression level of NLGN4X might detect new cases.

Effect of the oligodendrocyte myelin glycoprotein (OMgp) on the expansion and neuronal differentiation of rat neural stem cells.

The oligodendrocyte myelin glycoprotein (OMgp) inhibits axon regeneration after injury in the adult mammalian central nervous system. However its function during brain development remains largely unknown. The present study aims to analyze a possible role for OMgp during neurogenesis. We showed that neural stem cells (NSC) extracted from the whole mesencephalon of rat embryos (E14) and cultured as free floating neurospheres expressed both OMgp and its receptor Nogo-R1. An over-expression of OMgp affected NSC expansion by reducing cell proliferation, but did not affect their differentiation into neurons. These findings indicate a new role for OMgp during brain development as a possible regulator of neurogenesis. Moreover, they suggest a possible implication for OMG gene in the etiology of neurofibromatosis type 1 forms characterized by a deletion of the NF1 gene locus containing OMG.

Haploinsufficiency of the GPD2 gene in a patient with nonsyndromic mental retardation.

We have investigated the chromosome abnormalities in a female patient exhibiting mild nonsyndromic mental retardation. The patient carries a de novo balanced reciprocal translocation 46,XX,t(2;7)(q24.1;q36.1). Physical mapping of the breakpoints by fluorescent in situ hybridization experiments revealed the disruption of the GPD2 gene at the 2q24.1 region. This gene encodes the mitochondrial glycerophosphate dehydrogenase (mGPDH), which is located on the outer surface of the inner mitochondrial membrane, and catalyzes the unidirectional conversion of glycerol-3-phosphate (G3P) to dihydroxyacetone phosphate with concomitant reduction of the enzyme-bound FAD. Molecular and functional studies showed approximately a twofold decrease of GPD2 transcript level as well as decreased activity of the coded mGPDH protein in lymphoblastoid cell lines of the patient compared to controls. Bioinformatics analysis allowed us to confirm the existence of a novel transcript of the GPD2 gene, designated GPD2c, which is directly disrupted by the 2q breakpoint. To validate GPD2 as a new candidate gene for mental retardation, we performed mutation screening of the GPD2 gene in 100 mentally retarded patients; however, no mutations have been identified. Nevertheless, our results propose that a functional defect of the mGPDH protein could be associated with mental retardation, suggesting that GPD2 gene could be involved in mental retardation in some cases.

Transmission disequilibrium study of an oligodendrocyte and myelin glycoprotein gene allele in 431 families with an autistic proband.

Autistic disorder is a neurodevelopmental disorder where genetic factors play an important role. We previously described an association between a subgroup of French autistic patients and an allele of a non-synonymous single nucleotide polymorphism (nsSNP: OMGP62 G>A or rs11080149) in the gene coding for the oligodendrocyte and myelin glycoprotein (OMG), located at 7Mb from the marker D17S250, linked to autism in two independent genome scan studies. We report a study on 431 families with 1 affected child from different origins: French Canada (n=262), Italy (n=123) and United States (n=46). We analyzed the transmission of the rs11080149 alleles from parents to their affected children. There was a preferential transmission of the G allele from parents to affected children (p=0.0017) in the overall sample. Paternal and maternal transmission rates were both skewed. Taking into account our previous results obtained in a French group of patients, where we observed an association with allele A, a direct role of this polymorphism is improbable in autism. The associations observed in Japanese and French patients, the linkage studies and the present work speak in favor of the existence of a susceptibility gene for autism in the NF1 locus.

The oligodendrocyte-myelin glycoprotein gene is highly expressed during the late stages of myelination in the rat central nervous system.

Oligodendrocyte-myelin glycoprotein (OMgp) is expressed on the surface of oligodendrocytes and neurones and is thought to inhibit axonal regeneration after brain injury in adult, like Nogo and myelin-associated glycoprotein (MAG). We previously observed that the OMgp gene locus on chromosome 17 could be associated with autism, a developmental disorder. The aim of the present study was to characterise the developmental expression of OMgp mRNA in the central nervous system. First we determined the rat OMgp gene sequence and compared it with the human and mouse sequences. Several regions, putative sites for the fixation of transcription factors, are conserved between these three species in the unique intron of this gene. Using quantitative and semi-quantitative RT-PCR, we studied OMgp gene expression in rat brain during post-natal development. We found that OMgp mRNA expression was developmentally regulated, with a peak of expression in the late stages of myelination. We observed a similar profile in oligodendrocyte cultures, in absence of neurones, suggesting that OMgp mRNA expression by oligodendrocytes was independent of axonal influence. Our observations suggest that OMgp is a late marker of myelination, which could be implicated in the arrest of oligodendrocyte proliferation, arrest of myelination or compaction of myelin.

Oligodendrocyte myelin glycoprotein growth inhibition function requires its conserved leucine-rich repeat domain, not its glycosylphosphatidyl-inositol anchor.

The oligodendrocyte myelin glycoprotein (OMgp) inhibits neurite outgrowth and axonal regeneration after brain injury, but its normal function remains unknown. Several observations suggest its implication in cell growth regulation. Here we report an analysis of the domain requirement in OMgp proliferation inhibitory function. We first studied the OMgp protein sequence in 14 mammal species and observed a high conservation of its leucine-rich repeat (LRR) domain. The deletion of this LRR domain is responsible for a total loss of function in an in vitro expression system. The possible three-dimensional structure of the LRR domain of OMgp was modelled using the structure of Yersinia pestis YopM cytotoxin as a template. The predicted arrangement of the LRR segments is compatible with a function of OMgp as a binding protein. The OMgp is a glycosylphosphatidyl-inositol-linked protein anchored in the plasma membrane of oligodendrocytes and neurones. Using deletion mutagenesis, we demonstrated the dispensability of the glycosylphosphatidyl-inositol anchor for OMgp proliferation inhibition function. Our results suggest that OMgp is part of a receptor complex, either as a coreceptor or as a membrane-bound or soluble ligand, involved in the transmission of a growth suppressive signal.