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Biomedical subjects

B Chabot

Publications and source records attributed to B Chabot.

At least 19 recordsLinked to original sources
PubMed2005

Are eye movement abnormalities indicators of genetic vulnerability to schizophrenia?

UNLABELLED: Fifty to eighty-five percent of schizophrenic patients are impaired on ocular pursuit paradigms. However, results regarding the relatives are more discordant. The aim of this study was to investigate whether eye movement disorders could be a vulnerability marker of schizophrenia. METHOD: Twenty-one schizophrenic patients (DSM-IV), 31 first-degree relatives of those patients without schizophrenic spectrum disorders, and two groups of healthy controls matched by age and sex were included. Three oculomotor tasks (smooth pursuit, reflexive saccades and antisaccades) were used. RESULTS: Patients had a lower averaged gain (P= 0.035) during smooth pursuit than controls, made less correct visually guided saccades (P< 0.001) and more antisaccades errors (P= 0.002) than controls. In contrast, none of the comparison between the relatives and their controls was significant. CONCLUSION: Schizophrenic patients were impaired on smooth pursuit and antisaccade paradigms. None of these impairments was, however, observed in their first-degree relatives. Our results suggest that the eye movement parameters tested could not be considered as vulnerability markers for schizophrenia.

Adult↗
PubMed2003

[Familial idiopathic striato-pallido-dentate calcifications: clinical and brain imaging study in a family].

Familial idiopathic basal ganglia calcification (FIBGC) is a rare condition and its pathophysiology has not so far been elucidated. We report the results of a clinical study in two patients of a family affected with FIBGC. Brain imaging with 18-FDG-PET was performed in one. Psychiatric and cognitive troubles were the main clinical symptoms. Basal ganglia calcifications were associated with white matter lesions. The PET study performed in one patient revealed a striatal and a posterior cingulate hypometabolism. Posterior cingulate gyrus is involved in episodic memory processing, and could be involved in episodic memory deficit observed in this patient. These results suggest that a cortical dysfunction could be associated to the disease. The underlying mechanism, that could be a neuronal loss, a cortical deafferentation or an alteration of synaptic transmission, remains to be elucidated.

Adult↗
PubMed2001

hnRNP A1 may interact simultaneously with telomeric DNA and the human telomerase RNA in vitro.

The hnRNP A1 protein and a shortened derivative (UP1) promote telomere elongation in mammalian cells. In support of a direct role for A1 in telomere biogenesis, we have shown that the recombinant UP1 protein binds to telomeric DNA sequences in vitro, and pulls down telomerase activity from a cell extract. Here we show that A1/UP1 can interact directly with the RNA component of human telomerase (hTR). A portion of A1/UP1 that contains RNA recognition motif 2 (RRM2) is sufficient for an interaction with the first 208 nt of hTR. Given that the portion of A1/UP1 that contains RRM1 is sufficient for binding to a telomeric DNA oligonucleotide, we have tested whether A1/UP1 can interact simultaneously with both nucleic acids. Using a chromatography assay, we find that A1/UP1 bound to hTR can interact with telomeric DNA. Notably, these interactions are sufficiently robust to withstand incubation in a cell extract. Our results suggest that hnRNP A1 may help recruit telomerase to the ends of chromosomes.

Binding Sites↗
PubMed2001

A novel mutation in the neurofibromatosis type 1 (NF1) gene promotes skipping of two exons by preventing exon definition.

Using a protein truncation assay, we have identified a new mutation in the neurofibromatosis type 1 (NF1) gene that causes a severe defect in NF1 pre-mRNA splicing. The mutation, which consists of a G to A transition at position +1 of the 5' splice site of exon 12a, is associated with the loss of both exons 11 and 12a in the NF1 mRNA. Through the use of in vivo and in vitro splicing assays, we show that the mutation inactivates the 5' splice site of exon 12a, and prevents the definition of exon 12a, a process that is normally required to stimulate the weak 3' splice site of exon 12a. Because the 5' splice site mutation weakens the interaction of splicing factors with the 3' splice site of exon 12a, we propose that exon 11/exon 12a splicing is also compromised, leading to the exclusion of both exons 11 and 12a. Our results provide in vivo support for the importance of the exon definition model during NF1 splicing, and suggest that the NF1 region containing exons 11 and 12a plays an important role in the activity of neurofibromin.

Alternative Splicing↗
PubMed2001

Dimethyl sulfoxide affects the selection of splice sites.

Depending on the cell lines and cell types, dimethyl sulfoxide (Me2SO) can induce or block cell differentiation and apoptosis. Although Me2SO treatment alters many levels of gene expression, the molecular processes that are directly affected by Me2SO have not been clearly identified. Here, we report that Me2SO affects splice site selection on model pre-mRNAs incubated in a nuclear extract prepared from HeLa cells. A shift toward the proximal pair of splice sites was observed on pre-mRNAs carrying competing 5'-splice sites or competing 3'-splice sites. Because the activity of recombinant hnRNP A1 protein was similar when added to extracts containing or lacking Me2SO, the activity of endogenous A1 proteins is probably not affected by Me2SO. Notably, in a manner reminiscent of SR proteins, Me2SO activated splicing in a HeLa S100 extract. Moreover, the activity of recombinant SR proteins in splice site selection in vitro was improved by Me2SO. Polar solvents like DMF and formamide similarly modulated splice site selection in vitro but formamide did not activate a HeLa S100 extract. We propose that Me2SO improves ionic interactions between splicing factors that contain RS-domains. The direct impact of Me2SO on alternative splicing may explain, at least in part, the different and sometimes opposite effects of Me2SO on cell differentiation and apoptosis.

3' Untranslated Regions↗
PubMed2001

Schizophrenia and familial idiopathic basal ganglia calcification: a case report.

BACKGROUND: Familial idiopathic basal ganglia calcification (FIBGC) is generally associated with neurological and psychiatric symptoms. An association between FIBGC and schizophrenia has been described but it remains uncertain. We studied the relationship between the presence and extent of basal ganglia calcification and schizophrenia in a multiply affected family. METHOD: Symmetrical basal ganglia calcifications (BGC) were detected on computerized tomography (CT) in a schizophrenic proband and led us to carry out CTs and standardized psychiatric evaluations (SADS--Endicott & Spitzer, 1978) in all available first-degree relatives (mother and six siblings). RESULTS: Five subjects had BGC, including three subjects diagnosed as schizophrenic. Three subjects had no BGC and none of them was diagnosed as schizophrenic. We subdivided the BGC into three groups: massive (pallidum, striatum and dentate nuclei affected); medium (pallidum and striatum); and mild (pallidum only). The two subjects with massive BGC and one of the two with medium BGC had schizophrenia. The subject with mild BGC had no psychotic symptoms. CONCLUSION: Our results are consistent with the hypothesis that BGC favours the occurrence of a schizophrenia-like syndrome and that the risk of occurrence of this syndrome is proportional to the extent of calcification. These findings support the hypothesis that schizophrenia is determined by a disruption of thalamo-cortico-striatal circuits.

Adult↗
PubMed2000

Heterogeneous nuclear ribonucleoprotein A1 and UP1 protect mammalian telomeric repeats and modulate telomere replication in vitro.

The heterogeneous nuclear ribonucleoprotein A1 protein and a shortened derivative (UP1) promote telomere elongation in mammalian cells. To gain insights into the function of A1/UP1 in telomere biogenesis, we have investigated the binding properties of recombinant A1/UP1 and derivatives to single-stranded DNA oligonucleotides. Our results indicate that UP1 prefers to bind to DNA carrying single-stranded telomeric extensions at the 3' terminus. The RNA recognition motif 1 is sufficient for strong and specific binding to oligomers carrying vertebrate telomeric repeats. We find that the binding of A1/UP1 protects telomeric sequences against degradation by endo- and exonucleases. Moreover, A1/UP1 binding prevents extension by telomerase and terminal deoxynucleotidyltransferase and inhibits rNTP-dependent DNA synthesis in vitro. These observations are consistent with the hypothesis that A1/UP1 is a telomere end-binding protein that plays a role in the maintenance of long 3' overhangs.

Base Sequence↗
PubMed2000

Control of hnRNP A1 alternative splicing: an intron element represses use of the common 3' splice site.

Alternative splicing of exon 7B in the hnRNP A1 pre-mRNA produces mRNAs encoding two proteins: hnRNP A1 and the less abundant A1B. We have reported the identification of several intron elements that contribute to exon 7B skipping. In this study, we report the activity of a novel element, conserved element 9 (CE9), located in the intron downstream of exon 7B. We show that multiple copies of CE9 inhibit exon 7B-exon 8 splicing in vitro. When CE9 is inserted between two competing 3' splice sites, a single copy of CE9 decreases splicing to the distal 3' splice site. Our in vivo results also support the conclusion that CE9 is a splicing modulator. First, inserting multiple copies of CE9 into an A1 minigene compromises the production of fully spliced products. Second, one copy of CE9 stimulates the inclusion of a short internal exon in a derivative of the human beta-globin gene. In this case, in vitro splicing assays suggest that CE9 decreases splicing of intron 1, an event that improves splicing of intron 2 and decreases skipping of the short internal exon. The ability of CE9 to act on heterologous substrates, combined with the results of a competition assay, suggest that the activity of CE9 is mediated by a trans-acting factor. Our results indicate that CE9 represses the use of the common 3' splice site in the hnRNP A1 alternative splicing unit.

Alternative Splicing↗
PubMed1999

An element in the 5' common exon of the NCAM alternative splicing unit interacts with SR proteins and modulates 5' splice site selection.

The neural cell adhesion molecule (NCAM) gene contains an 801 nt exon that is included preferentially in neuronal cells. We have set up an in vitro splicing system that mimics the neuro-specific alternative splicing profile of NCAM exon 18. Splicing regulation is observed using model pre-mRNAs that contain competing 5' or 3' splice sites, suggesting that distinct pathways regulate NCAM 5' and 3' splice site selection. While inclusion of exon 18 is the predom-inant choice in neuronal cells, an element in the 5' common exon 17 improves exon 17/exon 19 splicing in a neuronal cell line. A similar behavior is observed in vitro as the element can stimulate the 5' splice site of exon 17 or a heterologous 5' splice site. The minimal 32 nt sequence of the exon 17 enhancer consists of purine stretches and A/C motifs. Mutations in the purine stretches compromise the binding of SR proteins and decreases splicing stimulation in vitro. Mutations in the A/C motifs do not affect SR protein binding but reduce enhancing activity. Our results suggest that the assembly of an enhancer complex containing SR proteins in a 5' common exon ensures that NCAM mRNAs lacking exon 18 are made in neuronal cells.

Alternative Splicing↗
PubMed1999

Modulation of exon skipping by high-affinity hnRNP A1-binding sites and by intron elements that repress splice site utilization.

The RNA-binding protein hnRNP A1 is a splicing regulator produced by exclusion of alternative exon 7B from the A1 pre-mRNA. Each intron flanking exon 7B contains a high-affinity A1-binding site. The A1-binding elements promote exon skipping in vivo, activate distal 5' splice site selection in vitro and improve the responsiveness of pre-mRNAs to increases in the concentration of A1. Whereas the glycine-rich C-terminal domain of A1 is not required for binding, it is essential to activate the distal 5' splice site. Because A1 complexes can interact simultaneously with two A1-binding sites, we propose that an interaction between bound A1 proteins facilitates the pairing of distant splice sites. Based on the distribution of putative A1-binding sites in various pre-mRNAs, an A1-mediated change in pre-mRNA conformation may help define the borders of mammalian introns. We also identify an intron element which represses the 3' splice site of exon 7B. The activity of this element is mediated by a factor distinct from A1. Our results suggest that exon 7B skipping results from the concerted action of several intron elements that modulate splice site recognition and pairing.

Alternative Splicing↗
PubMed1999

Month of birth in deficit and non-deficit schizophrenic patients.

In order to test the hypothesis that an excess of summer births is a risk factor for deficit syndrome, the month of birth was studied in 53 deficit schizophrenic patients compared to 158 non-deficit patients. No significant difference in terms of month of birth or season of birth was observed between deficit and non-deficit patients, suggesting that summer births might not be a risk factor for deficit schizophrenia.

Adult↗
PubMed1998

Telomere elongation by hnRNP A1 and a derivative that interacts with telomeric repeats and telomerase.

Telomeric DNA of mammalian chromosomes consists of several kilobase-pairs of tandemly repeated sequences with a terminal 3' overhang in single-stranded form. Maintaining the integrity of these repeats is essential for cell survival; telomere attrition is associated with chromosome instability and cell senescence, whereas stabilization of telomere length correlates with the immortalization of somatic cells. Telomere elongation is carried out by telomerase, an RNA-dependent DNA polymerase which adds single-stranded TAGGGT repeats to the 3' ends of chromosomes. While proteins that associate with single-stranded telomeric repeats can influence tract lengths in yeast, equivalent factors have not yet been identified in vertebrates. Here, it is shown that the heterogeneous nuclear ribonucleoprotein A1 participates in telomere biogenesis. A mouse cell line deficient in A1 expression harbours telomeres that are shorter than those of a related cell line expressing normal levels of A1. Restoring A1 expression in A1-deficient cells increases telomere length. Telomere elongation is also observed upon introduction of exogenous UP1, the amino-terminal fragment of A1. While both A1 and UP1 bind to vertebrate single-stranded telomeric repeats directly and with specificity in vitro, only UP1 can recover telomerase activity from a cell lysate. These findings establish A1/UP1 as the first single-stranded DNA binding protein involved in mammalian telomere biogenesis and suggest possible mechanisms by which UP1 may modulate telomere length.

Animals↗
PubMed1997

An intron element modulating 5' splice site selection in the hnRNP A1 pre-mRNA interacts with hnRNP A1.

The hnRNP A1 pre-mRNA is alternatively spliced to yield the A1 and A1b mRNAs, which encode proteins differing in their ability to modulate 5' splice site selection. Sequencing a genomic portion of the murine A1 gene revealed that the intron separating exon 7 and the alternative exon 7B is highly conserved between mouse and human. In vitro splicing assays indicate that a conserved element (CE1) from the central portion of the intron shifts selection toward the distal donor site when positioned in between the 5' splice sites of exon 7 and 7B. In vivo, the CE1 element promotes exon 7B skipping. A 17-nucleotide sequence within CE1 (CE1a) is sufficient to activate the distal 5' splice site. RNase T1 protection/immunoprecipitation assays indicate that hnRNP A1 binds to CE1a, which contains the sequence UAGAGU, a close match to the reported optimal A1 binding site, UAGGGU. Replacing CE1a by different oligonucleotides carrying the sequence UAGAGU or UAGGGU maintains the preference for the distal 5' splice site. In contrast, mutations in the AUGAGU sequence activate the proximal 5' splice site. In support of a direct role of the A1-CE1 interaction in 5'-splice-site selection, we observed that the amplitude of the shift correlates with the efficiency of A1 binding. Whereas addition of SR proteins abrogates the effect of CE1, the presence of CE1 does not modify U1 snRNP binding to competing 5' splice sites, as judged by oligonucleotide-targeted RNase H protection assays. Our results suggest that hnRNP A1 modulates splice site selection on its own pre-mRNA without changing the binding of U1 snRNP to competing 5' splice sites.

Alternative Splicing↗
PubMed1997

A highly stable duplex structure sequesters the 5' splice site region of hnRNP A1 alternative exon 7B.

Exon 7B in the hnRNP A1 pre-mRNA is alternatively spliced to yield A1 and A1(B), two proteins that differ in their ability to modulate 5' splice site selection. Sequencing the murine intron downstream of exon 7B revealed the existence of several regions of similarity to the corresponding human intron. In vitro splicing assays indicate that an 84-nt region (CE6IO) decreases splicing to the proximal 5' splice site in a pre-mRNA carrying the 5' splice sites of exon 7 and 7B. In vivo, the CE6IO element promotes exon 7B skipping in pre-mRNAs expressed from a mini-gene containing the hnRNP A1 alternative splicing unit. Using oligonucleotide-targeted RNase H cleavage assays, we provide support for the existence of highly stable base pairing interactions between CE6IO and the 5' splice site region of exon 7B. Duplex formation occurs in naked pre-mRNA, resists incubation in splicing extracts, and is associated with a reduction in the assembly of U1 snRNP-dependent complexes to the 5' splice site of exon 7B. Our results demonstrate that pre-mRNA secondary structure plays an important role in promoting exon 7B skipping in the A1 pre-mRNA.

Alternative Splicing↗
PubMed1997

Natural base-pairing interactions between 5' splice site and branch site sequences affect mammalian 5' splice site selection.

In the murine gene encoding the neuronal cell adhesion molecule (NCAM), the integrity of the 5' splice site of exon 18 (E18) is essential for regulation of alternative splicing. To further study the contribution of 5' splice site sequences, we used a simple NCAM pre-mRNA containing a portion of E18 fused to E19 and separated by a shortened intron. This RNA is spliced in vitro to produce five sets of lariat intermediates and products, the most abundant set displaying aberrant migration in acrylamide/urea gels. Base pairing interactions between positions +5 and +8 of the intron and positions -3 and -6 from the branch point were responsible for the faster migration of this set of lariat molecules. To test whether the duplex structure forms earlier and contributes to 5' splice site selection, we used NCAM substrates carrying the 5' splice sites of E17 and E18 in competition for the 3' splice site of E19. Mutations upstream of the major branch site improve E18/E19 splicing in NIH3T3 extracts, whereas compensatory mutations at positions +7 and +8 neutralize the effect of branch site mutations and curtail E18/E19 splicing. Our data suggest that duplex formation occurs early and interferes with the assembly of complexes initiated on the 5' splice site of NCAM E18. This novel type of intron interaction may exist in the introns of other mammalian pre-mRNAs.

3T3 Cells↗
PubMed1996

The nuclear matrix protein p255 is a highly phosphorylated form of RNA polymerase II largest subunit which associates with spliceosomes.

The monoclonal antibody CC-3 recognizes a phosphodependent epitope on a 255 kDa nuclear matrix protein (p255) recently shown to associate with splicing complexes as part of the [U4/U6.U5] tri-snRNP particle [Chabot et al. (1995) Nucleic Acids Res. 23, 3206-3213]. In mouse and Drosophila cultured cells the electrophoretic mobility of p255, faster in the latter species, was identical to that of the hyperphosphorylated form of RNA polymerase II largest subunit (IIo). The CC-3 immunoreactivity of p255 was abolished by 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole, which is known to cause the dephosphorylation of the C-terminal domain of subunit IIo by inhibiting the TFIIH-associated kinase. The identity of p255 was confirmed by showing that CC-3-immunoprecipitated p255 was recognized by POL3/3 and 8WG16, two antibodies specific to RNA polymerase II largest subunit. Lastly, the recovery of RNA polymerase II largest subunit from HeLa splicing mixtures was compromised by EDTA, which prevents the interaction of p255 with splicing complexes and inhibits splicing. Our results indicate that p255 represents a highly phosphorylated form of RNA polymerase II largest subunit physically associated with spliceosomes and possibly involved in coupling transcription to RNA processing.

3T3 Cells↗
PubMed1996

Autoantigenic epitopes on hY5 Ro RNA are distinct from regions bound by the 60-kDa Ro and La proteins.

We recently reported the identification in human anti-Ro serum of Abs specifically immunoprecipitating deproteinized hY5 RNA. In the present report, we characterized the epitopes recognized by anti-hY5 RNA Abs. Using deletion and site-directed mutagenesis of hY5 cDNA and in vitro transcribed RNAs with intact and 3'-shortened ends, we have defined two conformational antigenic determinants distinct from the regions known to bind Ro and La proteins. One of these epitopes (epitope A) is present in the middle portion of hY5 RNA and is dependent on the presence of a four-nucleotide sequence (AACC at position 58-61) that may form a single-stranded loop. Deleting these four nucleotides or modifying the stem structures proximal or distal to this loop abolishes recognition of the mutated RNAs by Abs. The second epitope (epitope B) requires the presence of another four-nucleotide sequence (CUUG at position 74-77) in between the Ro and La binding sites. Deleting this CUUG sequence or modifying nucleotides on the 5' side of the stem structure below the Ro60 binding site severely compromises the interaction with Abs. Since Abs to deproteinized hY RNAs are restricted to anti-hY5 RNA and target determinants not involved in interactions with known hY5 RNA-binding proteins, human RohY5 particles may play a direct role in the immunization process, leading to the production of anti-hY5 RNA autoantibodies.

Antibody Specificity↗