Biomedical subjects

P Pontarotti

Publications and source records attributed to P Pontarotti.

At least 37 records · Page 2Linked to original sources

PubMed1996

Expressed sequence tags identify human isologs of the ARF-dependent phospholipase D.

By searching into Expressed Sequence Tags databases (dbEST) using Blast X algorithm software and a plant phospholipase D as template, we have identified a cDNA from human brain (Z45777) which encodes for a protein similar to the amino acid region 743-929 of the human phospholipase D1 (PLD1), and a cDNA from human liver (R93485) which encodes for a protein similar to region 815-932 of PLD1. Sequence comparison between cloned phospholipases showed the presence of 3 conserved amino acid sequences: AFVGGIDLAYGRWD (box A), IIGSANINDRS (box B), and YIYIENQFFI (box C). Phylogenic analysis indicated that the cDNA from brain and liver encoded for human isologs of PLD1.

ADP-Ribosylation Factors↗

PubMed1996

Cloning of a human homologue of the mouse Tctex-5 gene within the MHC class I region.

Using a positional cloning strategy to identify the hemochromatosis gene (HFE), we isolated seven cDNAs by cDNA selection from a region of 400 kilobases (kb) located near the HLA-A and HLA-F loci. In this paper, we report the study of one of the corresponding genes, referred to as HCG V (hemochromatosis candidate gene), localized 150 kb centromeric to HLA-A. This gene was found to be expressed ubiquitously in the form of a 1.8 kb transcript, and to be apparently well conserved during evolution. The gene spanned 3.1 kb and is organized in three exons and two introns. The cDNA of 1620 base pairs (bp) showed an open reading frame of 378 bp, encoding for a 126 amino acid polypeptide which displayed a strong identity with the predicted product of a mouse Tctex-5 gene (t complex, testis expressed) localized in the t complex on chromosome 17. The HCG V gene was assessed as a potential candidate for hemochromatosis in regard to its localization in the linkage disequilibrium area between HFE and polymorphic markers. The study of deletions and point mutations in hemochromatosis patients revealed a single bp polymorphism within the coding region; however, no associated disease changes were found. Therefore we conclude that HCG V is unlikely to be involved in the pathogenesis of hemochromatosis.

Amino Acid Sequence↗

PubMed1996

New highly polymorphic microsatellite marker in linkage disequilibrium with HLA-B.

The difficulty of molecular typing of the HLA class I genes and the relevance of the genes of this region to disease susceptibility and transplantation have provided an impetus to develop useful typing markers. We have characterized by polymerase chain reaction analysis a new highly informative CA repeat localized approximately 25-kb centromeric to the gene HLA-B and 10-kb telomeric to the gene MICA. Twelve alleles defined by length were found in a sample of French Basques, with the PIC being 0.82. A detailed haplotype analysis was performed to investigate the association between this microsatellite and two others markers of the region (HLA-B gene and TNF region microsatellite). The 10 haplotypes with the highest estimated frequencies show evidence of a gametic association or linkage disequilibrium. A very strong association between the expressed HLA-B polymorphism and microsatellite alleles was also revealed in this sample and confirmed in the workshop cells lines of the Fourth Asia-Oceania Histocompatibility Workshop. This marker can be used in the fine mapping of this region and the association with some alleles of HLA-B may allow the replacement of HLA-B typing at least in a preliminary study. Moreover, these studies support the hypothesis of a high mutability for large alleles in microsatellite loci.

Alleles↗

PubMed1995

A precise meiotic map in the class I region of the human major histocompatibility complex.

Human families in which recombinant meiotic event(s) are known to have occurred are powerful tools with which to analyze more precisely the structures of defined genomic regions, especially unstable areas. Such families allow the determination of the haplotypes of each member and, taking into account the recombinant event, it is possible to localize very precisely the point of crossover. Using families in which crossovers between the genes HLA-A and -B have occurred, we have constructed a meiotic map localizing the meiotic breakpoint events with respect to both anonymous markers and the principal genes of the region. Such mapping, which depends on the direct analysis of genomic DNA, is essential for fine structural analysis and is a powerful means of verification of the order and the localization of markers: physical mapping alone, using yeast artificial chromosomes, presents some uncertainties due to the numerous chimeras and inversions that can be produced. The establishment of this map will allow us to determine efficiently the precise location for new markers already localized to the map region. Three microsatellites (D6S265, D6S276, and D6S306), localized in the HLA region by linkage analysis, have been precisely located with respect to the points of recombination in the class I region. The sites of meiotic recombination in the MHC class I region seem to be not randomly distributed but in the majority of cases occurred between HLA-C and the microsatellite D6S265. This study also shows two cases of abnormal segregation of alleles. We discuss how these mutations correspond to a spontaneous mutation event at the somatic or germinal level.

Binding Sites↗

PubMed1995

The human myelin oligodendrocyte glycoprotein (MOG) gene: complete nucleotide sequence and structural characterization.

Human myelin oligodendrocyte glycoprotein (MOG), a myelin component of the central nervous system, is a candidate target antigen for autoimmune-mediated demyelination. We have isolated and sequenced part of a cosmid clone that contains the entire human MOG gene. The primary nuclear transcript, extending from the putative start of transcription to the site of poly(A) addition, is 15,561 nucleotides in length. The human MOG gene contains 8 exons, separated by 7 introns; canonical intron/exon boundary sites are observed at each junction. The introns vary in size from 242 to 6484 bp and contain numerous repetitive DNA elements, including 14 Alu sequences within 3 introns. Another Alu element is located in the 3'-untranslated region of the gene. Alu sequences were classified with respect to subfamily assignment. Seven hundred sixty-three nucleotides 5' of the transcription start and 1214 nucleotides 3' of the poly(A) addition sites were also sequenced. The 5'-flanking region revealed the presence of several consensus sequences that could be relevant in the transcription of the MOG gene, in particular binding sites in common with other myelin gene promoters. Two polymorphic intragenic dinucleotide (CA)n and tetranucleotide (TAAA)n repeats were identified and may provide genetic marker tools for association and linkage studies.

Base Sequence↗

PubMed1995

Localization of new genes and markers to the distal part of the human major histocompatibility complex (MHC) region and comparison with the mouse: new insights into the evolution of mammalian genomes.

We have refined and extended the map of the distal half of the human major histocompatibility complex. The map is continuous from HLA-E to 1000 kb telomeric of HLA-F and includes six new markers and genes. In addition, the corresponding sequences that were not previously mapped in the mouse genome have been located. The human and the mouse organizations have therefore been compared. This comparison allows us to demonstrate that the structure of the distal part of the MHC is similar in the two species. In addition, this comparison shows the presence of a breakpoint of synteny telomeric of the distal part of the H-2 region. Indeed, the region telomeric of HLA in human is found on a chromosome different from that carrying H-2 in mouse. The mapping analysis of paralogous genes (structurally related genes) around the breakpoint shows that the human organization probably represents the putative human/mouse ancestral one. This evolutionary breakpoint was precisely mapped in human, and the surrounding region was cloned into yeast artificial chromosomes. Finally, we show that the region found around the breakpoint was involved several times in chromosome recombinations in the mouse lineage, as it seems to correspond also to the t-complex distal inversion point.

Animals↗

PubMed1995

Myelin oligodendrocyte glycoprotein (MOG) gene polymorphisms and multiple sclerosis: no evidence of disease association with MOG.

The region surrounding the myelin oligodendrocyte glycoprotein (MOG) gene, located telomeric to the major histocompatibility complex on chromosome 6, was shown to contain three highly informative microsatellites. To examine the potential role of variants of the MOG gene in susceptibility to multiple sclerosis, these CA-repeat polymorphic markers were characterized on a sample of 169 multiple sclerosis patients and 173 healthy unrelated individuals by a method combining fluorescence labelling of PCR products and use of an automated DNA sequencer. Both patients and controls lived in the southwest of France (in the Pyrénées-Atlantiques) and had similar ethnic background. The distribution of the MOG haplotypes was not significantly different in the two groups (P = 0.38). This is not in favour of the implication of the MOG gene in the genetic component of multiple sclerosis, unless different independent mutations have occurred within this gene.

Base Sequence↗

PubMed1995

Three highly polymorphic microsatellites at the human myelin oligodendrocyte glycoprotein locus, 100 kb telomeric to HLA-F. Characterization and relation to HLA haplotypes.

The MOG locus, located on chromosomal bands 6p21.3-p22 and mapped about 100 kb telomeric to HLA-F, was isolated from cosmid ICRFc109A2434 and shown to contain three microsatellites. These CA-repeat polymorphic markers were characterized in a sample of 173 healthy unrelated individuals and 84 DNAs from the HLA Workshop reference panel, by a method combining fluorescence labeling of PCR products and use of an automated DNA sequencer. For the three markers, frequencies of heterozygotes are well predicted from allele frequencies by the Hardy-Weinberg rule, which suggests that problems of allele nonamplification are unlikely. Typing of cell lines homozygous in the HLA region allowed unambiguous definition of 81 HLA-MOG haplotypes and showed that several HLA ancestral haplotypes extended to the MOG region. The high degree of polymorphism (59%, 51%, and 81% at the three loci, respectively, and 87% at the haplotype level) makes these new markers informative for association or linkage studies with diseases such as hemochromatosis or multiple sclerosis, and for studies aimed at precisely delineating the site of crossover in chromosomes in which recombination occurred in the distal part of the HLA class I region.

B-Lymphocytes↗

PubMed1994

Localization of the OTF3 gene within the human MHC class I region by physical and meiotic mapping.

OTF3 (octamer transcription factor 3) is a transcription factor containing a POU-specific domain and a homeodomain that could play a role in early development. In situ hybridization and pairwise linkage analysis showed that OTF3 gene maps close to the human MHC (major histocompatibility complex). In this paper, we define its localization within the MHC, around 100 kb telomeric to HLA-C, using a combination of physical and genetic analyses.

Alleles↗

PubMed1994

Non-homologous recombination within the major histocompatibility complex creates a transcribed hybrid sequence.

The P5-1 cDNA clone maps to the human MHC class I region (Vernet et al. 1993a). In this paper, we show that the P5-1 cDNA represents a chimeric transcript in which the first exon of an MHC class I gene has been spliced to an unrelated sequence. The corresponding gene P5-1 is composed of the 5' sequence of an MHC class I gene including the promoter region, the first exon, and the half of the first intron fused to an unrelated intron, followed by a large exon. Furthermore, the non-class I part of P5-1 is present within the MHC class I region in multiple copies, defining the P5 family. Another member of the P5 family is fused to a class I gene, although by a type of rearrangement different from P5-1. These two fusion events between members of HLA class I and P5 families reflect the existence of a duplication unit including two class I genes and a P5 sequence. These data shed light on the MHC class I evolution and on the creation and evolution of new genes.

Base Sequence↗

PubMed1994

Structure and evolution of a member of a new subfamily of GTP-binding proteins mapping to the human MHC class I region.

A gene coding for a putative GTP-binding protein, MMR1, has been localized on band C of the murine Chr 17 within or close to the MHC (Denizot et al. 1992). Its human homolog, HSR1, localized to the human MHC class I region, is described in this paper. Its sequence, compared with MMR1, shows that the conceptual proteins encoded by these genes are highly homologous and have thus been subjected to high constraints during evolution. Furthermore, a detailed databank search with HSR1 leads to the characterization of a new subfamily of GTP-binding proteins, of which HSR1 and MMR1 are the only eukaryotic members. The precise localization of HSR1 within the human MHC class I region is also presented.

Amino Acid Sequence↗

PubMed1994

Anonymous marker loci within 400 kb of HLA-A generate haplotypes in linkage disequilibrium with the hemochromatosis gene (HFE)

The hemochromatosis gene (HFE) maps to 6p21.3 and is less than 1 cM from the HLA class I genes; however, the precise physical location of the gene has remained elusive and controversial. The unambiguous identification of a crossover event within hemochromatosis families is very difficult; it is particularly hampered by the variability of the phenotypic expression as well as by the sex- and age-related penetrance of the disease. For these practical considerations, traditional linkage analysis could prove of limited value in further refining the extrapolated physical position of HFE. We therefore embarked upon a linkage-disequilibrium analysis of HFE and normal chromosomes from the Brittany population. In the present report, 66 hemochromatosis families yielding 151 hemochromatosis chromosomes and 182 normal chromosomes were RFLP-typed with a battery of probes, including two newly derived polymorphic markers from the 6.7 and HLA-F loci located 150 and 250 kb telomeric to HLA-A, respectively. The results suggest a strong peak of existing linkage disequilibrium focused within the i82-to-6.7 interval (approximately 250 kb). The zone of linkage disequilibrium is flanked by the i97 locus, positioned 30 kb proximal to i82, and the HLA-F gene, found 250 kb distal to HLA-A, markers of which display no significant association with HFE. These data support the possibility that HFE resides within the 400-kb expanse of DNA between i97 and HLA-F. Alternatively, the very tight association of HLA-A3 and allele 1 of the 6.7 locus, both of which are comprised by the major ancestral or founder HFE haplotype in Brittany, supports the possibility that the disease gene may reside immediately telomeric to the 6.7 locus within the linkage-disequilibrium zone. Additionally, hemochromatosis haplotypes possessing HLA-A11 and the low-frequency HLA-F polymorphism (allele 2) are supportive of a separate founder chromosome containing a second, independently arising mutant allele. Overall, the establishment of a likely "hemochromatosis critical region" centromeric boundary and the identification of a linkage-disequilibrium zone both significantly contribute to a reduction in the amount of DNA required to be searched for novel coding sequences constituting the HFE defect.

Alleles↗

PubMed1993

Myelin/oligodendrocyte glycoprotein is a member of a subset of the immunoglobulin superfamily encoded within the major histocompatibility complex.

Myelin/oligodendrocyte glycoprotein (MOG) is found on the surface of myelinating oligodendrocytes and external lamellae of myelin sheaths in the central nervous system, and it is a target antigen in experimental autoimmune encephalomyelitis and multiple sclerosis. We have isolated bovine, mouse, and rat MOG cDNA clones and shown that the developmental pattern of MOG expression in the rat central nervous system coincides with the late stages of myelination. The amino-terminal, extracellular domain of MOG has characteristics of an immunoglobulin variable domain and is 46% and 41% identical with the amino terminus of bovine butyrophilin (expressed in the lactating mammary gland) and B-G antigens of the chicken major histocompatibility complex (MHC), respectively; these proteins thus form a subset of the immunoglobulin superfamily. The homology between MOG and B-G extends beyond their structure and genetic mapping to their ability to induce strong antibody responses and has implications for the role of MOG in pathological, autoimmune conditions. We colocalized the MOG and BT genes to the human MHC on chromosome 6p21.3-p22. The mouse MOG gene was mapped to the homologous band C of chromosome 17, within the M region of the mouse MHC.

Aging↗

PubMed1993

Evolutionary study of multigenic families mapping close to the human MHC class I region.

During a search for novel coding sequences within the human MHC class I region (chromosome 6p21.3), we found an exon (named B30-2) coding for a 166-amino-acid peptide which is very similar to the C-terminal domain of several coding sequences: human 52-kD Sjögren's syndrome nuclear antigen A/Ro (SS-A/Ro) and ret finger protein (RFP), Xenopus nuclear factor 7 (XNF7), and bovine butyrophilin. The first three of these proteins share similarities over the whole length of the molecule whereas butyrophilin is similar in the C-terminal domain. The N-terminal domain of butyrophilin is similar to rat myelin/oligodendrocyte glycoprotein (MOG) and chicken B blood group system (B-G) protein. These domains are components of a new subfamily of the immunoglobulin superfamily (IgSF). Butyrophilin is thus a mosaic protein composed of the MOG/B-G Ig-like domain and the C-terminal domain of 52-kD SS-A/Ro, RFP, and XNF7 (B30-2-like domain). Moreover, in situ hybridization shows that RFP, butyrophilin, and MOG map to the human chromosome 6p21.3-6p22 region and are thus close to the MHC class I genes. It is therefore possible that the butyrophilin gene is the product of an exon shuffling event which occurred between ancestors of the RFP and MOG genes. To our knowledge, this is the first example of the colocalization of a chimeric gene and its putative progenitors. Finally, regulatory protein T-lymphocyte 1 (Rpt-1) shares similarities with the N-terminal halves of RFP, 52-kD SS-A/Ro, and XNF7, but not with the B30-2-like domain. We show that the ancestral Rpt-1 gene evolved by overprinting.

Amino Acid Sequence↗

PubMed1993

HLA-E is the only class I gene that escapes CpG methylation and is transcriptionally active in the trophoblast-derived human cell line JAR.

Polymorphic as well as HLA-F and -G genes are repressed in the human cell line JAR, derived from a tumor of trophoblast origin. By contrast, the HLA-E gene as well as the non-HLA novel coding-sequence, R1, located 5' to HLA-E, both remain transcriptionally active. We first demonstrated the role of DNA methylation in the repression of class I genes (except HLA-E) in JAR by the use of the 5-Azacytidine demethylating agent. Following treatment, JAR clones reexpressed polymorphic class I transcripts and cell surface alpha chains. Using methylation-sensitive rare cutter enzymes on JAR genomic DNA, followed by classical or pulse field gel electrophoresis and hybridization with HLA locus-specific probes, we found methylated CpG islands in the 5' region of all class I genes, except for HLA-E. These results, establishing an inverse relationship between states of methylation and transcriptional activity within the MHC class I chromosomal region in JAR, and the observations that the HLA-E and R1 genes were ubiquitously expressed, suggest that the HLA-E chromosomal domain might have functional importance including the presence of housekeeping genes.

Azacitidine↗

PubMed1993

A novel coding sequence belonging to a new multicopy gene family mapping within the human MHC class I region.

The human major histocompatibility complex (MHC) region is a genomic region spanning about 4000 kilobases (kb) including the class I, class II, and class III subregions. The class I subregion is larger than the two others but with fewer genes described to date. It includes a) classical human leucocyte antigen (HLA) class I genes (HLA-A, HLA-B, HLA-C) which are highly polymorphic and encode products presenting the endogenous antigenic peptides to the T-cell receptors, and b) non-classical class I genes (HLA-E, HLA-F, HLA-G) whose function is still unknown. In this study, we describe the first coding sequence which is not structurally related to the class I genes, although it is localized within the MHC class I region. This novel gene, P5-1, belongs to a multiple copy family, all members of which map within the MHC. Although the P5-1 sequence showed no similarity to sequences in different databanks, its transcription, which is restricted to lymphoid tissues, argues for an immunological function of its product.

Amino Acid Sequence↗

PubMed1993

Analysis of HLA-A/B recombinant families with new polymorphic markers.

The MHC in humans is a much studied region of the genome, the genes of which display a high rate of polymorphism and a high rate of linkage disequilibrium. Four families in which intra-class-I recombination has occurred have been analyzed with six polymorphic markers between HLA-A and -B in order to determine the full haplotypes of the whole families and to localize the points of crossover. The previously proposed order of the markers was confirmed by recombination mapping. In one family, the crossover was shown to have occurred in the 20-kb stretch of DNA bounded by the two markers (P3B and P5) between which no evidence of linkage disequilibrium was found, a region which constitutes of only about 1% of the distance between HLA-A and HLA-B. Although supportive of the suggestion of a hot spot of recombination in this region, based on the apparent lack of linkage disequilibrium between the markers P3B and P5, more such families need to be tested in order to confirm or refute this hypothesis.

Crossing Over, Genetic↗

PubMed1993

Differences between human sperm and somatic cell DNA in CpG methylation within the HLA class I chromosomal region.

PROBLEM: We investigated the possible negative regulatory mechanisms that repress classical human leukocyte antigen (HLA) class I gene expression in human spermatozoa and searched for novel testis-specific coding sequences that might be present in MHC class I chromosomal region. METHOD: We performed a comparative DNA methylation analysis of this genomic region in both purified human spermatozoa and mononuclear blood cells from the same donors, using methylation-sensitive restriction enzymes followed by classical or pulsed field gel electrophoresis and hybridization with HLA class I locus-specific probes. RESULTS: Unmethylated CpG sites were detected in the 3' part of HpaII tiny fragments of the HLA-F and HLA-G genes in spermatozoal DNA. In contrast, no difference was observed in the methylation status of the HLA-B, HLA-C, and HLA-E genes between germ and somatic cells. CpG unmethylation events were also detected in several parts of this chromosomal region (outside the known loci) in spermatozoal DNA. CONCLUSIONS: These results suggest that this genomic region undergoes changes in its DNA methylation pattern during the developmental process. We hypothesize that these dynamic changes have functional importance, including a possible transcriptional activity of nonclassical class I genes and/or as yet undescribed testis-specific coding sequences.

Base Sequence↗