The critical region for Angelman syndrome lies between D15S122 and D15S113.
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Angelman Syndrome↗
Biomedical subjects
M Lalande
Publications and source records attributed to M Lalande.
Inhibition of the G1-S transition of the cell cycle by inhibitors of deoxyhypusine hydroxylation.
The formation of the unusual amino-acid hypusine in eIF-5A (eukaryotic initiation factor 5A) is associated with cellular proliferation. We used a panel of compounds, including mimosine, to probe the relationship between the exit from the G1 phase of the cell cycle, i.e., the onset of DNA replication, and the formation of hypusine by the enzyme deoxyhypusyl hydroxylase (DOHH). These two parameters displayed the same dose dependency and structure-activity relationship. Only compounds that inhibited DOHH also suppressed proliferation. This effect was observed: (i) in spontaneously proliferating, virally transformed, and mitogen-stimulated cells; (ii) for both anchorage-dependent and anchorage-independent proliferation; and (iii) with normal and malignant cell lines. DOHH reactivation occurred rapidly after inhibitor withdrawal and correlated with synchronized entry into S. The changes in the expression of specific genes during the G1-to-S transition mimicked the physiological pattern. These findings suggest that hypusine formation in eIF-5A which occurs in a specific, invariant sequence motif acquired early in evolution, may be involved in the G1-to-S transition in the eukaryotic cells tested.
Alanine↗
The human gamma-aminobutyric acid receptor subunit beta 3 and alpha 5 gene cluster in chromosome 15q11-q13 is rich in highly polymorphic (CA)n repeats.
The gamma-aminobutyric acid (GABAA) receptor beta 3 (GABRB3) and alpha 5 (GABRA5) subunit genes have been localized to the Angelman and Prader-Willi syndrome region of chromosome 15q11-q13. GABRB3, which encompasses 250 kb, is located 100 kb proximal of GABRA5, with the two genes arranged in head-to-head transcriptional orientation. In screening 135 kb of cloned DNA within a 260-kb interval extending from within GABRB3 to the 5' end of GABRA5, 10 new (CA)n repeats have been identified. Five of these have been analyzed in detail and found to be highly polymorphic, with the polymorphism information content (PIC) ranging from 0.7 to 0.85 and with heterozygosities of 67 to 94%. In the clones from GABRB3/GABRA5 region, therefore, the frequency of (CA)n with PICs > or = 0.7 is 1 per 27 kb. Previous estimates of the density of (CA)n with PICs > or = 0.7 in the human genome have been approximately 10-fold lower. The GABRB3/GABRA5 region appears, therefore, to be enriched for highly informative (CA)n. This set of closely spaced, short tandem repeat polymorphisms will be useful in the molecular analyses of Prader-Willi and Angelman syndromes and in high-resolution studies of genetic recombination within this region.
Angelman Syndrome↗
Allele specificity of DNA replication timing in the Angelman/Prader-Willi syndrome imprinted chromosomal region.
DNA replication within chromosome 15q11-q13, a region subject to genomic imprinting, was examined by fluorescence in situ hybridization. Asynchronous replication between homologues was observed in cells from normal individuals and in Prader-Willi (PWS) and Angelman syndrome (AS) patients with chromosome 15 deletions but not in PWS patients with maternal uniparental disomy. Opposite patterns of allele-specific replication timing between homologous loci were observed; paternal early/maternal late at D15S63, D15S10 and the gamma-aminobutyric acid receptor beta 3 subunit gene (GABRB3); and maternal early/paternal late at the more distal gamma-aminobutyric acid receptor alpha 5 subunit gene (GABRA5). At the most distal locus examined, D15S12, both patterns of allele-specific replication timing were detected.
Alleles↗
In and around SNRPN.
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Angelman Syndrome↗
Imprinting mutations suggested by abnormal DNA methylation patterns in familial Angelman and Prader-Willi syndromes.
The D15S9 and D15S63 loci in the Prader-Willi/Angelman syndrome region on chromosome 15 are subject to parent-of-origin-specific DNA methylation. We have found two Prader-Willi syndrome families in which the patients carry a maternal methylation imprint on the paternal chromosome. In one of these families, the patients have a small deletion encompassing the gene for the small nuclear ribonucleoprotein polypeptide N, which maps 130 kb telomeric to D15S63. Furthermore, we have identified a pair of nondeletion Angelman syndrome sibs and two isolated Angelman syndrome patients who carry a paternal methylation imprint on the maternal chromosome. These Angelman and Prader-Willi syndrome patients may have a defect in the imprinting process in 15q11-13. We propose a model in which a cis-acting mutation prevents the resetting of the imprinting signal in the germ line and thus disturbs the expression of imprinted genes in this region.
Angelman Syndrome↗
A cluster of three GABAA receptor subunit genes is deleted in a neurological mutant of the mouse p locus.
The mouse pink-eyed cleft-palate (p(cp)) mutation is characterized by hypopigmentation associated with cleft palate, neurological disorders and runting. Most p(cp) homozygotes are born with cleft palate and die shortly after birth, presumably as a result of feeding problems. A few exceptional p(cp) mutants live beyond this stage but display tremor and jerky gait. We report here that the genes encoding the gamma-aminobutyric acid type A (GABAA) receptor subunits alpha 5 (originally described as alpha 4; ref. 4), beta 3 and gamma 3 are disrupted by a deletion in p(cp) mice. We also show that the alpha 5 and gamma 3 genes are located between the p and beta 3 genes on mouse chromosome 7. The p(cp) deletion leads to alterations of binding properties of the GABAA receptors in the brain, providing an in vivo model system for studying GABAA receptor function. The human homologue of the region deleted in p(cp) mice is associated with Angelman syndrome. Thus, p(cp) mice may be useful in defining the region containing the gene(s) for this syndrome.
Animals↗
Determination of DNA replication kinetics in synchronized human cells using a PCR-based assay.
Studies on the temporal order of DNA replication are difficult due to the lack of sensitivity of methods available for replication kinetic analysis. To overcome problems associated with the current techniques, we propose a PCR-based assay to determine the replication time of any single-copy DNA sequence in complex genomes. Human cells labeled with 5-bromodeoxyuridine (BrdU) were flow sorted, according to their DNA content, at different times after synchronous release from the G1/S phase boundary. The selective removal of newly-replicated BrdU-substituted DNA was achieved by UV light irradiation followed by S1 nuclease treatment. The timing of replication of selected DNA sequences (housekeeping, tissue-specific, and non-coding loci) was determined by polymerase chain reaction (PCR) amplification using appropriate primers. DNA sequences localized in inactive replication units allowed amplification whereas those that have replicated will not be amplified by PCR. Using this sensitive and quantitative assay the replication kinetic analysis of a number of different DNA sequences can be performed from a single sorting experiment.
Bromodeoxyuridine↗
High-performance liquid chromatographic method for the determination of indapamide in human whole blood.
A sensitive, accurate, and reproducible high-performance liquid chromatographic procedure for the analysis of indapamide in human whole blood is reported. After a single-step liquid-liquid extraction at pH 6.6 using diethyl ether, indapamide was eluted from a Nucleosil C18 5-microns column with 80 mM ammonium acetate, pH 3.5-acetonitrile-2-propanol (65:30:5, v/v/v). The peak height versus whole blood concentration was linear over the range 10.0-500 ng/ml using ultraviolet detection. The mean absolute recovery of indapamide using the described assay was 87.4%. The inter- and intra-day accuracy and precision were within 9.6% of the actual values for all concentrations investigated. Furthermore, this procedure was applied to the analysis of whole blood samples from healthy subjects receiving a single 2.5-mg oral dose of indapamide.
Chromatography, High Pressure Liquid↗
Cytogenetic and molecular studies in the Prader-Willi and Angelman syndromes: an overview.
The majority of patients with Angelman syndrome and Prader-Willi syndrome have a cytogenetic and molecular deletion of chromosome 15q11q13 with the primary difference being in the parental origin of deletion. Our current understanding of the cytogenetics and molecular genetics of these 2 clinically distinct syndromes will be discussed in this review.
Angelman Syndrome↗
Molecular definition of the Prader-Willi syndrome chromosome region and orientation of the SNRPN gene.
The Prader-Willi syndrome and the Angelman syndrome are caused by the loss of function of distinct but closely linked genes on human chromosome 15. Based on a yeast artificial chromosome restriction map and two key patients we have determined that the shortest region of deletion overlap in the Prader-Willi syndrome comprises 320 kb. The region includes the anonymous DNA marker PW71 (D15S63) and the gene for the small nuclear ribonucleoprotein N (SNRPN). The SNRPN gene maps 130 kb distal to PW71 and is transcribed from centromere to telomere.
Autoantigens↗
FISH ordering of reference markers and of the gene for the alpha 5 subunit of the gamma-aminobutyric acid receptor (GABRA5) within the Angelman and Prader-Willi syndrome chromosomal regions.
We have established a probe order within the Angelman/Prader-Willi chromosomal regions by multicolor fluorescence in situ hybridization (FISH). The probe [locus] order extending distally from the centromere is 34[D15S9]-IR4-3R[D15S11]-189-1[D15S13]-PW71++ + [D15S63]-3-21[D15S10]-28 beta 3-H3[GABRB3]-IR10-1 [D15S12]. This order agrees with that recently reported (1) with the exception of PW71 [D15S63]. In addition, a second gamma-aminobutyric acid (GABAA) receptor, the alpha 5 subunit, has been localized within the reference map to between GABRB3 and D15S12. The locus order was further confirmed by DNA hybridization analysis of two patients, one with Angelman syndrome and one with Prader-Willi syndrome, with different unbalanced translocations and molecular extents of deletion. Our results provide a framework map of chromosome 15q11-q13 into which additional markers can be oriented and allow a further differentiation of the critical genetic regions of the two syndromes.
Amino Acid Sequence↗
Cloning of the breakpoints of a submicroscopic deletion in an Angelman syndrome patient.
The majority of cases of the two distinct disorders Prader-Willi syndrome (PWS) and Angelman syndrome (AS) result from cytogenetic deletions of chromosome 15q11-q13. These deletions are exclusively of maternal origin in AS but of paternal origin in PWS indicating that the 15q11-q13 region is subject to genomic imprinting. Transmission of a submicroscopic deletion in one three generation family resulted in AS only upon maternal transmission of the deletion with no clinical phenotype associated with paternal transmission (1,2). The breakpoint of this submicroscopic deletion has been cloned and sequenced. This is the first deletion junction from the AS/PWS region which has been so characterized. The nucleotide sequence of the deletion junction revealed a 19 bp insertion of unknown origin with no evidence of repetitive elements. A probe from the proximal deletion breakpoint, PB11, lies within the currently defined minimum region of deletion overlap in PWS, which contains the SNRPN and D15S63 loci. Our results suggest that the imprinted gene(s) responsible for the PWS phenotype are proximal of pB11 in this deletion overlap region.
Angelman Syndrome↗
Isolation of new probes in the region of the Wilson disease locus, 13q14.2-->q14.3.
A hybrid panel was used to refine the localizations of eight established markers useful for the diagnosis of Wilson disease. Two of the markers, D13S59 and D13S31, that map very close to the Wilson disease locus (WND) were localized to the region 13q14.2-->q14.3. We report the isolation of seven new probes from this region, using two different approaches. First, 16 clones from a chromosome 13-specific library were mapped using the hybrid panel. Three of the clones mapped to 13q14.2-->q14.3. As a second approach, Alu element-mediated PCR (Alu-PCR) was used to generate clones from a hybrid (ICD) that contains the proximal half of chromosome 13 as the only human component. To select for those that potentially mapped within the region 13q14.2-->q14.3, the clones were screened by differential hybridization using the labeled Alu-PCR products from a hybrid (KSF39) that is similar to ICD but has a deletion in the region 13q14.2-->q14.3. The procedure was successful even though KSF39 contains five additional human chromosomes. Six independent clones were selected. Five of these were found to be nonrepetitive, and four were found to map correctly to 13q14.2-->q14.3 when localized using the hybrid panel.
Chromosomes, Human, Pair 13↗
Mapping of the Angelman and Prader-Willi syndromes.
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Angelman Syndrome↗
Linkage analysis in familial Angelman syndrome.
Familial Angelman syndrome (AS) can result from mutations in chromosome 15q11q13 that, when transmitted from father to child, result in no phenotypic abnormality but, when transmitted from mother to child, cause AS. These mutations therefore behave neither as dominant nor as recessive mutations but, rather, show an imprinted mode of inheritance. We have analyzed two sibling pairs with AS and a larger family with four AS offspring of three sisters with several recently described microsatellite polymorphisms in the AS region. AS siblings inherited the same maternal alleles at the GABRB3 and GABRA5 loci, and the unaffected siblings of AS individuals inherited the other maternal alleles at these loci. In one of the AS sibling pairs, analysis of a recombination event indicates that the mutation responsible for AS is distal to locus D15S63. This result is consistent with a previously described imprinted submicroscopic deletion causing AS, a deletion that includes loci D15S10, D15S113, and GABRB3, all distal to D15S63. The analysis of the larger AS family provides the first clear demonstration of a new mutation in nondeletion AS. Analysis of linkage of AS to GABRB3 in these three families, on the assumption of imprinted inheritance (i.e., penetrance of an AS mutation is 1 if transmitted maternally and is 0 if transmitted paternally), indicates a maximum lod score of 3.52 at theta = 0.
Angelman Syndrome↗
High-resolution mapping of the gamma-aminobutyric acid receptor subunit beta 3 and alpha 5 gene cluster on chromosome 15q11-q13, and localization of breakpoints in two Angelman syndrome patients.
The gamma-aminobutyric acid (GABAA) receptors are a family of ligand-gated chloride channels constituting the major inhibitory neurotransmitter receptors in the nervous system. In order to determine the genomic organization of the GABAA receptor beta 3 subunit gene (GABRB3) and alpha 5 subunit gene (GABRA5) in chromosome 15q11-q13, we have constructed a high-resolution physical map using the combined techniques of field-inversion gel electrophoresis and phage genomic library screening. This map, which covers nearly 1.0 Mb, shows that GABRB3 and GABRA5 are separated by less than 100 kb and are arranged in a head-to-head configuration. GABRB3 encompasses approximately 250 kb, while GABRA5 is contained within 70 kb. This difference in size is due in large part to an intron of 150 kb within GABRB3. We have also identified seven putative CpG islands within a 600-kb interval. Chromosomal rearrangement breakpoints--in one Angelman syndrome (AS) patient with an unbalanced translocation and in another patient with a submicroscopic deletion--are located within the large GABRB3 intron. These findings will facilitate chromosomal walking strategies for cloning the regions disrupted by the DNA rearrangements in these AS patients and will be valuable for mapping new genes to the AS chromosomal region.
Angelman Syndrome↗
A duplicated region is responsible for the poly(ADP-ribose) polymerase polymorphism, on chromosome 13, associated with a predisposition to cancer.
The poly(ADP-ribose) polymerase (PADPRP) gene (13q33-qter) depicts a two-allele (A/B) polymorphism. In the noncancer population, the frequency of the B allele is higher among blacks than among whites. Since the incidence of multiple myeloma and prostate and lung cancer is higher in the U.S. black population, we have analyzed the B-allele frequency in germ-line DNA to determine whether the PADPRP gene correlates with a polymorphic susceptibility to these diseases. For multiple myeloma and prostate cancer, an increased frequency of the B allele appeared to be striking only in black patients. In contrast, the distribution of the B allele in germ-line DNA did not differ among white patients with these diseases, when compared with the control group. An elevated B-allele frequency was also found in germ-line DNA in blacks with colon cancer. These observations suggest that the PADPRP polymorphism may provide a valid marker for a predisposition to these cancers in black individuals. To determine the genomic structure of the polymorphic PADPRP sequences, a 2.68-kb HindIII clone was isolated and sequenced from a chromosome 13-enriched library. Sequence analysis of this clone (A allele) revealed a close sequence similarity (91.8%) to PADPRP cDNA (1q42) and an absence of introns, suggesting that the gene on 13q exists as a processed pseudogene. A 193-bp conserved duplicated region within the A allele was identified as the source of the polymorphism. The nucleotide differences between the PADPRP gene on chromosome 13 and related PADPRP genes were exploited to develop oligonucleotides that can detect the difference between the A/B genotypes in a PCR. This PCR assay offers the opportunity for analyzing additional black cancer patients, to determine how the PADPRP processed pseudogene or an unidentified gene that cosegregates with the PADPRP gene might be involved with the development of malignancy.
Alleles↗