Linkage analysis with chromosome 15q11-13 markers shows genomic imprinting in familial Angelman syndrome.

Angelman syndrome (AS) and Prader-Willi syndrome (PWS) have become the classical examples of genomic imprinting in man, as completely different phenotypes are generated by the absence of maternal (AS) or paternal (PWS) contributions to the q11-13 region of chromosome 15 as a result of deletion or uniparental disomy. Apparently, most patients are sporadic cases. The genetic mechanism underlying familial AS has remained enigmatic for a long time. Recently, evidence has been emerging suggesting autosomal dominant inheritance of a detectable or undetectable defect in a gene or genes at 15q11-13, subject to genomic imprinting. The present report describes an unusually large pedigree with segregation of AS through maternal inheritance and apparent asymptomatic transmission through several male ancestors. Deletion and paternal disomy at 15q11-13 were excluded. However, the genetic defect is still located in this region, as we obtained a maximum lod score of 5.40 for linkage to the GABA receptor locus GABRB3 and the anonymous DNA marker D15S10, which have been mapped within or adjacent to the AS critical region at 15q11-13. The size of the pedigree allowed calculation of an odds ratio in favour of genomic imprinting of 9.25 x 10(5). This family illustrates the necessity of extensive pedigree analysis when considering recurrence risks for relatives of AS patients, those without detectable deletion or disomy in particular.     

Further evidence for dominant inheritance at the chromosome 15q11-13 locus in familial Angelman syndrome.

Eleven patients with Angelman syndrome (AS) and their parents from 5 families have been studied with high resolution chromosome analysis and molecular probes from region 15q11-13 in an attempt to elucidate the mode of inheritance in familial AS. No deletions were detected. All families were informative with a combination of different short arm cytogenetic markers. All sets of sibs inherited the same maternal chromosome 15, but in 3 families sibs inherited different paternal 15s. Analysis of 6 polymorphic DNA markers supported the conclusion that AS sibs inherit the same maternal 15, but often different paternal 15s. These data make autosomal recessive inheritance at a 15q11-13 locus very unlikely and support the hypothesis that familial AS is due to maternal transmission of a mutation within 15q11-13.     

Angelman syndrome with a chromosomal inversion 15 inv(p11q13) accompanied by a deletion in 15q11q13.

A family is described in which an inversion of chromosome 15, 15 inv(p11q13), is segregating. All family members are healthy except the proband who is a 10 year old boy with Angelman syndrome. Although the chromosomal inversion has been passed from the grandfather to both his son and his daughter with no ill effect, passage from daughter to grandson has resulted in a deletion of chromosome 15 material which is presumed to be the cause of Angelman syndrome in this boy. The probabilities of an inversion of this type being instrumental in causing the syndrome are discussed.     

A boy with developmental delay and a maternally inherited deletion in 15q11q13.

A boy was referred at 8 weeks of age for failure to thrive. Cytogenetic and molecular studies showed that he had a large proximal deletion of the maternally derived chromosome 15q. He did not have Angelman syndrome, but at 2 years of age was severely globally delayed. He died at 2 1/2 years of age.     

Supernumerary inv dup(15) in a patient with Angelman syndrome and a deletion of 15q11–q13

We have studied a patient with Angelman syndrome (AS) and a 47,XY,+inv dup(15) (pter→q11::q11→pter) karyotype. Molecular cytogenetic studies demonstrated that one of the apparently normal 15s was deleted at loci D15S9, GABRB3, and D15S12. There were no additional copies of these loci on the inv dup (15). The inv dup (15) contained only the pericentromeric sequence D15Z1. Quantitative DNA analysis confirmed these findings and documented a standard large deletion of sequences from 15q11-q13, as usually seen in patients with AS. DNA methylation testing at D15S63 showed a deletion of the maternally derived chromosome. AS in this patient can be explained by the absence of DNA sequences from chromosome 15q11-q13 on one of the apparently cytogenetically normal 15s, and not by the presence of an inv dup (15). This is the fourth patient with an inv dup (15) and AS or Prader Willi syndrome, who has been studied at the molecular level. In all cases an additional alteration of chromosome 15 was identified, which was hypothesized to be the cause of the disease. Patients with inv dup (15)s may be at increased risk for other chromosome abnormalities involving 15q11-q13. © 1995 Wiley-Liss, Inc.     

Angelman syndrome associated with a maternal 15q11-13 deletion of less than 200 kb

Angelman syndrome (AS) is a neurogenetic disorder arising froma lack of genetic contribution from the maternal chromosome15q11–13. To date, the AS critical region has been definedby an inherited deletion of approximately 1.5Mb, spanning the3–21 (D15S10), LS6–1 (D15S113) and GABRB3 loci.We have Identified an individual with the typical features ofAS who has a deletion of the maternal chromosome which encompassesLS6–1, but does not extend to either flanking marker.This deletion, initially detected by (CA)n repeat analysis,was further characterised by fluorescence In situ hybridisation(FISH) using cosmids derived from a 260 kb LS6–1 yeastartificial chromosome (YAC). Neither end cosmid from this YACclone falls within the deletion, suggesting that the minimalAS region Is less than 200 kb. We also studied three loci within15q11–13 which detect parent-of-origin specific DNA methylationimprints, and found that both normal maternal and paternal patternswere present in this patient.     

Large genomic duplicons map to sites of instability in the Prader-Willi/Angelman syndrome chromosome region (15q11-q13).

The most common etiology for Prader-Willi syndrome and Angelman syndrome is de novo interstitial deletion of chromosome 15q11-q13. Deletions and other recurrent rearrangements of this region involve four common 'hotspots' for breakage, termed breakpoints 1-4 (BP1-BP4). Construction of an approximately 4 Mb YAC contig of this region identified multiple sequence tagged sites (STSs) present at both BP2 and BP3, suggestive of a genomic duplication event. Interphase FISH studies demonstrated three to five copies on 15q11-q13, one copy on 16p11.1-p11.2 and one copy on 15q24 in normal controls, while analysis on two Class I deletion patients showed loss of approximately three signals at 15q11-q13 on one homolog. Multiple FISH signals were also observed at regions orthologous to both human chromosomes 15 and 16 in non-human primates, including Old World monkeys, suggesting that duplication of this region may have occurred approximately 20 million years ago. A BAC/PAC contig for the duplicated genomic segment (duplicon) demonstrated a size of approximately 400 kb. Surprisingly, the duplicon was found to contain at least seven different expressed sequence tags representing multiple genes/pseudogenes. Sequence comparison of STSs amplified from YAC clones uniquely mapped to BP2 or BP3 showed two different copies of the duplicon within BP3, while BP2 comprised a single copy. The orientation of BP2 and BP3 are inverted relative to each other, whereas the two copies within BP3 are in tandem. The presence of large duplicated segments on chromosome 15q11-q13 provides a mechanism for homologous unequal recombination events that may mediate the frequent rearrangements observed for this chromosome.     

Triplication of 15q11-q13 with inv dup(15) in a female with developmental delay.

A 4 year old female referred with developmental delay was found to have two de novo abnormal derivatives of chromosome 15, a supernumerary inverted duplicated marker chromosome (inv dup(15)) and an interstitial triplication of proximal 15q11-q13 or 14 in one of the two 15 homologues (trip(15)). Fluorescence in situ hybridisation (FISH) using probes within and flanking the Prader-Willi/Angelman syndrome critical region (PWASCR) confirmed the triplication in the abnormal 15 homologue. The inv dup(15) was dicentric, positive for IR39d which maps proximal to the PWASCR, but was negative for all the PWASCR FISH probes used. Results using polymorphic microsatellite repeats confirmed that the additional material in the trip(15) was maternal in origin and included several PWASCR loci. The presence of two de novo abnormalities involving the proximal region of 15q suggests a linked mechanism of origin.     

Molecular dissection of the Prader-Willi/Angelman syndrome region (15q11-13) by YAC cloning and FISH analysis.

Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are distinct mental retardation disorders associated with deletions of proximal 15q (q11-q13) of different parental origin. Yeast artificial chromosome (YAC) clones were isolated for 9 previously mapped DNA probes from this region, and for one newly derived marker, LS6-1 (D15S113). A YAC contig of 1-1.5 Mb encompassing four markers (ML34, IR4-3R, PW71, and TD189-1) was constructed. Multi-color fluorescence in situ hybridization (FISH) analysis of interphase nuclei was combined with YAC contig information to provide the following order of markers: cen-IR39-ML34-IR4-3R-PW71-TD189-1-LS6++ +-1-TD3-21-GABRB3-IR10-1-CMW1-tel. FISH analysis was performed on 8 cases of PWS and 3 cases of AS, including 5 patients with normal karyotypes. All eleven patients were deleted for YACs in the interval from IR4-3R to GABRB3. On the proximal side of the deletion interval, 10/10 breakpoints fell within a single ML34 YAC of 370 kb. On the distal side, 8/9 breakpoints fell within a single IR10-1 YAC of 200 kb. These results indicate a striking consistency in the location of the proximal and distal breakpoints in PWS and AS patients. FISH analysis on a previously reported case of familial AS confirmed a submicroscopic deletion including YACs corresponding to LS6-1, TD3-21 and GABRB3 and supports the separation of the PWS and AS critical regions. Since these three YACs do not overlap each other, the minimum size of the AS critical region is > or = 650 kb.     

Maternal origin of 15q11-13 deletions in Angelman syndrome suggests a role for genomic imprinting

Six persons with the classical Angelman syndrome (AS) phenotype and de novo deletions of chromosome 15q11-q13 were studied to determine the parental origin of the chromosome deletion. Four of the 6 patients had informative cytogenetic studies and all demonstrated maternal inheritance of the deletion. These findings, together with other reported cases of the origin of the chromosome 15 deletion in AS, suggest that deletion of the maternally contributed chromosome leads to the AS phenotype. This contrasts with the Prader-Willi syndrome (PWS) in which a similar deletion of the paternally contributed chromosome 15 is observed. In deletion cases, a parental gamete effect such as genomic imprinting may be the best model to explain why apparently identical 15q11-q13 deletions may develop the different phenotypes of AS or PWS.     

Molecular dissection of the Prader-Willi/Angelman syndrome region (15q11-13) by YAC cloning and FISH analysis

The authors wish to note an error in the relative order of probesPW71 and TD189-1. The order of PWS/AS probes should be revisedas follows: cen-IR39-ML34-IR4-3R-TD189-1-PW71-LS6-1-TD3-21-GABRB3-IR10-1-CMW1-tel.Thecorrected map of the PWS/AS critical region (Figure 4) summarizingprobe order from interphase FISH analysis and YAC contig informationis provided below. The reversed order of TD189-1 and PW71 wasdiscovered by analysis of YAC 71B11, a non-chimeric YAC of 700kb from the CEPH library which was identified with the STS fromIR4-3R as indicated in Table 1. This YAC was also positive foran STS from the left end of YAC 307A12, identified with theSTS from TD 189-1. In confirmation of these results, the rightend of 71B11 was positive for 307A12 by hybridization, and anSTS from the left end of 71B11 was positive for YACs 172A10and 495D1. The Alu-PCR dot-blot hybridization experiment inFigure 1a appears to represent a false positive overlap betweenYACs A156E1 (used as probe) and B58C7 (labeled 58 on the figure).This is possibly due to homologous sequences contained withinthese two YACs. All other Alu-PCR dot-blot experiments havebeen confirmed by either YAC end hybridization by STS analysis.     

Deletion of chromosome 15 (q11-13) in a Prader-Labhart-Willi syndrome clinic population

Deletion of the long arm of chromosome 15 has recently been reported in a number of patients with the Prader-Labhart-Willi syndrome who were studied with prometaphase banding. We performed cytogenetic analysis on 12 patients with this disorder in whom the clinical diagnosis was certain. A specific cytogenetic anomaly, del(15q11-13) was found in all of the 12 patients. In nine of the 12, the deletion was noted in all cells examined; in two, there was mosaicism, some cells having the deletion and others being normal; one patient had a 7;15 translocation. No clinical differences were evident between individuals with mosaicism for the translocation and those with the typical deletion in all cells examined. The finding that all of our patients with Prader-Labhart-Willi syndrome have a cytogenetic anomaly, with some patients having mosaicism, distinguishes the results of this study from those of previous reports. Prometaphase chromosome analysis is recommended in all individuals clinically suspected of having Prader-Labhart-Willi syndrome and should be considered in hypotonic infants without a specific diagnosis.     

Is Angelman syndrome an alternate result of del(15)(q11q13)?

Two unrelated females, age 15 and 5 years respectively, were studied cytogenetically because of severe mental retardation, seizures and ataxia-like incoordination. A similar deletion of the proximal long arm of chromosome 15 was found in both patients. Re-evaluation showed no voracious appetite or obesity; normal size of hands and feet, minimal to no hypotonia by history or examination and facial features not typical of the Prader-Willi syndrome. However, the facial appearance of the girls was similar to each other with mild hypertelorism. The similarity of these girls and dissimilarity to Prader-Willi syndrome suggest a different syndrome, perhaps the result of deletion of a different segment of 15q. The findings of ataxic-like movements, frequent, unprovoked and prolonged bouts of laughter and facial appearance are more compatible with the diagnosis of Angelman syndrome.     

Unusual clinical features in an Angelman syndrome patient with uniparental disomy due to a translocation 15q15q

We had previously described a patient with an overgrowth syndrome and the chromosome constitution 45,XY,t(15q15q) (Wajntal et al., DNA Cell Biol 1993: 12: 227–231). Clinical reassessment and the use of molecular studies, including methylation analysis with an SNRPN probe, microsatellite analyses of D15S11 , GABRB3 and D15S113 loci, and fluorescence in situ hybridization (FISH) using the SNRPN and GABRB3 probes, are consistent with a diagnosis of Angelman syndrome (AS) due to paternal isodisomy. This is the fourth report case of a translocation 15q15q with paternal uniparental disomy (UPD). Our findings suggest that some patients with clinical features of AS have hyperphagia and obesity with overgrowth, and that these features should not rule out a diagnosis of AS.     

An association analysis of microsatellite markers across the Prader-Willi/Angelman critical region on chromosome 15 (q11-13) and autism spectrum disorder.

Autism (OMIM 209850) is a neurodevelopmental disorder with a significant genetic component of a complex nature. Cytogenetic abnormalities in the Prader-Willi/Angelman syndrome critical region (PWACR) on chromosome 15 (q11-13) have been described in several individuals with autism. We have examined five microsatellite markers spread across the 4 Mb PWACR for linkage disequilibrium (LD) in 148 families with autism spectrum disorder (ASD) and a subset of 82 families with autism using the extended transmission disequilibrium test (ETDT). The markers examined were D15S11, D15S128, D15S1506, GABRB3, and D15S1002. In addition we have examined the microsatellite D15S822 for hemizygous deletion status in our sample as it had been previously reported to be increased in autism. We found no significant LD with any of the markers tested either in the ASD or autism families when looking at paternal and maternal meioses combined. However, as there are known imprinted genes in the region, including possibly GABRB3, we also examined for LD in paternal and maternal meioses separately. Examining paternal transmissions only, we found marginal evidence for LD with a protective allele at marker D15S11 in the ASD families (Chi-sq 7 df, P = 0.05) and marginal evidence for risk alleles at markers D15S1506 (Chi-sq 13.7, 6 df, P = 0.06), GABRB3 (Chi-sq 15.9, 8 df, P = 0.11) and D15S1002 (Chi-sq 17.7, 9 df, P = 0.08) in the autism only families. The allele responsible for the association with GABRB3 is the 191 allele which was previously reported to be overtransmitted. Hemizygous deletion of the microsatellite D15S822 was found in 3 out of 340 independent chromosomes in our sample; a rate of 0.8%. This is not significantly different to the frequency in the general population. In conclusion, our results did not rule out the involvement of this chromosomal region, but provided further evidence, albeit very limited, to implicate GABRB3. Further more systematic work in larger samples is required and confirmation that GABRB3 is imprinted is desirable.     

Interstitial duplications of chromosome region 15q11q13: Clinical and molecular characterization

Duplications of chromosome region 15q11q13 often occur as a supernumerary chromosome 15. Less frequently they occur as interstitial duplications [dup(15)]. We describe the clinical and molecular characteristics of three patients with de novo dup(15). The patients, two males and one female (ages 3–21 years), had nonspecific findings that included autistic behavior, hypotonia, and variable degrees of mental retardation. The extent, orientation, and parental origin of the duplications were assessed by fluorescent in situ hybridization, microsatellite analyses, and methylation status at D15S63. Two patients had large direct duplications of 15q11q13 [dir dup(15)(q11q13)] that extended through the entire Angelman syndrome/Prader-Willi syndrome (AS/PWS) chromosomal region. Their proximal and distal breaks, at D15S541 or D15S9 and between D15S12 and D15S24, respectively, were comparable to those found in the common AS/PWS deletions. This suggests that duplications and deletions may be the reciprocal product of an unequal recombination event. These two duplications were maternally derived, but the origin of the chromatids involved in the unequal crossing over in meiosis differs. In one patient, the duplication originated from two different maternal chromosomes, while in the other patient it arose from the same maternal chromosome. The third patient had a much smaller duplication that involved only D15S11 and parental origin could not be determined. There was no obvious correlation between phenotype and extent of the duplication in these patients. Am. J. Med. Genet. 79:82–89, 1998. © 1998 Wiley-Liss, Inc.     

Angelman syndrome in a daughter with del(15) (q11q13) associated with brachycephaly, hearing loss, enlarged foramen magnum, and ataxia in the mother

We report on a 4-year-old girl with Angelman syndrome who has an apparent de-novo del(15) (q11q13) originating from a maternally derived chromosome. Her mother had severe brachycephaly, sensorineural hearing loss, speech impediment, and mild ataxia. CT brain scans showed an enlarged foramen magnum in the mother and daughter but magnetic resonance imaging (MRI) showed no brainstem abnormality in either. This family demonstrates that some Angelman syndrome cases may be dominantly transmitted with variable expression and associated with abnormal or cytogenetically apparently normal chromosome 15.     

Angelman syndrome without detectable chromosome 15q11–13 anomaly: Clinical study of familial and isolated cases

The clinical findings in 12 Angelman syndrome (AS) patients (4 sib pairs and 4 sporadic cases, aged 12–55 years) without a cytogenetic or molecular detectable defect at the AS locus were compared to those of 28 AS patients (aged 11–50 years) with a deletion, in order to determine whether the clinical spectrum differed between the two groups. There were only two minor differences, i.e., mandibular prognathism was always found in the patients with a defect (100% vs. 58%), whereas truncal hypotonia was found less frequently in the group with a detectable genetic defect (54% vs. 91%). All other clinical and physical characteristics were equally represented in the two groups. Epileptic seizures occurred in 93% and 75%, respectively, of patients with and without a detectable chromosome 15 defect. Specific EEG patterns were found in 90% of both groups. The clinical signs and symptoms of our patients closely resemble those in familial AS cases reported in the literature, with the exception of scoliosis, which was present in 55% of the patients in our study. We conclude that the absence of a detectable cytogenetic or molecular defect at the AS locus is not associated with a strikingly different AS phenotype, compared to those with such a defect. Mutation analysis of the UBE3A gene in our patients without a detectable genetic defect, especially in the familial cases, is currently underway. Am. J. Med. Genet. 76:262–268, 1998. © 1998 Wiley-Liss, Inc.     

A 5-year-old white girl with Prader-Willi syndrome and a submicroscopic deletion of chromosome 15q11q13

We report on a 5-year-old white girl with Prader-Willi syndrome (PWS) and a submicroscopic deletion of 15q11q13 of approximately 100–200 kb in size. High resolution chromosome analysis was normal but fluorescence in situ hybridization (FISH), Southern hybridization, and microsatellite data from the 15q11q13 region demonstrated that the deletion was paternal in origin and included the SNRPN, PAR-5, and PAR-7 genes from the proximal to distal boundaries of the deletion segment. SNRPN and PW71B methylation studies showed an abnormal pattern consistent with the diagnosis of PWS and supported the presence of a paternal deletion of 15q11q13 or an imprinting mutation. Biparental (normal) inheritance of PW71B (D15S63 locus) and a deletion of the SNRPN gene were observed by microsatellite, quantitative Southern hybridization, and/or FISH analyses. Our patient met the diagnostic criteria for PWS, but has no reported behavior problems, hyperphagia, or hypopigmentation. Our patient further supports SNRPN and possibly other genomic sequences which are deleted as the cause of the phenotype recognized in PWS patients. © 1996 Wiley-Liss, Inc.