Magda Montfort

DNA Hypomethylation at ALOX12 Is Associated with Persistent Wheezing in Childhood

RATIONALE Epigenetic changes may play a role in the occurrence of asthma-related phenotypes. OBJECTIVES To identify epigenetic marks in terms of DNA methylation of asthma-related phenotypes in childhood, and to assess the effect of prenatal exposures and genetic variation on these epigenetic marks. METHODS Data came from two cohorts embedded in the Infancia y Medio Ambiente (INMA) PROJECT: Menorca (n = 122) and Sabadell (n = 236). Wheezing phenotypes were defined at age 4-6 years. Cytosine-guanine (CpG) dinucleotide site DNA methylation differences associated with wheezing phenotypes were screened in children of the Menorca study using the Illumina GoldenGate Panel I. Findings were validated and replicated using pyrosequencing. Information on maternal smoking and folate supplement use was obtained through questionnaires. Dichlorodiphenyldichloroethylene was measured in cord blood or maternal serum. Genotypes were extracted from genome-wide data. MEASUREMENT AND MAIN RESULTS Screening identified lower DNA methylation at a CpG site in the arachidonate 12-lipoxygenase (ALOX12) gene in children having persistent wheezing compared with those never wheezed (P = 0.003). DNA hypomethylation at ALOX12 loci was associated with higher risk of persistent wheezing in the Menorca study (odds ratio per 1% methylation decrease, 1.13; 95% confidence interval, 0.99-1.29; P = 0.077) and in the Sabadell study (odds ratio, 1.16; 95% confidence interval, 1.03-1.37; P = 0.017). Higher levels of prenatal dichlorodiphenyldichloroethylene were associated with DNA hypomethylation of ALOX12 in the Menorca study (P = 0.033), but not in the Sabadell study (P = 0.377). ALOX12 DNA methylation was strongly determined by underlying genetic polymorphisms. CONCLUSIONS DNA methylation of ALOX12 may be an epigenetic biomarker for the risk of asthma-related phenotypes.

Mutation 1091delC is highly prevalent in Spanish Sanfilippo syndrome type A patients

The gene resposible for Sanfilippo syndrome type A, a lysosomal disorder caused by deficiency of sulfamidase, was recently cloned and more than 40 mutations were identified. This paper presents the mutation analysis and clinical findings in 11 Spanish patients in whom 19 of the 22 mutant alleles have been identified. This is the first report on mutations in Spanish Sanfilippo A patients. Seven different mutations were found, four of which (Q85R, R206P, A354P, and L386R) were not previously described. Mutation 1091del C was the most prevalent, accounting for nearly one‐half of the mutated alleles, while mutations R245H and R74C were not found. Haplotype analysis suggests a founder effect as the cause of the high frequency of 1091del C in this population. Hum Mutat 12:274–279, 1998.

Design and evaluation of a panel of single-nucleotide polymorphisms in microRNA genomic regions for association studies in human disease.

MicroRNAs (miRNA) are recognized posttranscriptional gene repressors involved in the control of almost every biological process. Allelic variants in these regions may be an important source of phenotypic diversity and contribute to disease susceptibility. We analyzed the genomic organization of 325 human miRNAs (release 7.1, miRBase) to construct a panel of 768 single-nucleotide polymorphisms (SNPs) covering ∼1 Mb of genomic DNA, including 131 isolated miRNAs (40%) and 194 miRNAs arranged in 48 miRNA clusters, as well as their 5-kb flanking regions. Of these miRNAs, 37% were inside known protein-coding genes, which were significantly associated with biological functions regarding neurological, psychological or nutritional disorders. SNP coverage analysis revealed a lower SNP density in miRNAs compared with the average of the genome, with only 24 SNPs located in the 325 miRNAs studied. Further genotyping of 340 unrelated Spanish individuals showed that more than half of the SNPs in miRNAs were either rare or monomorphic, in agreement with the reported selective constraint on human miRNAs. A comparison of the minor allele frequencies between Spanish and HapMap population samples confirmed the applicability of this SNP panel to the study of complex disorders among the Spanish population, and revealed two miRNA regions, hsa-mir-26a-2 in the CTDSP2 gene and hsa-mir-128-1 in the R3HDM1 gene, showing geographical allelic frequency variation among the four HapMap populations, probably because of differences in natural selection. The designed miRNA SNP panel could help to identify still hidden links between miRNAs and human disease.

Genetic fine localization of the β-glucocerebrosidase (GBA) and prosaposin (PSAP) genes: implications for Gaucher disease

Abstract Mutations in the glucocerebrosidase (GBA) and prosaposin (PSAP) genes are responsible for Gaucher disease, the most prevalent sphingolipidosis. Somatic cell hybrid analysis and in situ hybridization experiments have localized the GBA gene to 1q21 and the PSAP gene to 10q21-q22. We performed pairwise and multi-point linkage analyses between the two genes and several highly polymorphic markers from the Généthon human linkage map. Our results show that six markers cosegregate with the GBA gene (Zmax = 8.73 at θ = 0.00 for marker D1S2714) and define a 3.2-cM interval between D1S305 and D1S2624 as the most probable location for the gene. Three of these markers (D1S2777, D1S303, and D1S2140), as well as the gene encoding pyruvate kinase (PKLR), are contained in a single YAC clone together with the GBA gene. A new polymorphism was identified within the PSAP gene (C16045T) and used for linkage studies. The multi-point analysis places the gene in a 9.8-cM interval between D10S1688 and D10S607. The fine localization of these genes provides a useful tool for cosegregation analysis, indirect molecular diagnosis, and population genetic studies.

Familial hemiplegic migraine: linkage to chromosome 14q32 in a Spanish kindred

We sought to map the disease-causing gene in a large Spanish kindred with familial hemiplegic migraine (FHM). Patients were classified according to the ICHD-II criteria. After ruling out linkage to known migraine genetic loci, a single nucleotide polymorphism-based, 0.62-cM density genome-wide scan was performed. Among 13 affected subjects, FHM was the prevailing migraine phenotype in six, migraine with aura in four and migraine without aura in three. Linkage analysis revealed a disease locus in a 4.15-Mb region on 14q32 with a maximum two-point logarithm of odds (LOD) score of 3.1 and a multipoint parametric LOD score of 3.8. This genomic region does not overlap with the reported migraine loci on 14q21–22. Sequence analysis of three candidate genes in the region, SLC24A4, ATXN3 and ITPK1, failed to show disease-causing mutations in our patients. Genetic heterogeneity in FHM may be greater than previously suspected.

Gaucher Disease: The N370S Mutation in Ashkenazi Jewish and Spanish Patients has a Common Origin and Arose Several Thousand Years Ago

This work was supported by Comision Interministerial de Ciencia y Technologia (SAF 97-0074). We are grateful to Dr. E. Beutler for critical reading of the manuscript; to L. Gort, M. Martinez, and J. Armstrong for technical assistance; to R. Rycroft for revising the English; and to Dr. N. Espiro and Dr. J. Israel for helpful comments on Jewish history.

A de novo nonsense mutation in MAGEL2 in a patient initially diagnosed as Opitz-C: similarities between Schaaf-Yang and Opitz-C syndromes

Opitz trigonocephaly C syndrome (OTCS) is a rare genetic disorder characterized by craniofacial anomalies, variable intellectual and psychomotor disability, and variable cardiac defects with a high mortality rate. Different patterns of inheritance and genetic heterogeneity are known in this syndrome. Whole exome and genome sequencing of a 19-year-old girl (P7), initially diagnosed with OTCS, revealed a de novo nonsense mutation, p.Q638*, in the MAGEL2 gene. MAGEL2 is an imprinted, maternally silenced, gene located at 15q11-13, within the Prader-Willi region. Patient P7 carried the mutation in the paternal chromosome. Recently, mutations in MAGEL2 have been described in Schaaf-Yang syndrome (SHFYNG) and in severe arthrogryposis. Patient P7 bears resemblances with SHFYNG cases but has other findings not described in this syndrome and common in OTCS. We sequenced MAGEL2 in nine additional OTCS patients and no mutations were found. This study provides the first clear molecular genetic basis for an OTCS case, indicates that there is overlap between OTCS and SHFYNG syndromes, and confirms that OTCS is genetically heterogeneous. Genes encoding MAGEL2 partners, either in the retrograde transport or in the ubiquitination-deubiquitination complexes, are promising candidates as OTCS disease-causing genes.

On the Age of the Most Prevalent Gaucher Disease–Causing Mutation, N370S

To the Editor:We have recently described a common origin for the most prevalent mutation (N370S) observed among Gaucher disease (GD) patients of Ashkenazi Jewish (AJ) and Spanish descent (Diaz et al. 1999xGaucher disease: the N370S mutation in Ashkenazi Jewish and Spanish patients has a common origin and arose several thousand years ago. Diaz, A, Montfort, M, Cormand, B, Zeng, BJ, Pastores, GM, Chabas, A, Vilageliu, L et al. Am J Hum Genet. 1999; 64: 1233–1238Abstract | Full Text | Full Text PDF | PubMed | Scopus (15)See all References1999). We also estimated the age of this mutation, using a formula described by Risch et al. (1995bxGenetic analysis of idiopathic torsion dystonia in Ashkenazi Jews and their recent descent from a small founder population. Risch, N, DeLeon, D, Ozelius, L, Kramer, P, Almasy, L, Singer, B, Fahn, S et al. Nat Genet. 1995b; 9: 152–159Crossref | PubMed | Scopus (299)See all References1995b). Unfortunately, as R. Colombo pointed out in a recent report (Colombo 2000xAge estimate of the N370S mutation causing Gaucher disease in Ashkenazi Jews and European populations: a reappraisal of haplotype data. Colombo, R. Am J Hum Genet. 2000; 66: 692–697Abstract | Full Text | Full Text PDF | PubMed | Scopus (22)See all References2000), there was an error in the formula presented in the original publication that was never rectified. In a reply (Risch et al. 1995axITD in Ashkenezi Jews—genetic drift or selection? In reply. Risch, N, DeLeon, D, Fahan, S, Ozelius, L, Breakfield, X, Kramer, P, Almasy, L et al. Nat Genet. 1995a; 11: 14–15CrossrefSee all References1995a) to criticisms raised by Zoossmann-Diskin (1995xITD in Ashkenazi Jews—genetic drift or selection?. Zoossmann-Diskin, A. Nat Genet. 1995; 11: 13–14Crossref | PubMed | Scopus (4)See all References1995), Risch et al. made no mention of an error in the formula. The continued application of the formula by researchers who may not be well versed in the field may lead to repeated mistakes if the error remains uncorrected. We apologize for our failure to recognize the error, but we are pleased that it has been identified by Colombo, who had no difficulty in re-estimating the age of the mutation, using our data. This reappraisal indicates that the N370S mutation may have occurred between the 11th and 13th centuries or even in the 10th century, considering 30 years per generation as suggested recently by Tremblay and Vezina (2000xNew estimates of intergenerational time intervals for the calculation of age and origin of mutations. Tremblay, M and Vezina, H. Am J Hum Genet. 2000; 66: 651–658Abstract | Full Text | Full Text PDF | PubMed | Scopus (117)See all References2000). This new estimated date for the mutation suggests that it occurred (or entered) the AJ population after the separation of the Ashkenazi and Sephardic Jewish traditions, which would be consistent with the apparent absence of this mutation among Sephardic patients. Using a totally different approach based on mutation detection in healthy Roman Jews, Oddoux et al. (1999xMendelian diseases among Roman Jews: implications for the origins of disease alleles. Oddoux, C, Guillen-Navarro, E, Ditivoli, C, Dicave, E, Cilio, MR, Clayton, CM, Nelson, H et al. J Clin Endocrinol Metab. 1999; 84: 4405–4409Crossref | PubMedSee all References1999) also got to the conclusion that the N370S is an old mutation. Further information on the origin of a mutation could be provided from the length of the chromosomal region noted with linkage disequilibrium (LD) and the strength of the LD. In this case, the data can be used to determine whether the N370S mutation had a Jewish or non-Jewish origin. The 3.2-cM region in LD in AJ (Diaz et al. 1999xGaucher disease: the N370S mutation in Ashkenazi Jewish and Spanish patients has a common origin and arose several thousand years ago. Diaz, A, Montfort, M, Cormand, B, Zeng, BJ, Pastores, GM, Chabas, A, Vilageliu, L et al. Am J Hum Genet. 1999; 64: 1233–1238Abstract | Full Text | Full Text PDF | PubMed | Scopus (15)See all References1999) seems to be shorter in Spanish chromosomes, as the flanking marker D1S2624 is in LD among AJ, whereas it is not in Spanish GD patients. Moreover, LD values are stronger in AJ than in Spanish chromosomes. These observations suggest that the mutation was introduced later in the AJ population, a statement that is independent of the actual age of the mutation. However, these results should be taken with caution because of the small number of Spanish chromosomes included in the study. Further work would be necessary to settle this issue which, to date, remains an open question.

Reliable co-segregation analysis for prenatal diagnosis and heterozygote detection in Gaucher disease

Mutations in the gene encoding β‐glucocerebrosidase are the main cause of Gaucher disease. The identification of some of these mutations in prenatal tests is a good complement to enzymatic assay and allows diagnosis and, in some cases, prognosis of the disease to be made. DNA analysis is particularly useful for carrier detection since the results of biochemical analyses are often ambiguous. The main drawback of mutation analysis for prenatal diagnosis and carrier detection in Gaucher disease is that rare mutations account for more than 30 per cent of the mutant alleles in most populations. The individual detection of these mutations is too expensive and time‐consuming for routine use. Here we present a diagnostic protocol based on co‐segregation analysis, using highly polymorphic markers, to be applied when at least one disease allele does not correspond to the most common mutations. Because of the frequency of the N370S mutation and its relevance for prognosis, an improved PCR detection method is included.