[Show abstract][Hide abstract] ABSTRACT: Holoprosencephaly (HPE) is the most common developmental brain anomaly in human, associated with a wide spectrum of presentations. The etiology is heterogeneous, due to environmental and genetic factors. Out of 12 cytogenetic candidate loci previously reported, eight were subtelomeric, including the loci in which two of the four major HPE genes were identified (SHH and TGIF). Recently, we reported that these two genes could be mutated or microdeleted. Therefore, we hypothesized that subtelomeres screening in HPE patients could refine the known subtelomeric candidate loci and identify novel ones. In this study, 181 samples, 72 fetuses and 109 live-born infants, with HPE and a normal karyotype, and 10 patients deleted for SHH or TGIF (3.5 Mb from telomeres) were screened for subtelomeric rearrangements using the multiplex ligation probe-dependent amplification (MLPA) method with two kits. Quantitative PCR was performed when discrepancies were observed between these two kits. We found that known SHH and TGIF microdeletions on 7q and 18p, encompassed their subtelomeric region (3.5 Mb) and were often associated with cryptic gains. Out of the 181 samples, we detected rearrangements in known candidate HPE loci (1q, 20p, and 21q) as well as in other novel subtelomeric locations (1p, 5q, 8p, 17q, 18q, 22q, and Xq) and in the subcentromeric 15q. We also found associations between cryptic subtelomeric gain and loss that may be inherited from a parental balanced translocation, which is helpful for genetic counseling. These findings reinforce the multihit origin for HPE and contribute to the explanation of the wide phenotypic spectrum described in this developmental disorder.
Human Mutation 12/2007; 28(12):1189-97. DOI:10.1002/humu.20594 · 5.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Holoprosencephaly (HPE), the most common structural malformation of the forebrain in humans, can be detected early during pregnancy using prenatal ultrasonography . Among foetuses with a normal karyotype, 14% have mutations in the four main HPE genes (SHH, ZIC2, SIX3 and TGIF). Genomic rearrangements have now been implicated in many genetic diseases, so we hypothesized that microdeletions in the major HPE genes may also be common in HPE foetuses with severe phenotype or other associated malformations. We screened the DNA obtained from 94 HPE foetuses with a normal karyotype for the presence of microdeletions involving the four major HPE genes (SHH, ZIC2, SIX3 and TGIF). Thirteen of the foetuses had a point mutation in one of the 4 genes and 81 had no known mutations. Quantitative multiplex PCR of short fluorescent fragments (QMPSF) analysis was used for rapid determination of HPE genes copy numbers and the identified microdeletions were confirmed by real time quantitative PCR, or fluorescent in situ hybridization (FISH) (if a cell line was available). Microdeletions were detected in 8 of 94 foetuses (8.5%) (2 in SHH, 2 in SIX3, 3 in ZIC2 and 1 in TGIF genes), and only among the 81 foetuses with a normal karyotype and no point mutations. These data suggest that microdeletions in the four main HPE genes are a common cause of prenatal HPE, as well as point mutations, and increase the total diagnosis rate close to approximately 22.3% of foetuses with normal karyotype. Detection can be achieved by the QMPSF testing method that proved to be efficient for testing several genes in a single assay.
Human Genetics 04/2006; 119(1-2):1-8. DOI:10.1007/s00439-005-0097-6 · 4.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mutations of the developmental gene Sonic hedgehog (SHH) and alterations of SHH signaling have been associated with holoprosencephaly (HPE), a rare disorder characterized by a large spectrum of brain and craniofacial anomalies. Based on the crystal structure of mouse N-terminal and Drosophila C-terminal hedgehog proteins, we have developed three-dimensional models of the corresponding human proteins (SHH-N, SHH-C) that have allowed us to identify within these two domains crucial regions associated with HPE missense mutations. We have further characterized the functional consequences linked to 11 of these mutations. In transfected HEK293 cells, the production of the active SHH-N fragment was dramatically impaired for eight mutants (W117R, W117G, H140P, T150R, C183F, L271P, I354T, A383T). The supernatants from these cell cultures showed no significant SHH-signaling activity in a reporter cell-based assay. Two mutants (G31R, D222N) were associated with a lower production of SHH-N and signaling activity. Finally, one mutant harboring the A226T mutation displays an activity comparable with the wild-type protein. This work demonstrates that most of the HPE-associated SHH mutations analyzed have a deleterious effect on the availability of SHH-N and its biological activity. However, because of the lack of correlation between genotype and phenotype for SHH-associated mutations, our study suggests that other factors intervene in the development of the spectrum of HPE anomalies.
[Show abstract][Hide abstract] ABSTRACT: Since 1996, a European network has been organized from Rennes, France and holoprosencephalic files were collected for clinical and molecular study. Familial instances of typical and atypical holoprosencephaly (HPE) were found in 30% of cases. All affected children had psychomotor delay with microcephaly, often associated with endocrine, digestive, and respiratory abnormalities, and thermal dysregulation. Among 173 subjects in the molecular study, 28 heterozygous mutations were identified (16%): 15 SHH mutations, 6 ZIC2 mutations, 5 SIX3 mutations, and 2 TGIF mutations.
American Journal of Medical Genetics Part A 09/2004; 129A(1):21-4. DOI:10.1002/ajmg.a.30110 · 2.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Holoprosencephaly (HPE; 1 out of 16,000 live births; 1 out of 250 conceptuses) is a complex brain malformation resulting from incomplete cleavage of the prosencephalon, affecting both the forebrain and the face. Clinical expressivity is variable, ranging from a single cerebral ventricle and cyclopia to clinically unaffected carriers in familial dominant autosomic HPE. The disease is genetically heterogeneous, but additional environmental agents also contribute to the etiology of HPE. In our cohort of 200 patients, 34 heterozygous mutations were identified, 24 of them being novel ones: 13 out of 17 in the Sonic hedgehog gene (SHH); 4 out of 7 in ZIC2; and 7 out of 8 in SIX3. The two mutations identified in TGIF have already been reported. Novel phenotypes associated with a mutation have been described, such as abnormalities of the pituitary gland and corpus callosum, colobomatous microphthalmia, choanal aperture stenosis, and isolated cleft lip. This study confirms the great genetic heterogeneity of the disease, the important phenotypic variability in HPE families, and the difficulty to establish genotype-phenotype correlations.
Human Mutation 08/2004; 24(1):43-51. DOI:10.1002/humu.20056 · 5.14 Impact Factor