Ann Haskins Olney

Refinement of a 400-kb Critical Region Allows Genotypic Differentiation between Isolated Lissencephaly, Miller-Dieker Syndrome, and Other Phenotypes Secondary to Deletions of 17p13.3

Deletions of 17p13.3, including the LIS1 gene, result in the brain malformation lissencephaly, which is characterized by reduced gyration and cortical thickening; however, the phenotype can vary from isolated lissencephaly sequence (ILS) to Miller-Dieker syndrome (MDS). At the clinical level, these two phenotypes can be differentiated by the presence of significant dysmorphic facial features and a more severe grade of lissencephaly in MDS. Previous work has suggested that children with MDS have a larger deletion than those with ILS, but the precise boundaries of the MDS critical region and causative genes other than LIS1 have never been fully determined. We have completed a physical and transcriptional map of the 17p13.3 region from LIS1 to the telomere. Using fluorescence in situ hybridization, we have mapped the deletion size in 19 children with ILS, 11 children with MDS, and 4 children with 17p13.3 deletions not involving LIS1. We show that the critical region that differentiates ILS from MDS at the molecular level can be reduced to 400 kb. Using somatic cell hybrids from selected patients, we have identified eight genes that are consistently deleted in patients classified as having MDS. In addition, deletion of the genes CRK and 14-3-3 epsilon delineates patients with the most severe lissencephaly grade. On the basis of recent functional data and the creation of a mouse model suggesting a role for 14-3-3 epsilon in cortical development, we suggest that deletion of one or both of these genes in combination with deletion of LIS1 may contribute to the more severe form of lissencephaly seen only in patients with MDS.

De novo ACTA2 mutation causes a novel syndrome of multisystemic smooth muscle dysfunction

Smooth muscle cells (SMCs) contract to perform many physiological functions, including regulation of blood flow and pressure in arteries, contraction of the pupils, peristalsis of the gut, and voiding of the bladder. SMC lineage in these organs is characterized by cellular expression of the SMC isoform of α‐actin, encoded by the ACTA2 gene. We report here on a unique and de novo mutation in ACTA2, R179H, that causes a syndrome characterized by dysfunction of SMCs throughout the body, leading to aortic and cerebrovascular disease, fixed dilated pupils, hypotonic bladder, malrotation, and hypoperistalsis of the gut and pulmonary hypertension.

Mutations in ZIC2 in Human Holoprosencephaly: Description of a Novel ZIC2-Specific Phenotype and Comprehensive Analysis of 157 Individuals

Background Holoprosencephaly (HPE), the most common malformation of the human forebrain, may be due to mutations in genes associated with non-syndromic HPE. Mutations in ZIC2, located on chromosome 13q32, are a common cause of non-syndromic, non-chromosomal HPE. Objective To characterise genetic and clinical findings in patients with ZIC2 mutations. Methods Through the National Institutes of Health and collaborating centres, DNA from approximately 1200 individuals with HPE spectrum disorders was analysed for sequence variations in ZIC2. Clinical details were examined and all other known cases of mutations in ZIC2 were included through a literature search. Results By direct sequencing of DNA samples of an unselected group of unrelated patients with HPE in our NIH laboratory, ZIC2 mutations were found in 8.4% (49/582) of probands. A total of 157 individuals from 119 unrelated kindreds are described, including 141 patients with intragenic sequence determined mutations in ZIC2. Only 39/157 patients have previously been clinically described. Unlike HPE due to mutations in other genes, most mutations occur de novo and the distribution of HPE types differs significantly from that of non-ZIC2 related HPE. Evidence is presented for the presence of a novel facial phenotype which includes bitemporal narrowing, upslanting palpebral fissures, a short nose with anteverted nares, a broad and well demarcated philtrum, and large ears. Conclusions HPE due to ZIC2 mutations is distinct from that due to mutations in other genes. This may shed light on the mechanisms involved in formation of the forebrain and face and will help direct genetic counselling and diagnostic strategies.

Report from the workshop on Pallister-Hall syndrome and related phenotypes

A one day workshop was convened on the NIH campus on March 1, 1996, in Bethesda, Maryland to discuss emerging clinical and molecular information on Pallister-Hall syndrome (PHS) and related disorders. PHS is a pleiotropic autosomal dominant disorder comprising hypothalamic hamartoma, pituitary dysfunction, central polydactyly, and visceral malformations. The goals of the meeting were to update participants in the latest clinical and research findings in the disorder, review the history and evolution of the understanding of the phenotype, determine diagnostic criteria for PHS, and make recommendations for clinical evaluation of individuals affected by PHS. These topics were addressed by several speakers and data were displayed from several of the large pedigrees of autosomal dominant PHS. 37 refs., 4 tabs.

Diamond–Blackfan anemia with mandibulofacial dystostosis is heterogeneous, including the novel DBA genes TSR2 and RPS28

Patients with physical findings suggestive of Treacher Collins syndrome (TCS) or mandibulofacial dysostosis (MFD) and macrocytic anemia diagnostic of Diamond–Blackfan anemia (DBA) have been reported. Disease‐causing genes have been identified for TCS and other MFDs. Mutations in several ribosomal protein genes and the transcription factor GATA1 result in DBA. However, no disease‐causing mutation had been identified in the reported patients with the combination of TCS/MFD and DBA phenotype, and we hypothesized that pathogenic mutations in a single gene could be identified using whole exome analysis. We studied probands from six unrelated families. Combining exome analysis and Sanger sequencing, we identified likely pathogenic mutations in 5/6 families. Two mutations in unrelated families were seen in RPS26, the known DBA10 gene. One variant was predicted to affect mRNA splicing, and the other to lead to protein truncation. In another family a likely pathogenic X‐linked mutation affecting a highly conserved residue was found in TSR2, which encodes a direct binding partner of RPS26. De novo mutations affecting the RPS28 start codon were found in two unrelated probands, identifying RPS28 as a novel disease gene. We conclude that the phenotype combining features of TCS with DBA is genetically heterogeneous. Each of the pathogenic variants identified is predicted to impede ribosome biogenesis, which in turn could result in altered cell growth and proliferation, causing abnormal embryologic development, defective erythropoiesis and reduced growth. The phenotype combining TCS/MFD and DBA is highly variable, overlaps with DBA and lies within the phenotypic spectrum of ribosomopathies.

Array-based comparative genomic hybridization analysis of 1176 consecutive clinical genetics investigations

Purpose: Cytogenetic investigations are useful for etiologic determinations of mental retardation, developmental delay, multiple congenital anomalies, and pregnancy complications; however, the causes remain elusive in a majority of cases despite high-resolution cytogenetic studies and multiple fluorescence in situ hybridization examinations. Array-based comparative genomic hybridization has the ability to examine the genome at a higher resolution and may yield an increased detection of genetic abnormalities. The purpose of this study was to assess the use of array-based comparative genomic hybridization in a clinical genetics setting.Methods: DNA from 1176 patients was analyzed using a bacterial artificial chromosome array-based comparative genomic hybridization platform. All abnormal cases were confirmed by fluorescence in situ hybridization and parental studies were completed when possible.Results: Of the 1176 patients included in this survey, 163 showed a genomic imbalance identified by array-based comparative genomic hybridization. Of these 163 cases, 116 had a clinically relevant genetic abnormality. A total of 9.8% (116 of 1176 cases) were determined to exhibit a causative genomic imbalance. Twenty-five of the 116 abnormal cases had a previously identified cytogenetic abnormality yielding an increased detection rate of 7.9% (91 of 1146) in cases with normal or no cytogenetics.Conclusion: Array-based comparative genomic hybridization increases the overall abnormality detection rate, thus improving the diagnostic potential of clinical cytogenetics investigations.

Mapping of the human complement factor I gene to 4q25

A detailed genetic and physical map of human complement factor I (IF) using somatic cell hybrids, in situ hybridization, and genetic linkage is reported. The gene has been localized to band 4q25. The order GC-INP10-ADH3-EGF-IF-IL2-MNS is proposed for genes on 4q on the basis of genetic and physical mapping techniques. A BclI polymorphism found with the IF probe demonstrated a maternal origin for a de novo deletion of chromosome 4 that was used in physically mapping the gene. The genetic and physical distances around band 4q24 suggest that 1 cM is approximately 1.2 million bp of DNA. This work provides a useful addition to the map of 4q.

SMARCAL1 deficiency predisposes to non‐Hodgkin lymphoma and hypersensitivity to genotoxic agents in vivo

Schimke immuno‐osseous dysplasia (SIOD) is a multisystemic disorder with prominent skeletal, renal, immunological, and ectodermal abnormalities. It is caused by mutations of SMARCAL1 (SWI/SNF‐related, matrix‐associated, actin‐dependent regulator of chromatin, subfamily a‐like 1), which encodes a DNA stress response protein. To determine the relationship of this function to the SIOD phenotype, we profiled the cancer prevalence in SIOD and assessed if defects of nucleotide excision repair (NER) and nonhomologous end joining (NHEJ), respectively, explained the ectodermal and immunological features of SIOD. Finally, we determined if Smarcal1del/del mice had hypersensitivity to irinotecan (CPT‐11), etoposide, and hydroxyurea (HU) and whether exposure to these agents induced features of SIOD. Among 71 SIOD patients, three had non‐Hodgkin lymphoma (NHL) and one had osteosarcoma. We did not find evidence of defective NER or NHEJ; however, Smarcal1‐deficient mice were hypersensitive to several genotoxic agents. Also, CPT‐11, etoposide, and HU caused decreased growth and loss of growth plate chondrocytes. These data, which identify an increased prevalence of NHL in SIOD and confirm hypersensitivity to DNA damaging agents in vivo, provide guidance for the management of SIOD patients.

Collagen XVIII mutation in Knobloch syndrome with acute lymphoblastic leukemia

Knobloch syndrome (KNO) is caused by mutations in the collagen XVIII gene (COL18A1) and patients develop encephalocele and vitreoretinal degeneration. Here, we report an El Salvadorian family where two sisters showed features of KNO. One of the siblings also developed acute lymphoblastic leukemia. DNA sequencing of COL18A1 revealed a homozygous, 2‐bp deletion (c3514‐3515delCT) in exon 41, which leads to abnormal collagen XVIII and deficiency of its proteolytic cleavage product endostatin. KNO patients with mutations in COL18A1 may be at risk for endostatin‐related conditions including malignancy.

Carriers and patients with muscle–eye–brain disease can be rapidly diagnosed by enzymatic analysis of fibroblasts and lymphoblasts

We report a new fibroblast and lymphoblast based protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 enzymatic assay, which allows rapid and accurate diagnosis of carriers and patients with muscle-eye-brain type of congenital muscular dystrophy. Seven patients with genetically confirmed muscle-eye-brain disease were assayed for protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 enzyme activity. In three patients and their heterozygous parents, the assays were done on EBV-transformed lymphoblasts, in another three patients they were done on cultured fibroblasts and in the last patient on both fibroblasts and lymphoblasts. Cultured fibroblasts and lymphoblasts from the muscle-eye-brain patients showed a highly significant decrease in protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 activity relative to controls. The residual protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 level in fibroblasts (average 0.11 nmoles/h per mg) was about 13% of normal controls. The ratio of protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 activity to the activity of a glycosyltransferase control (N-acetylglucosaminyltransferase 1; GnT1) in fibroblasts was on average 0.006 in muscle-eye-brain patients and 0.045 in controls. The average residual protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 level in lymphoblasts was 15% of normal controls. The average ratio of protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1/GnT1 activity was 0.007 in muscle-eye-brain patients, 0.026 in heterozygous carriers and 0.046 in normal controls. Assay of protein O-mannosyl beta-1,2-N-acetylglucosaminyltransferase 1 activity in fibroblasts and lymphoblasts from muscle-eye-brain carriers and patients provides a rapid and relatively simple diagnostic test for this disease and could be used as a screening test in carriers and patients with complex congenital muscular dystrophy.