A spectrum of FOXC1 mutations suggests gene dosage as a mechanism for developmental defects of the anterior chamber of the eye.

Mutations in the forkhead transcription-factor gene (FOXC1), have been shown to cause defects of the anterior chamber of the eye that are associated with developmental forms of glaucoma. Discovery of these mutations was greatly facilitated by the cloning and characterization of the 6p25 breakpoint in a patient with both congenital glaucoma and a balanced-translocation event involving chromosomes 6 and 13. Here we describe the identification of novel mutations in the FOXC1 gene in patients with anterior-chamber defects of the eye. We have detected nine new mutations (eight of which are novel) in the FOXC1 gene in patients with anterior-chamber eye defects. Of these mutations, five frameshift mutations predict loss of the forkhead domain, as a result of premature termination of translation. Of particular interest is the fact that two families have a duplication of 6p25, involving the FOXC1 gene. These data suggest that both FOXC1 haploinsufficiency and increased gene dosage can cause anterior-chamber defects of the eye.

Developmental delay and multiple congenital anomalies in a child with a unique combination of partial monosomy 18 and partial trisomy 16.

A male child with multiple congenital anomalies and developmental delay is described. Cytogenetic evaluation showed that the patient was partially monosomic for the short arm of chromosome 18 and partially trisomic for the short arm of chromosome 16: a combination of chromosomal syndromes not previously described.

Nonsyndromic congenital retinal nonattachment gene maps to human chromosome band 10q21.

Nonsyndromic congenital retinal nonattachment (NCRNA) comprises congenital insensitivity to light, massive retrolental mass, shallow anterior chamber, microphthalmia, and nystagmus. We searched for the location of the gene responsible for an autosomal recessive form of NCRNA by using DNA samples from 36 individuals from a founding population. To this end we applied homozygosity mapping and a DNA pooling strategy using genomewide screen polymorphic microsatellite markers. We used two DNA pools, one pool contained DNA from 16 individuals affected with NCRNA. The second pool contained DNA from 20 normal carrier individuals, the parents of the patients. The polymorphic microsatellite markers were polymerase chain reaction (PCR)-amplified in each DNA pool; the PCR products were electrophoresed on polyacrylamide gels and visualized by silver staining. The banding patterns from DNA pools of affected and unaffected persons were compared, and linkage was detected between the NCRNA and D10S1225, a marker in 10q21. To confirm linkage of NCRNA to chromosome 10q21 by homozygosity mapping, the patients and their carrier parents were genotyped for a number of other microsatellite polymorphic markers in the 10q21region. Statistically significant linkage was observed with multiple polymorphic markers in the 10q21 region. At straight theta = 0 with markers D10S1225, D10S1428, D10S1422, and D10S1418 maximum LOD scores of 3.74, 3.58, 3. 79, and 3.48 were generated, respectively. TDT P values for markers D10S1225 and D10S1418 were 0.0000021 and 0.000021, respectively.

Exclusion of AR-CHED from the chromosome 20 region containing the PPMD and AD-CHED loci.

Congenital hereditary endothelial dystrophy (CHED) is a disorder of the corneal endothelium and has been recognized to segregate in families with both autosomal dominant (AD) and autosomal recessive (AR) modes of transmission. AD-CHED has been previously linked to the pericentric region of chromosome 20. Posterior polymorphous dystrophy (PPMD), a corneal endothelial disorder showing phenotypic overlap with CHED, has also been previously genetically mapped to this region. The genetic interval containing AD-CHED is within the larger genetic interval containing the PPMD locus. This study sought to determine whether AR-CHED segregating in a consanguineous Saudi Arabian pedigree is linked to the previously mapped and overlapping loci for AD-CHED and PPMD on the pericentric region of chromosome 20. Forty members of a consanguineous Saudi Arabian pedigree segregating AR-CHED were ascertained. Short tandem-repeat polymorphic markers from the 20 cM interval on chromosome 20 containing both the PPMD and AD-CHED loci were used to genotype these individuals. LOD score analysis of the genotype data with the MENDEL software package utilizing a model of autosomal recessive inheritance with complete penetrance showed exclusion of CHED from the entire PPMD/AD-CHED interval by utilizing overlapping intervals of LOD scores of at least -2. The results obtained demonstrate that AR-CHED is not allelic to either AD-CHED or PPMD, although it has been proposed that AD-CHED may be allelic to PPMD. Thus, there are at least two genes responsible for CHED and PPMD.

Antenatal sonographic diagnosis of club foot with particular attention to the implications and outcomes of isolated club foot.

OBJECTIVE: There are no studies to date on the implications and outcomes of antenatally detected isolated club foot. The purpose of this study was to perform a contemporary evaluation of club foot diagnosed in the antenatal period. DESIGN: We performed a retrospective analysis of all ultrasound examinations performed in 1989-96 in the Fetal Diagnosis and Treatment Unit of the University of Iowa Hospitals and Clinics (n = 23,863). SUBJECTS AND METHODS: All cases of club foot (n = 35) were evaluated for the presence of other detectable abnormalities and karyotype results if available. Postnatal follow-up was performed until over 1 year of age. RESULTS: We diagnosed unilateral (n = 18) and bilateral (n = 17) club foot from 17.4 to 37.0 weeks. Defects involving other systems were found in 28 of 35 cases. Of the seven cases considered to be isolated antenatally, three were diagnosed with additional malformations in the neonatal period. CONCLUSION: Most cases of antenatally diagnosed club foot were not isolated. Even when they were thought to be isolated on antenatal ultrasound, over half of them were later found to be associated with additional severe abnormalities that were detectable only in the neonatal period.

The forkhead transcription factor gene FKHL7 is responsible for glaucoma phenotypes which map to 6p25.

A number of different eye disorders with the presence of early-onset glaucoma as a component of the phenotype have been mapped to human chromosome 6p25. These disorders have been postulated to be either allelic to each other or associated with a cluster of tightly linked genes. We have identified two primary congenital glaucoma (PCG) patients with chromosomal anomalies involving 6p25. In order to identify a gene involved in PCG, the chromosomal breakpoints in a patient with a balanced translocation between 6p25 and 13q22 were cloned. Cloning of the 6p25 breakpoint led to the identification of two candidate genes based on proximity to the breakpoint. One of these, FKHL7, encoding a forkhead transcription factor, is in close proximity to the breakpoint in the balanced translocation patient and is deleted in a second PCG patient with partial 6p monosomy. Furthermore, FKHL7 was found to harbour mutations in patients diagnosed with Rieger anomaly (RA), Axenfeld anomaly (AA) and iris hypoplasia (IH). This study demonstrates that mutations in FKHL7 cause a spectrum of glaucoma phenotypes.

Short tandem repeat polymorphic markers for the rat genome from marker-selected libraries.

In an effort to generate a genome-wide set of high-quality polymorphic markers for the rat, we used the marker-selection method, which has already been proven useful for the development of markers, especially for the human genome. Small-insert (300-900 bp) rat genomic libraries were constructed with an estimated complexity of three genome equivalents and enriched for short tandem repeat sequences (STRs). The enriched libraries were found to contain 45% (CA)n and 27% (GATA)n, representing at least a 50-fold enrichment over unselected small insert genomic libraries. A subset of 2160 STR-containing clones, primarily of the (GATA)n class of repeats, were sequenced. PCR primers flanking the repeats were synthesized from some of the sequences from the (CA)n and (GATA)n classes of STRs and tested for polymorphism in a panel of eight inbred rat strains. This strategy yielded 147 polymorphic markers, which mapped with high odds to all chromosomes by linkage in three F2 populations. The integration of these STR markers with other rat genetic markers and mapping reagents will facilitate the mapping of disease genes in the rat and the identification of loci associated with complex mammalian phenotypes.

Genomic organization, 5'-flanking region, and chromosomal localization of the human RGS3 gene.

RGS3 is the largest member of a recently discovered family of proteins (RGS proteins) that appear to function as negative regulators of heterotrimeric G-protein signaling. Seventeen mammalian RGS proteins have been identified by cloning or by comparison to expressed sequence tags, and several of these proteins have been shown recently to function as GTPase-activating proteins for G-protein alpha subunits. Despite the intense interest in RGS proteins as physiological regulators of G-protein signaling, there is little understanding of the structure and regulation of mammalian RGS genes. Using long-distance PCR, we amplified and characterized the entire coding and 5'-untranslated region of the human RGS3 gene. The coding region of the human RGS3 gene spans 14.7 kb and contains six exons, and the 5'-untranslated region spans 3.2 kb and contains two exons. Mapping of the exons revealed that the RGS domain, conserved among all RGS proteins, was encoded by three exons, while the unique amino-terminal domain of RGS3 was encoded by a single exon. Comparison of the location of the intron-exon boundaries of the human RGS3 gene to that of the human RGS2 gene, the only mammalian RGS gene described previously, revealed a remarkable similarity, providing the first conceptual support for a common ancestral mammalian RGS gene. 5'-RACE analysis was used to map the transcription start site 517 bp upstream of the translation start site, and anchored PCR was performed to amplify 1.0 kb of genomic DNA upstream of the transcription start site. Analysis of the 5'-flanking region revealed the presence of many potential regulatory elements, the presence of an initiator (Inr) element overlapping the transcription start site, and the absence of a TATA or a CCAAT box at the usual positions. By radiation hybrid mapping, the RGS3 gene was assigned to human chromosome 9q31-q33. This study is the first to elucidate the structure, chromosomal location, and regulatory sequences of the RGS3 gene, and it establishes the genetic basis for RGS3 gene research in humans.

Homozygosity mapping of achromatopsia to chromosome 2 using DNA pooling.

Achromatopsia is an autosomal recessive disease of the retina, characterized clinically by an inability to distinguish colors, impaired visual acuity, nystagmus and photophobia. A genome-wide search for linkage was performed using an inbred Jewish kindred from Iran. To facilitate the genome-wide search, we utilized a DNA pooling strategy which takes advantage of the likelihood that the disease in this inbred kindred is inherited by all affected individuals from a common founder. Equal molar amounts of DNA from all affected individuals were pooled and used as the PCR template for short tandem repeat polymorphic markers (STRPs). Pooled DNA from unaffected members of the kindred was used as a control. A reduction in the number of alleles in the affected versus control pool was observed at several loci. Upon genotyping of individual family members, significant linkage was established between the disease phenotype and markers localized on chromosome 2. The highest LOD score observed was 5.4 (theta = 0). When four additional small unrelated families were genotyped, the combined peak LOD score was 8.2. Analysis of recombinant chromosomes revealed that the disease gene lies within a 30 cM interval which spans the centromere. Additional fine-mapping studies identified a region of homozygosity in all affected individuals, narrowing the region to 14 cM. A candidate gene for achromatopsia was excluded from this disease interval by radiation hybrid mapping. Linkage of achromatopsia to chromosome 2 is an essential first step in the identification of the disease-causing gene.

Prenatal ultrasound findings in hydrolethalus: continuing difficulties in diagnosis.

We present the prenatal ultrasound findings in a case of hydrolethalus. This case illustrates ongoing problems in differentiating hydrolethalus, both pre- and postnatally, from other midline malformation syndromes including Pallister-Hall, Smith-Lemli-Opitz, pseudo-trisomy 13, oral facial-digital syndrome, and Meckel syndrome. Hydrolethalus can also be difficult to distinguish from certain skeletal dysplasias such as the short rib-polydactyly syndromes and campomelic dysplasia. Tests which can aid in diagnosis are presented.