Different RPGR exon ORF15 mutations in Canids provide insights into photoreceptor cell degeneration.
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The canine disease, X-linked progressive retinal atrophy (XLPRA), is similar to human RP3, an X-linked form of retinitis pigmentosa, and maps to the same region in the X chromosome. Analysis of the physical map of the XLPRA and RP3 intervals shows a high degree of conservation in terms of genes and their order. We have found different mutations in exon ORF15 of the RPGR gene in two distinct mutant dog strains (XLPRA1, XLPRA2). Microdeletions resulting in a premature stop or a frameshift mutation result in very different retinal phenotypes, which are allele-specific and consistent for each mutation. The phenotype associated with the frameshift mutation in XLPRA2 is very severe and manifests during retinal development; the phenotype resulting from the XLPRA1 nonsense mutation is expressed only after normal photoreceptor morphogenesis. Splicing of RPGR mRNA transcripts in retina is complex, and either exon ORF15 or exon 19 can be a terminal exon. The retina-predominant transcript contains ORF15 as a terminal exon, and is expressed in normal and mutant retinas. The frameshift mutation dramatically alters the deduced amino acid sequence, and the protein aggregates in the endoplasmic reticulum of transfected cells. The cellular and molecular results in the two canine RPGR exon ORF15 mutations have implications for understanding the phenotypic variability found in human RP3 families that carry similar mutations. (+info)
Foxe3 haploinsufficiency in mice: a model for Peters' anomaly.
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PURPOSE: To evaluate the importance in anterior segment dysgenesis of genetic variation in Foxe3, a gene encoding a forkhead transcription factor specifically expressed in the lens. METHODS: The phenotype of mice heterozygous for a mutation in the DNA-binding domain of Foxe3 was examined from histologic sections, and DNA binding by the encoded protein was investigated by gel-shift assay. FOXE3 from human patients with Peters' anomaly was PCR amplified and sequenced. RESULTS: The dysgenetic lens (dyl) allele of Foxe3 was found to encode a protein unable to bind DNA. Approximately 40% of mice heterozygous for Foxe3(dyl) have corneal and lenticular defects. The phenotype is variable but typically consists of the equivalent of Peters' anomaly in humans, with central corneal opacity, keratolenticular adhesion, and, in some cases, anterior polar cataract. In a small cohort (n = 13) of patients with Peters' anomaly, shown to be normal in the PAX6 locus, one individual was found to be heterozygous for a nonconservative missense mutation in FOXE3. The mutation, which does not occur in 116 chromosomes from a control population, substitutes leucine for arginine 90 at a highly conserved position in the forkhead domain. CONCLUSIONS: Haploinsufficiency of Foxe3 in a mouse model causes anterior segment dysgenesis similar to Peters' anomaly. Although causality could not be shown in the human case, the presence of a rare, nonconservative substitution in FOXE3 of a patient with Peters' anomaly is interesting, in light of the phenotypic similarities with the mutant mice. (+info)
Systematic evaluation of map quality: human chromosome 22.
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Marker positions on nine genetic linkage, radiation hybrid, and integrated maps of human chromosome 22 were compared with their corresponding positions in the completed DNA sequence. The proportion of markers whose map position is <250 kb from their respective sequence positions ranges from 100% to 35%. Several discordant markers were identified, as well as four regions that show common inconsistencies across multiple maps. These shared discordant regions surround duplicated DNA segments and may indicate mapping or assembly errors due to sequence homology. Recombination-rate distributions along the chromosome were also evaluated, with male and female meioses showing significantly different patterns of recombination, including an 8-Mb male recombination desert. The distributions of radiation-induced chromosome breakage for the GB4 and the G3 radiation hybrid panels were also evaluated. Both panels show fluctuations in breakage intensity, with different regions of significantly elevated rates of breakage. These results provide support for the common assumption that radiation-induced breaks are generally randomly distributed. The present studies detail the limitations of these important map resources and should prove useful for clarifying potential problems in the human maps and sequence assemblies, as well as for mapping and sequencing projects in and across other species. (+info)
The LGI1 gene involved in lateral temporal lobe epilepsy belongs to a new subfamily of leucine-rich repeat proteins.
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Recently mutations in the LGI1 (leucine-rich, glioma-inactivated 1) gene have been found in human temporal lobe epilepsy. We have now identified three formerly unknown LGI-like genes. Hydropathy plots and pattern analysis showed that LGI genes encode proteins with large extra- and intracellular domains connected by a single transmembrane region. Sequence analysis demonstrated that LGI1, LGI2, LGI3, and LGI4 form a distinct subfamily when compared to other leucine-rich repeat-containing proteins. In silico mapping and radiation hybrid experiments assigned LGI2, LGI3, and LGI4 to different chromosomal regions (4p15.2, 8p21.3, 19q13.11), some of which have been implicated in epileptogenesis and/or tumorigenesis. (+info)
Conserved synteny in rat and mouse for a blood pressure QTL on human chromosome 17.
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Evidence for blood pressure quantitative trait loci (QTLs) on rat chromosome 10 has been found in multiple independent studies. Analysis of the homologous region on human chromosome 17 revealed significant linkage to blood pressure. The critical segment on human chromosome 17 spans a large interval containing the genes Itga2b, Gfap, and Itgb3. Therefore, findings in the rat may help to refine the position of blood pressure-regulating loci, assuming a common molecular cause across species. However, it has recently been suggested that the gene order in human, rat, and mouse is not conserved in this region, leaving uncertainty about the overlap of the blood pressure- regulating region between human chromosome 17 and rat chromosome 10. We have performed a detailed comparative analysis among human, mouse, and rat, defining the segment in question, by obtaining gene structure information in silico and by radiation hybrid mapping. It is of interest that this region also contains Wnk4, a gene previously identified to cause pseudohypoaldosteronism type II and human hypertension. Our results definitively show that the conserved synteny extends among human chromosome 17, rat chromosome 10, and mouse chromosome 11, demonstrating an overlap between previously localized blood pressure QTLs in humans and rats. (+info)
A higher resolution radiation hybrid map of bovine chromosome 13.
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In this paper, we present a radiation hybrid framework map of BTA13 composed of nine microsatellite loci, six genes and one EST. The map has been developed using a recently constructed 12'000 rad bovine-hamster whole-genome radiation hybrid panel. Moreover, we present a comprehensive map of BTA13 comprising 72 loci, of which 45 are microsatellites, 20 are genes and seven are ESTs. The map has an estimated length of 2694.7 cR(12'000). The proposed order is in general agreement with published maps of BTA13. Our results only partially support previously published information of five blocks of conserved gene order between cattle and man. We found no evidence for the existence of an HSA20 homologous segment of coding DNA on BTA13 located centromeric of a confirmed HSA10 homologous region. The present map increases the marker density and the marker resolution on BTA13 and enables further insight into the evolutionary development of the chromosome as compared to man. (+info)
Physical mapping of genes in somatic cell radiation hybrids by comparative genomic hybridization to cDNA microarrays.
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BACKGROUND: Somatic cell mutants can be informative in the analysis of a wide variety of cellular processes. The use of map-based positional cloning strategies in somatic cell hybrids to analyze genes responsible for recessive mutant phenotypes is often tedious, however, and remains a major obstacle in somatic cell genetics. To fulfill the need for more efficient gene mapping in somatic cell mutants, we have developed a new DNA microarray comparative genomic hybridization (array-CGH) method that can rapidly and efficiently map the physical location of genes complementing somatic cell mutants to a small candidate genomic region. Here we report experiments that establish the validity and efficacy of the methodology. RESULTS: CHO cells deficient for hypoxanthine:guanine phosphoribosyl transferase (HPRT) were fused with irradiated normal human fibroblasts and subjected to HAT selection. Cy5-labeled genomic DNA from the surviving hybrids containing the HPRT gene was mixed with Cy3-labeled genomic DNA from normal CHO cells and hybridized to a microarray containing 40,185 cDNAs, representing 29,399 genes (UniGene clusters). The DNA spots with the highest Cy5:Cy3 fluorescence ratios corresponded to a group of genes mapping within a 1 Mb interval centered near position 142.7 Mb on the X chromosome, the genomic location of HPRT. CONCLUSION: The results indicate that our physical mapping method based on radiation hybrids and array-CGH should significantly enhance the speed and efficiency of positional cloning in somatic cell genetics. (+info)
Expressed sequence tag analysis of human RPE/choroid for the NEIBank Project: over 6000 non-redundant transcripts, novel genes and splice variants.
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PURPOSE: The retinal pigment epithelium (RPE) and choroid comprise a functional unit of the eye that is essential to normal retinal health and function. Here we describe expressed sequence tag (EST) analysis of human RPE/choroid as part of a project for ocular bioinformatics. METHODS: A cDNA library (cs) was made from human RPE/choroid and sequenced. Data were analyzed and assembled using the program GRIST (GRouping and Identification of Sequence Tags). Complete sequencing, Northern and Western blots, RH mapping, peptide antibody synthesis and immunofluorescence (IF) have been used to examine expression patterns and genome location for selected transcripts and proteins. RESULTS: Ten thousand individual sequence reads yield over 6300 unique gene clusters of which almost half have no matches with named genes. One of the most abundant transcripts is from a gene (named "alpha") that maps to the BBS1 region of chromosome 11. A number of tissue preferred transcripts are common to both RPE/choroid and iris. These include oculoglycan/opticin, for which an alternative splice form is detected in RPE/choroid, and "oculospanin" (Ocsp), a novel tetraspanin that maps to chromosome 17q. Antiserum to Ocsp detects expression in RPE, iris, ciliary body, and retinal ganglion cells by IF. A newly identified gene for a zinc-finger protein (TIRC) maps to 19q13.4. Variant transcripts of several genes were also detected. Most notably, the predominant form of Bestrophin represented in cs contains a longer open reading frame as a result of splice junction skipping. CONCLUSIONS: The unamplified cs library gives a view of the transcriptional repertoire of the adult RPE/choroid. A large number of potentially novel genes and splice forms and candidates for genetic diseases are revealed. Clones from this collection are being included in a large, nonredundant set for cDNA microarray construction. (+info)