Mbp Interval Research Articles

Genomewide linkage analysis in Costa Rican families implicates chromosome 15q14 as a candidate region for OCD

Obsessive compulsive disorder (OCD) has a complex etiology that encompasses both genetic and environmental factors. However, to date, despite the identification of several promising candidate genes and linkage regions, the genetic causes of OCD are largely unknown. The objective of this study was to conduct linkage studies of childhood-onset OCD, which is thought to have the strongest genetic etiology, in several OCD-affected families from the genetically isolated population of the Central Valley of Costa Rica (CVCR). The authors used parametric and non-parametric approaches to conduct genome-wide linkage analyses using 5,786 single nucleotide repeat polymorphisms (SNPs) in three CVCR families with multiple childhood-onset OCD-affected individuals. We identified areas of suggestive linkage (LOD score ≥ 2) on chromosomes 1p21, 15q14, 16q24, and 17p12. The strongest evidence for linkage was on chromosome 15q14 (LOD = 3.13), identified using parametric linkage analysis with a recessive model, and overlapping a region identified in a prior linkage study using a Caucasian population. Each CVCR family had a haplotype that co-segregated with OCD across a ~7 Mbp interval within this region, which contains 18 identified brain expressed genes, several of which are potentially relevant to OCD. Exonic sequencing of the strongest candidate gene in this region, the ryanodine receptor 3 (RYR3), identified several genetic variants of potential interest, although none co-segregated with OCD in all three families. These findings provide evidence that chromosome 15q14 is linked to OCD in families from the CVCR, and supports previous findings to suggest that this region may contain one or more OCD susceptibility loci.

Genome-wide study of familial juvenile hyperuricaemic (gouty) nephropathy (FJHN) indicates a new locus, FJHN3, linked to chromosome 2p22.1-p21

Familial juvenile hyperuricaemic (gouty) nephropathy (FJHN), is an autosomal dominant disease associated with a reduced fractional excretion of urate, and progressive renal failure. FJHN is genetically heterogeneous and due to mutations of three genes: uromodulin (UMOD), renin (REN) and hepatocyte nuclear factor-1beta (HNF-1β) on chromosomes 16p12, 1q32.1, and 17q12, respectively. However, UMOD, REN or HNF-1β mutations are found in only approximately 45% of FJHN probands, indicating the involvement of other genetic loci in approximately 55% of probands. To identify other FJHN loci, we performed a single nucleotide polymorphism (SNP)-based genome-wide linkage analysis, in six FJHN families in whom UMOD, HNF-1β and REN mutations had been excluded. Parametric linkage analysis using a 'rare dominant' model established linkage in five of the six FJHN families, with a LOD score >+3, at 0% recombination, between FJHN and SNPs at chromosome 2p22.1-p21. Analysis of individual recombinants in two unrelated affected individuals defined a approximately 5.5 Mbp interval, flanked telomerically by SNP RS372139 and centromerically by RS896986 that contained the locus, designated FJHN3. The interval contains 28 genes, and DNA sequence analysis of the most likely candidate, solute carrier family 8 member 1 (SLC8A1), did not identify any abnormalities in the FJHN3 probands. FJHN3 is likely located within a approximately 5.5 Mbp interval on chromosome 2p22.1-p21, and identifying the genetic abnormality will help to further elucidate mechanisms predisposing to gout and renal failure.

The PPCD1 Mouse: Characterization of a Mouse Model for Posterior Polymorphous Corneal Dystrophy and Identification of a Candidate Gene

The PPCD1 mouse, a spontaneous mutant that arose in our mouse colony, is characterized by an enlarged anterior chamber resulting from metaplasia of the corneal endothelium and blockage of the iridocorneal angle by epithelialized corneal endothelial cells. The presence of stratified multilayered corneal endothelial cells with abnormal patterns of cytokeratin expression are remarkably similar to those observed in human posterior polymorphous corneal dystrophy (PPCD) and the sporadic condition, iridocorneal endothelial syndrome. Affected eyes exhibit epithelialized corneal endothelial cells, with inappropriate cytokeratin expression and proliferation over the iridocorneal angle and posterior cornea. We have termed this the “mouse PPCD1” phenotype and mapped the mouse locus for this phenotype, designated “Ppcd1”, to a 6.1 Mbp interval on Chromosome 2, which is syntenic to the human Chromosome 20 PPCD1 interval. Inheritance of the mouse PPCD1 phenotype is autosomal dominant, with complete penetrance on the sensitive DBA/2J background and decreased penetrance on the C57BL/6J background. Comparative genome hybridization has identified a hemizygous 78 Kbp duplication in the mapped interval. The endpoints of the duplication are located in positions that disrupt the genes Csrp2bp and 6330439K17Rik and lead to duplication of the pseudogene LOC100043552. Quantitative reverse transcriptase-PCR indicates that expression levels of Csrp2bp and 6330439K17Rik are decreased in eyes of PPCD1 mice. Based on the observations of decreased gene expression levels, association with ZEB1-related pathways, and the report of corneal opacities in Csrp2bptm1a(KOMP)Wtsi heterozygotes and embryonic lethality in nulls, we postulate that duplication of the 78 Kbp segment leading to haploinsufficiency of Csrp2bp is responsible for the mouse PPCD1 phenotype. Similarly, CSRP2BP haploinsufficiency may lead to human PPCD.

Mutation of a Gene Essential for Ribosome Biogenesis, EMG1, Causes Bowen-Conradi Syndrome

Bowen-Conradi syndrome (BCS) is an autosomal-recessive disorder characterized by severely impaired prenatal and postnatal growth, profound psychomotor retardation, and death in early childhood. Nearly all reported BCS cases have been among Hutterites, with an estimated birth prevalence of 1/355. We previously localized the BCS gene to a 1.9 Mbp interval on human chromosome 12p13.3. The 59 genes in this interval were ranked as candidates for BCS, and 35 of these, including all of the best candidates, were sequenced. We identified variant NM_006331.6:c.400A-->G, p.D86G in the 18S ribosome assembly protein EMG1 as the probable cause of BCS. This mutation segregated with disease, was not found in 414 non-Hutterite alleles, and altered a highly conserved aspartic acid (D) residue. A structural model of human EMG1 suggested that the D86 residue formed a salt bridge with arginine 84 that would be disrupted by the glycine (G) substitution. EMG1 mRNA was detected in all human adult and fetal tissues tested. In BCS patient fibroblasts, EMG1 mRNA levels did not differ from those of normal cells, but EMG1 protein was dramatically reduced in comparison to that of normal controls. In mammalian cells, overexpression of EMG1 harboring the D86G mutation decreased the level of soluble EMG1 protein, and in yeast two-hybrid analysis, the D86G substitution increased interaction between EMG1 subunits. These findings suggested that the D-to-G mutation caused aggregation of EMG1, thereby reducing the level of the protein and causing BCS.

Dissection of a locus on mouse chromosome 5 reveals arthritis promoting and inhibitory genes

IntroductionIn a cross between two mouse strains, the susceptible B10.RIII (H-2r) and resistant RIIIS/J (H-2r) strains, a locus on mouse chromosome 5 (Eae39) was previously shown to control experimental autoimmune encephalomyelitis (EAE). Recently, quantitative trait loci (QTL), linked to disease in different experimental arthritis models, were mapped to this region. The aim of the present study was to investigate whether genes within Eae39, in addition to EAE, control development of collagen-induced arthritis (CIA).MethodsCIA, induced by immunisation with bovine type II collagen, was studied in Eae39 congenic and sub-interval congenic mice. Antibody titres were investigated with ELISA. Gene-typing was performed by micro-satellite mapping and statistics was calculated by standard methods.ResultsExperiments of CIA in Eae39 congenic- and sub-interval congenic mice, carrying RIIIS/J genes on the B10.RIII genetic background, revealed three loci within Eae39 that control disease and anti-collagen antibody titres. Two of the loci promoted disease and the third locus was protected against CIA development. By further breeding of mice with small congenic fragments, we identified a 3.2 mega base pair (Mbp) interval that regulates disease.ConclusionsDisease-promoting and disease-protecting genes within the Eae39 locus on mouse chromosome 5 control susceptibility to CIA. A disease-protecting locus in the telomeric part of Eae39 results in lower anti-collagen antibody responses. The study shows the importance of breeding sub-congenic mouse strains to reveal genetic effects on complex diseases.

A Defect in the TUSC3 Gene Is Associated with Autosomal Recessive Mental Retardation

Recent studies have shown that autosomal recessive mental retardation (ARMR) is extremely heterogeneous, and there is reason to believe that the number of underlying gene defects goes into the thousands. To date, however, only four genes have been implicated in nonsyndromic ARMR (NS-ARMR): PRSS12 (neurotrypsin), CRBN (cereblon), CC2D1A, and GRIK2. As part of an ongoing systematic study aiming to identify ARMR genes, we investigated a large consanguineous family comprising seven patients with nonsyndromic ARMR in four sibships. Genome-wide SNP typing enabled us to map the relevant genetic defect to a 4.6 Mbp interval on chromosome 8. Haplotype analyses and copy-number studies led to the identification of a homozygous deletion partly removing TUSC3 (N33) in all patients. All obligate carriers of this family were heterozygous, but none of 192 unrelated healthy individuals from the same population carried this deletion. We excluded other disease-causing mutations in the coding regions of all genes within the linkage interval by sequencing; moreover, we verified the complete absence of a functional TUSC3 transcript in all patients through RT-PCR. TUSC3 is thought to encode a subunit of the endoplasmic reticulum-bound oligosaccharyltransferase complex that catalyzes a pivotal step in the protein N-glycosylation process. Our data suggest that in contrast to other genetic defects of glycosylation, inactivation of TUSC3 causes nonsyndromic MR, a conclusion that is supported by a separate report in this issue of AJHG. TUSC3 is only the fifth gene implicated in NS-ARMR and the first for which mutations have been reported in more than one family.

Ein neuer Genort für eine autosomal dominante, nichtsyndromale Hörstörung (DFNA57) kartiert auf Chromosom 19p13.2 und überlappt mit DFNB15

Non-syndromic hearing loss is the most genetically heterogeneous trait known in humans. To date, 54 loci for autosomal dominant non-syndromic sensorineural hearing loss (NSSHL) have been identified by linkage analysis. In this study a German pedigree has been identified segregating a progressive bilateral loss of lower and middle frequencies. A genome-wide screening and linkage analysis revealed the existence of a new NSSHL locus (DFNA57). The phenotype was mapped to a 10 degrees Mbp interval on chromosome 19p13.2 from 7.8 to 18.2 degrees Mbp, a maximum 2-point LOD score of 3.08 was obtained for the marker D19S586. The region overlaps with the recessive locus DFNB15. The results underline the heterogeneity of hereditary hearing disorders. Identification of genes can help to reach a better understanding of the molecular mechanism of hearing.

Positional Cloning: Regulation of Hematopoietic Stem and Progenitor Cell Mobilization through EGF Receptor Signaling.

There is significant individual variation in humans in their ability to mobilize hematopoietic stem and progenitor cells (HSPC) from bone marrow (BM) to peripheral blood upon G-CSF stimulation. Understanding the molecular mechanism underlying this variation may provide clinically significant targets to improve HSPC mobilization. Using forward genetics, we have previously demonstrated that a locus on chromosome 11 is responsible for murine inter-strain variation in G-CSF mediated HSPC mobilization (Geiger et. al., Exp. Hematol., 2004). Novel subcongenic animals were generated that further confined the locus to a 5 Mbp interval. Of the 12 genes in this interval, epidermal growth factor receptor (EGFR) was identified as a candidate gene for regulating inter-strain differences in mobilization. Using real-time PCR, we demonstrated that EGFR is expressed in both murine and human HSPC and that the level of EGFR expression is inversely correlated with GCSF-induced mobilization proficiency of murine HSPC. To further demonstrate a role of EGFR signaling in mobilization, C57BL/6 mice were treated with G-CSF and increasing concentrations of murine recombinant epidermal growth factor (EGF). Our results demonstrated that a single dose of EGF (0.8 ug/g) significantly inhibited mobilization efficiency (approx. 4-fold) (46.7±13 CFC/37.5ul PB with G-CSF treated mice vs. 12.67±1.5 CFC/37.5ul PB with G-CSF + EGF treated mice). To determine whether the reduction in mobilization efficiency is dependent upon EGFR activity, EGFR mutant mice (waved-2+/−) were treated with G-CSF and EGF. In contrast to control mice, EGF had no significant inhibitory effect on G-CSF-mediated mobilization in waved-2+/− mice indicating that EGFR signaling is necessary for inhibition of mobilization by EGF (46.7±13.0 vs. 12.67±1.5 CFC/37.5ul PB, G-CSF vs G-CSF+EGF treated control mice compared to 14.2±3.7 vs. 12.1±3.6 CFC/37.5ul PB with waved-2+/− mice). To determine if EGFR-signaling regulated mobilization is HSC intrinsic, we performed “competitive mobilization experiments” in which equal numbers of BM cells (4x106) from either control or waved-2−/− mice and competitor Ly5.1 cells were transplanted into lethally irradiated BoyJ mice and subsequently treated with G-CSF and EGF. Animals transplanted with control cells demonstrated a significant reduction in mobilization efficiency in response to EGF compared to animals transplanted with waved-2−/− cells (20.2±7.2 vs. 7.89±1.2 CFC/37.5ul PB, G-CSF vs G-CSF+EGF treated control transplanted mice compared to 20.3±4.5 vs. 18.67±1.2 CFC/37.5ul PB with waved-2−/− transplanted mice) indicating that EGFR activity in hematopoietic cells is necessary to confer inhibition of mobilization efficiency by EGF. Taken together, these findings identify EGFR signaling as a negative regulator and a genetic modifier of G-CSF induced mobilization proficiency and a potential novel target to enhance HSPC mobilization efficiency.

Mutations in the UBIAD1 Gene, Encoding a Potential Prenyltransferase, Are Causal for Schnyder Crystalline Corneal Dystrophy

Schnyder crystalline corneal dystrophy (SCCD, MIM 121800) is a rare autosomal dominant disease characterized by progressive opacification of the cornea resulting from the local accumulation of lipids, and associated in some cases with systemic dyslipidemia. Although previous studies of the genetics of SCCD have localized the defective gene to a 1.58 Mbp interval on chromosome 1p, exhaustive sequencing of positional candidate genes has thus far failed to reveal causal mutations. We have ascertained a large multigenerational family in Nova Scotia affected with SCCD in which we have confirmed linkage to the same general area of chromosome 1. Intensive fine mapping in our family revealed a 1.3 Mbp candidate interval overlapping that previously reported. Sequencing of genes in our interval led to the identification of five putative causal mutations in gene UBIAD1, in our family as well as in four other small families of various geographic origins. UBIAD1 encodes a potential prenyltransferase, and is reported to interact physically with apolipoprotein E. UBIAD1 may play a direct role in intracellular cholesterol biochemistry, or may prenylate other proteins regulating cholesterol transport and storage.

Adult‐onset autosomal dominant leukodystrophy with autonomic symptoms restricted to 1.5 Mbp on chromosome 5q23

Autosomal dominant leukodystrophy (ADLD) with autonomic symptoms is a slowly progressive leukodystrophy with an onset in the 4th to 6th decade of life. Early symptoms are derived from the autonomic nervous system with bladder dysfunction in the majority of patients. The disease progresses slowly with loss of fine motor skills, ataxia, and affected individuals may survive for two decades after onset of symptoms. The molecular basis behind ADLD remains unknown but the causative locus was previously mapped to a 4 cM region on chromosome 5. We have recently identified a large family of Swedish origin with this type of ADLD. Linkage analysis on samples from family members confirmed linkage to 5q23 and supports genetic homogeneity for the disease. We fine mapped and localized the ADLD gene to a 0.96 cM region between D5S1495 and CTT/CCT15. A maximum parametric multipoint location score of 9.45 was obtained for a position at the marker CTT55. From our results we conclude that the ADLD gene locus is restricted to a 1.47 Mbp interval containing 13 known or putative genes.

Human squamous cell carcinomas lose a mortality gene from chromosome 6q14.3 to q15.

Normal human keratinocytes possess a finite replicative lifespan. Most advanced squamous cell carcinomas (SCCs), however, are immortal, a phenotype that is associated with p53 and INK4A dysfunction, high levels of telomerase and loss of heterozygosity (LOH) at several genetic loci, suggestive of the dysfunction of other mortality genes. We show here that human chromosome 6 specifically reduces the proliferation or viability of a human SCC line, BICR31, possessing LOH across the chromosome. This was determined by an 88% reduction in colony yield (P<0.001), following the reintroduction of an intact normal chromosome 6 by monochromosome transfer. Deletion analysis of immortal segregants using polymorphic markers revealed the loss of a 2.9 Mbp interval, centred on marker D6S1045 at 6q14.3-q15, in 6/19 segregants. Crucially, allelic losses of this region were not identified in control hybrids constructed between chromosome 6 and the BICR6 SCC cell line that is heterozygous for chromosome 6 and which showed no reduction in colony formation relative to the control chromosome transfers. This indicates that the minimally deleted region at D6S1045 is not the result of fragile sites, a recombination hot spot, or a feature of the monochromosome transfer technique. LOH of D6S1045 was found in 2/9 immortal SCC lines and was part of a minimally deleted region of line BICR19. Furthermore, allelic imbalance, consistent with LOH, was detected in 3/17 advanced SCCs of the tongue. These results suggest the existence of a suppressor of SCC immortality and tumour development at chromosome 6q14.3-q15, which is important to a subset of human SCCs.

Refined mapping of the gene encoding the p127 kDa UV-damaged DNA-binding protein (DDB1) within 11q12-q13.1 and its exclusion in Best's vitelliform macular dystrophy.

Best's vitelliform macular dystrophy (Best's disease) is an autosomal dominant disorder of unknown causes and is typically characterised by an accumulation of lipofuscin-like material in the subretinal space of the macula. The disease gene has been localised to chromosome 11q12-13.1 within a 1.4 Mbp interval flanked by markers at D11S1765 and uteroglobin (UGB). Here we report the refined mapping of the gene encoding the p127 kDa subunit (DDB1) of a UV damage-specific DNA binding protein within the D11S1765-UGB region. Northern blot analysis demonstrates an abundant expression of the DDB1 transcript in the retina suggesting a functional role for DDB1 in this tissue. These considerations together with the chromosomal localisation have led us to evaluate the possible involvement of DDB1 in the pathogenesis of Best's disease.