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Heterozygous
Kinds of Heterozygous Terms modified by Heterozygous Selected AbstractsEctopic dermal ridge configurations on the interdigital webbings and postaxial marginal portion of the hindlimb in Hammertoe mutant mice (Hm),JOURNAL OF MORPHOLOGY, Issue 10 2008Sumiko Kimura Abstract The effects of the hereditary malformation of Hammertoe mutant mice (gene symbol Hm) on the digital pads and dermal ridge configurations on their hindlimbs were examined. In the wild-type (+/+) mice with normally separated digits, dermal ridges developed only on the pads. Heterozygous (Hm/+) and homozygous (Hm/Hm) mutant mice, however, had a broad big toe, fused interdigital soft tissues, reduced claws, an extra rudimentary postaxial digit and camptodactyly. The dermal ridges appeared not only on the pads, affected in their number and configurations, but also on the ventral surface of the interdigital webbings and postaxial marginal area exhibiting an extra rudimentary digit and webbing. These aberrant configurations may be related to the abnormal occurrence of programmed cell death (PCD) in the interdigital zones and the postaxial marginal portion in Hm/+ and Hm/Hm mice. That is, the diminished cell death may fail to decrease the cell density in the interdigital zones and postaxial marginal portion and result in the webbing and an extra rudimentary digit and webbing, respectively. Simultaneously, it could also interrupt the migration of surviving cells of these areas toward the neighboring digits and the distal area of the sole and produce the ectopic dermal ridges on the way to the as yet unformed (presumptive) digital and plantar volar pads. The present findings suggest that normal interdigital and pre/postaxial PCD contributes not only to the separation of digits, the initial formation of individual digits of different sizes, and the inhibition of the extra digit but also to the development of the presumptive digital and plantar pads, including dermal ridges. J. Morphol., 2008. © 2008 Wiley-Liss, Inc. [source] X-Irradiation Induces Up-regulation of ATM Gene Expression in Wild-type Lymphoblastoid Cell Lines, but Not in Their Heterozygous or Homozygous Ataxia-telangiectasia CounterpartsCANCER SCIENCE, Issue 6 2001Yuko Hirai Ataxia-telangiectasia (AT) is an autosomal recessive disease. The relevant gene has been cloned and designated ATM. We studied the expression of both ATM mRNA and the ATM protein in unirradi-ated and X-irradiated EBV (Epstein-Barr virus)-transformed lymphoblastoid cell lines (LCLs) derived from donors who were normal (ATM+/+), AT heterozygotes (ATM+/,), or AT homozy-gotes (ATM,/,), respectively. In ATM+/+ LCLs, the levels of ATM mRNA were found to have increased by approximately 1.5-fold within 1 h of exposure to 10 Gy of X-rays, while the ATM protein levels had increased by 1.5- to 2.0-fold within 2 to 3 h of irradiation. The wild-type mRNA and protein levels both returned to their basal values fairly quickly after this tune. The results obtained with the ATM+/, LCLs were quite different, however: neither the mRNA nor protein levels were found to have increased as a consequence of X-irradiation in any ATM+/, LCL. Twelve of the mutations in the ATM,/, LCLs we used were truncating mutations, and we suspected that the corresponding truncated ATM proteins would be too labile to be detected by western blot analysis. However, five of the ATM,/, LCLs produced mutant ATM proteins that were identical in molecular weight to the wild-type ATM protein. When cells from three of these five clones were exposed to X-rays, transcription of the mutant ATM genes appeared to reduce somewhat, as were the levels of protein being produced. These results suggest that the normal ATM gene responds to ionizing radiation by up-regulating its activity, whereas none of the mutant ATM genes we studied were able to respond in this way. [source] EndothelinA (ETA) and ETB receptor-mediated regulation of nitric oxide synthase 1 (NOS1) and NOS3 isoforms in the renal inner medullaACTA PHYSIOLOGICA, Issue 4 2007J. C. Sullivan Abstract Aim:, Our laboratory and others have shown that endothelin (ET)-1 directly stimulates nitric oxide (NO) production in inner medullary collecting duct (IMCD) cells. The goal of this study was to determine which NO synthase (NOS) isoforms in IMCD are sensitive to ET-1, and the role of ETA and ETB receptor activation in vivo and in vitro. Methods:, NOS enzymatic activity and NOS isoform protein expression were examined in cultured IMCD-3 cells and isolated renal inner medulla. ETB receptor-deficient homozygous rats (sl/sl) have elevated levels of circulating ET-1 and lack a functional ETB signalling pathway in kidneys, and furthermore provides a unique model to study ETA receptor signalling in the renal inner medulla in vivo. Results:, Incubation of IMCD-3 cells with exogenous ET-1 (50 nm) resulted in ETA -dependent increased NOS1 protein expression in IMCD-3 cells with no effect on NOS2 or NOS3 expression. ETB receptor antagonism has no effect on NOS expression in IMCD-3 cells. Consistent with in vitro results, cytosolic NOS1 protein expression was significantly greater in the renal inner medulla of sl/sl rats compared with heterozygous (sl/+) controls, with no alteration in NOS3 expression. In contrast to protein expression data, NOS1- and NOS3-specific enzymatic activities decreased in the cytosolic fraction from the renal inner medulla of sl/sl compared with sl/+. Conclusion:, These results provide evidence that both ETA and ETB receptors regulate NOS isoform activity in the renal inner medulla and specifically support the hypothesis that ETA receptor activation increases NOS1 expression. [source] Generation and expression of a Hoxa11eGFP targeted allele in miceDEVELOPMENTAL DYNAMICS, Issue 11 2008Lisa T. Nelson Abstract Hox genes are crucial for body axis specification during embryonic development. Hoxa11 plays a role in anteroposterior patterning of the axial skeleton, development of the urogenital tract of both sexes, and proximodistal patterning of the limbs. Hoxa11 expression is also observed in the neural tube. Herein, we report the generation of a Hoxa11eGFP targeted knock-in allele in mice in which eGFP replaces the first coding exon of Hoxa11 as an in-frame fusion. This allele closely recapitulates the reported mRNA expression patterns for Hoxa11. Hoxa11eGFP can be visualized in the tail, neural tube, limbs, kidneys, and reproductive tract of both sexes. Additionally, homozygous mutants recapitulate reported phenotypes for Hoxa11 loss of function mice, exhibiting loss of fertility in both males and females. This targeted mouse line will prove useful as a vital marker for Hoxa11 protein localization during control (heterozygous) or mutant organogenesis. Developmental Dynamics 237:3410,3416, 2008. © 2008 Wiley-Liss, Inc. [source] Potential roles for BMP and Pax genes in the development of iris smooth muscleDEVELOPMENTAL DYNAMICS, Issue 2 2005Abbie M. Jensen Abstract The embryonic optic cup generates four types of tissue: neural retina, pigmented epithelium, ciliary epithelium, and iris smooth muscle. Remarkably little attention has focused on the development of the iris smooth muscle since Lewis ([1903] J. Am. Anat. 2:405,416) described its origins from the anterior rim of the optic cup neuroepithelium. As an initial step toward understanding iris smooth muscle development, I first determined the spatial and temporal pattern of the development of the iris smooth muscle in the chick by using the HNK1 antibody, which labels developing iris smooth muscle. HNK1 labeling shows that iris smooth muscle development is correlated in time and space with the development of the ciliary epithelial folds. Second, because neural crest is the only other neural tissue that has been shown to generate smooth muscle (Le Lievre and Le Douarin [1975] J. Embryo. Exp. Morphol. 34:125,154), I sought to determine whether iris smooth muscle development shares similarities with neural crest development. Two members of the BMP superfamily, BMP4 and BMP7, which may regulate neural crest development, are highly expressed by cells at the site of iris smooth muscle generation. Third, because humans and mice that are heterozygous for Pax6 mutations have no irides (Hill et al. [1991] Nature 354:522,525; Hanson et al. [1994] Nat. Genet. 6:168,173), I determined the expression of Pax6. I also examined the expression of Pax3 in the developing anterior optic cup. The developing iris smooth muscle coexpresses Pax6 and Pax3. I suggest that some of the eye defects caused by mutations in Pax6, BMP4, and BMP7 may be due to abnormal iris smooth muscle. Developmental Dynamics 232:385,392, 2005. © 2004 Wiley-Liss, Inc. [source] Growth defect in Grg5 null mice is associated with reduced Ihh signaling in growth platesDEVELOPMENTAL DYNAMICS, Issue 1 2002Wen-Fang Wang Abstract Gene-targeted disruption of Grg5, a mouse homologue of Drosophila groucho (gro), results in postnatal growth retardation in mice. The growth defect, most striking in approximately half of the Grg5 null mice, occurs during the first 4,5 weeks of age, but most mice recover retarded growth later. We used the nonlinear mixed-effects model to fit the growth data of wild-type, heterozygous, and Grg5 null mice. On the basis of preliminary evidence suggesting an interaction between Grg5 and the transcription factor Cbfa1/Runx2, critical for skeletal development, we further investigated the skeleton in the mice. A long bone growth plate defect was identified, which included shorter zones of proliferative and hypertrophic chondrocytes and decreased trabecular bone formation. This decreased trabecular bone formation is likely caused by a reduced recruitment of osteoblasts into the growth plate region of Grg5 null mice. Like the growth defect, the growth plate and trabecular bone abnormality improved as the mice grew older. The growth plate defect was associated with reduced Indian hedgehog expression and signaling. We suggest that Grg5, a transcriptional coregulator, modulates the activities of transcription factors, such as Cbfa1/Runx2 in vivo to affect Ihh expression and the function of long bone growth plates. © 2002 Wiley-Liss, Inc. [source] Cerebral palsy in siblings caused by compound heterozygous mutations in the gene encoding protein CDEVELOPMENTAL MEDICINE & CHILD NEUROLOGY, Issue 5 2010CHOONG YI FONG We report two sisters with extensive bilateral periventricular haemorrhagic infarction (PVHI) causing cerebral palsy (CP). The older sister presented at 20 months with cortical visual blindness, spastic diplegia, and purpura fulminans. The younger sister presented aged 3 days old with apnoeas and multifocal seizures. She subsequently had global developmental delay, cortical visual blindness, spastic quadriplegia, epilepsy, and purpura fulminans at age 2 years. Neuroimaging of both siblings showed bilateral PVHI consistent with bilateral cerebral intramedullary venous thrombosis occurring at under 28 weeks' gestation for the older sister and around time of birth for the younger sister. At latest follow-up, the older sister (13y) has spastic diplegia at Gross Motor Function Classification System (GMFCS) level II, and the younger sister (10y) has spastic quadriplegia at GMFCS level IV. Both sisters showed partial quantitative reduction in plasma protein C antigen and severe qualitative reduction in plasma protein C anticoagulant activity. They were heterozygous for two independent mutations in the protein C gene (PROC). There was no other risk factor for CP. To our knowledge, this is the first family reported with compound heterozygous PROC mutations as the likely genetic cause of familial CP. This report adds to the list of known monogenic causes of CP. [source] Homozygous thermolabile variant of the methylenetetrahy-drofolate reductase gene: a potential risk factor for hyperhomo-cysteinaemia, CVD, and stroke in childhoodDEVELOPMENTAL MEDICINE & CHILD NEUROLOGY, Issue 4 2001Mara Prengler In this study of 118 children (median age 5.1 years; range 6 months to 17 years) with ischaemic stroke or transient ischaemic attack (TIA), 22 children (19%) were homozygous for the thermolabile variant of the methylenetetrahydrofolate reductase allele (t-MTHFR), compared with nine of 78 (12%) of a reference population (p=0.18, OR 1.76, 95% CI 0.76 to 4.04). Of those with cerebrovascular disease (CVD), 17 of 84 were homozygous for the t-MTHFR allele (p=0.13 compared with the reference population (OR 1.95, 95% CI 0.81 to 4.65). There was a significant (p<0.025) increment of plasma total homocysteine concentration in homozygotes for the t-MTHFR allele compared with heterozygotes, negatives for the t-MTHFR allele, and control children with no history of stroke. In four of 12 homozygotes for the t-MTHFR allele, plasma homocysteine levels were raised, compared with three of 38 of those who were negative or heterozygous (p=0.047; OR 5.8, 95% CI 1.1 to 31.2). Homozygotes for the t-MTHFR allele were significantly more likely to have a recurrent event than those who were negative or heterozygous (Cox regression p=0.031, hazard ratio 2.18, 95% CI 1.08 to 4.42). These data suggest that homozygosity for the t-MTHFR allele is associated with raised homocysteine levels in children and is a risk factor for primary and secondary stroke and TIA. [source] Reelin is essential for neuronal migration but not for radial glial elongation in neonatal ferret cortex,DEVELOPMENTAL NEUROBIOLOGY, Issue 5 2008Alisa Schaefer Abstract Numerous functions related to neuronal migration are linked to the glycoprotein reelin. Reelin also elongates radial glia, which are disrupted in mutant reeler mice. Our lab developed a model of cortical dysplasia in ferrets that shares features with the reeler mouse, including impaired migration of neurons into the cerebral cortex and disrupted radial glia. Explants of normal ferret cortex in coculture with dysplastic ferret cortex restore the deficits in this model. To determine if reelin is integral to the repair, we used explants of P0 mouse cortex either of the wild type (WT) or heterozygous (het) for the reelin gene, as well as P0 reeler cortex (not containing reelin), in coculture with organotypic cultures of dysplastic ferret cortex. This arrangement revealed that all types of mouse cortical explants (WT, het, reeler) elongated radial glia in ferret cortical dysplasia, indicating that reelin is not required for proper radial glial morphology. Migration of cells into ferret neocortex, however, did not improve with explants of reeler cortex, but was almost normal after pairing with WT or het explants. We also placed an exogenous source of reelin in ferret cultures at the pial surface to reveal that migrating cells move toward the reelin source in dysplastic cortex; radial glia in these cultures were also improved toward normal. Our results demonstrate that the normotopic position of reelin is important for proper neuronal positioning, and that reelin is capable of elongating radial glial cells but is not the only radialization factor. © 2008 Wiley Periodicals, Inc. Develop Neurobiol, 2008. [source] Caucasian patients with type 2 diabetes mellitus have elevated levels of monocyte chemoattractant protein-1 that are not influenced by the ,2518 A,G promoter polymorphismDIABETES OBESITY & METABOLISM, Issue 5 2005B. Zietz Aim:, To investigate the association of serum levels and the ,2518 A,G promoter polymorphism of the gene for chemokine monocyte chemoattractant protein-1 (MCP-1), a major chemoattractant of monocytes and activated lymphocytes, with metabolic parameters as well as insulin, leptin and the cytokines tumour necrosis factor-, (TNF-,) and interleukin-6 (IL-6) in 534 Caucasian patients with type 2 diabetes mellitus. Methods:, MCP-1 concentrations were measured by enzyme-linked immunosorbent assay. MCP-1 genotyping was performed by RFLP analysis in a subset of 426 patients. Results:, Two hundred and thirty-one (54.2%) patients were homozygous for the wildtype allele (AA), 156 (36.6%) were heterozygous (AG) and 39 (9.2%) were homozygous for the mutated allele (GG). Allelic frequency was similar to non-diabetic populations (wildtype allele A: 0.73; mutated allele G: 0.27). MCP-1 mean concentrations and percentiles were substantially higher in non-diabetic populations but were not influenced by the genotype (AA: 662.0 ± 323.0 pg/ml; AG: 730.6 ± 491.4 pg/ml; GG: 641.2 ± 323.8 pg/ml). MCP-1 serum levels and genotypes were only marginally related to hormones (insulin and leptin) and cytokines (TNF-, and IL-6). Conclusions:, This is the first study providing MCP-1 levels, percentiles and genotype frequency in a large and representative cohort of patients with type 2 diabetes mellitus. Compared to the literature, MCP-1 levels were found to be substantially higher in patients with type 2 diabetes mellitus. In contrast, genotype frequencies were similar compared to those in non-diabetic patients and were not related to MCP-1 levels. The mechanisms behind these elevated MCP-1 serum levels in type 2 diabetes are not to be explained by simple associations with hormones, cytokines or genotypes. [source] The MTHFR C677T polymorphism confers a high risk for stroke in both homozygous and heterozygous T allele carriers with Type 2 diabetesDIABETIC MEDICINE, Issue 5 2006M. P. Hermans Abstract Objective Individuals with Type 2 diabetes are at increased risk of stroke. Plasma homocysteine (tHcy) is an independent risk factor for cardiovascular (CV) disease. The methylene,tetrahydrofolate reductase (MTHFR) gene polymorphism (thermolabile variant C677T) is associated with CV risk, partly as a result of increased Hcy, especially in homozygous subjects. Aim To relate the occurrence of the MTHFR polymorphism with stroke prevalence by examining allelic frequency and genotype distribution in 165 subjects with Type 2 diabetes studied for the presence of thermolabile C677T MTHFR mutation. Results Mean age was 67.7 years, and tHcy 18.2 µmol/l. T allele frequency was 38.5%. MTHFR genotypes were: normal (CC) 40%; heterozygous (CT) 43%; homozygous (TT) 17%. Serum levels of folic acid and B12 vitamin were within normal limits. Stroke prevalence was 14%. Sixty-four per cent of stroke-free subjects had the normal C allele vs. 46% in stroke subjects. The frequencies of genotypes (CC-CT-TT) were (%): 44-41-15 in stroke-free vs. 17-57-26 in stroke patients. Coronary (CAD) and peripheral artery disease (PAD) were common in all groups, with no differences according to genotypes. Stroke prevalence was markedly higher in genotypes CT and TT (18 and 21%) compared with CC (6%). Mean tHcy levels were higher in TT subjects. Conclusion The allelic frequency of C677T MTHFR mutation in Type 2 diabetes subjects with stroke is markedly different from that of subjects without stroke. Genotypic characteristics suggest that C677T MTHFR mutation confers a higher risk for stroke to both homozygous and heterozygous T allele carriers that cannot be ascribed solely to raised tHcy and/or lower folate status in CT subjects, nor to phenotypic expression of conventional risk factors for stroke. The impact of the MTHFR polymorphism on stroke may result from T allele-linked deleterious effects, or C allele-linked protection. Confirmatory studies are warranted, as this cohort was not randomly selected, and a type 1 error cannot be ruled out. [source] Genetic variants of insulin receptor substrate-1 (IRS-1) in syndromes of severe insulin resistance.DIABETIC MEDICINE, Issue 10 2002Functional analysis of Ala513Pro, Gly1158Glu IRS- Abstract Aims To define further the role of IRS-1 mutations in human syndromes of severe insulin resistance. Methods The IRS-1 gene was scanned for mutations in 83 unrelated affected subjects and 47 unaffected individuals using fluorescent single-strand conformation polymorphism (fSSCP) analysis. A novel heterozygous mutation, Gly1158Glu, was found in one affected subject. Four and two subjects were heterozygous for the previously reported variants Gly972Arg and Ala513Pro, respectively. The previously identified variant Gly819Arg was found in one affected and one unaffected subject. While Gly972Arg has been described to alter the signalling properties of IRS-1, no functional studies of Ala513Pro or Gly1158Glu have been reported. Results Chinese hamster ovary (CHO) cells stably over-expressing the insulin receptor were transiently transfected with vectors expressing either wild-type, Glu1158 or Pro513 IRS-1. A modest increase in insulin-stimulated tyrosine phosphorylation of Glu1158 IRS-1 was observed. However, this did not result in any significant change in the association of Grb2 or the p85, subunit of PI3-kinase or of PI3-kinase activity. In parallel studies, the Pro513 IRS-1 variant was indistinguishable from wild-type IRS-1. Conclusions While subtle effects of these variants cannot be excluded in this system, it is unlikely that these variants are responsible for the extreme insulin resistance seen in the subjects harbouring them. Although IRS proteins play a central role in insulin signalling, functionally significant mutations in the IRS-1 gene are a rare cause of human syndromes of severe insulin resistance. [source] Microevolution in agricultural environments: how a traditional Amerindian farming practice favours heterozygosity in cassava (Manihot esculenta Crantz, Euphorbiaceae)ECOLOGY LETTERS, Issue 2 2005Benoît Pujol Abstract We demonstrate a novel case of selection for heterozygosity in nature, involving inadvertent human selection on a population of domesticated plants. Amerindian farmers propagate cassava (Manihot esculenta) clonally by cuttings. Seedlings also appear spontaneously in fields, and farmers allow them to grow, later using some for cuttings. These ,volunteers' contribute new genotypes. However, many are inbred, whereas multiplied clones are highly heterozygous. We demonstrate the selective retention of heterozygous volunteers. When farmers weeded fields, they killed small volunteers, but retained large ones. Plant size and heterozygosity were correlated, and both increased after weeding. The process we document allows maintenance of genotypically diverse and heterozygous clonal stocks. Demonstrating heterosis in nature usually requires large sample sizes, but novel features of our system allowed escape of this constraint. Traditional agroecosystems offer unusual opportunities for the microevolutionary studies required to give on-farm conservation of genetic resources a solid scientific basis. [source] DNA variants in coding region of EFHC1: SNPs do not associate with juvenile myoclonic epilepsyEPILEPSIA, Issue 5 2009Dongsheng Bai Summary Purpose:, Juvenile myoclonic epilepsy (JME) accounts for 3 to 12% of all epilepsies. In 2004, we identified a mutation-harboring Mendelian gene that encodes a protein with one EF-hand motif (EFHC1) in chromosome 6p12. We observed one doubly heterozygous and three heterozygous missense mutations in EFHC1 segregating as an autosomal dominant gene with 21 affected members of six Hispanic JME families from California and Mexico. In 2006, similar and three novel missense mutations were reported in sporadic and familial Caucasian JME from Italy and Austria. In this study, we asked if coding single nucleotide polymorphisms (SNPs) of EFHC1 also contribute as susceptibility alleles to JME with complex genetics. Methods:, We screened using denaturing high-performance liquid chromatography (DHPLC) and then directly sequenced the 11 exons of EFHC1 in 130 unrelated JME probands, their 352 family members, and seven exons of EFHC1 in 400,614 ethnically matched controls. We carried out case-control association studies between 124 unrelated Hispanic JME probands and 552,614 ethnically matched controls using four SNPs, rs3804506, rs3804505, rs1266787, and rs17851770. We also performed family-based association on SNPs rs3804506 and rs3804505 in 84 complete JME families using the Family-Based Association Test (FBAT) program. Results:, We found no statistically significant differences between JME probands and controls in case-control association and no genetic transmission disequilibria in family-based association for the tested SNPs. In addition, we identified four new DNA variants in the coding region of EFHC1. Conclusion:, The four coding SNPs, rs3804506, rs3804505, rs1266787, and rs17851770, of EFHC1 may not be susceptibility alleles for JME. [source] Steroid-responsive functional B12 deficiency in association with transcobalamin II polymorphism 776C,GEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 1 2006Daniel P. Gale Abstract:, We present a case of intracellular vitamin B12 deficiency presenting with confusion, subacute combined degeneration of the cord, megaloblastic anaemia and intrinsic factor antibodies in the serum. Diagnosis was delayed by a normal serum B12 level and was confirmed by a grossly elevated serum homocysteine. There was a dramatic response to steroids. The patient was heterozygous for the transcobalamin (TC) II polymorphism 776C,G. This case demonstrates the importance of functional assessment of intracellular B12 activity (e.g. serum homocysteine) in excluding B12 deficiency, the role of steroids in pernicious anaemia and a possible clinical correlation of a TCII polymorphism. [source] Detection of unique neutrophil non-muscle myosin heavy chain-A localization by immunofluorescence analysis in MYH9 disorder presented with macrothrombocytopenia without leukocyte inclusions and deafnessEUROPEAN JOURNAL OF HAEMATOLOGY, Issue 1 2005Shinji Kunishima Abstract:,MYH9 disorders are autosomal-dominant macrothrombocytopenias with leukocyte inclusions caused by mutations in the MYH9 gene, which encodes the non-muscle myosin heavy chain-A (NMMHCA). We report a patient with an MYH9 disorder who presented with macrothrombocytopenia without leukocyte inclusions and severe bilateral sensory deafness. Conventional May,Grünwald,Giemsa staining failed to detect granulocyte cytoplasmic inclusions, whereas immunofluorescence analysis clearly demonstrated abnormal neutrophil NMMHCA localization. Genetic analyses revealed a novel heterozygous 18 base deletion in MYH9, leading to a six-amino acid in-frame deletion (N76_S81del) in NMMHCA. These results further support the usefulness of immunofluorescence analysis in differential diagnosis of MYH9 disorders. [source] Low frequency of Parkin, Tyrosine Hydroxylase, and GTP Cyclohydrolase I gene mutations in a Danish population of early-onset Parkinson's DiseaseEUROPEAN JOURNAL OF NEUROLOGY, Issue 4 2006J. M. Hertz Autosomal recessive Parkinson's disease (PD) with early-onset may be caused by mutations in the parkin gene (PARK2). We have ascertained 87 Danish patients with an early-onset form of PD (age at onset ,40 years, or ,50 years if family history is positive) in a multicenter study in order to determine the frequency of PARK2 mutations. Analysis of the GTP cyclohydrolase I gene (GCH1) and the tyrosine hydroxylase gene (TH), mutated in dopa-responsive dystonia and juvenile PD, have also been included. Ten different PARK2 mutations were identified in 10 patients. Two of the patients (2.3%) were found to have homozygous or compound heterozygous mutations, and eight of the patients (9.2%) were found to be heterozygous. A mutation has been identified in 10.4% of the sporadic cases and in 15.0% of cases with a positive family history of PD. One patient was found to be heterozygous for both a PARK2 mutation and a missense mutation (A6T) in TH of unknown significance. It cannot be excluded that both mutations contribute to the phenotype. No other putative disease causing TH or GCH1 mutations were found. In conclusion, homozygous, or compound heterozygous PARK2 mutations, and mutations in GCH1 and TH, are rare even in a population of PD patients with early-onset of the disease. [source] Impaired water maze learning performance without altered dopaminergic function in mice heterozygous for the GDNF mutationEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 7 2001R. Gerlai Abstract Exogenous glial cell line-derived neurotrophic factor (GDNF) exhibits potent survival-promoting effects on dopaminergic neurons of the nigrostriatal pathway that is implicated in Parkinson's disease and also protects neurons in forebrain ischemia of animal models. However, a role for endogenous GDNF in brain function has not been established. Although mice homozygous for a targeted deletion of the GDNF gene have been generated, these mice die within hours of birth because of deficits in kidney morphogenesis, and, thus, the effect of the absence of GDNF on brain function could not be studied. Herein, we sought to determine whether adult mice, heterozygous for a GDNF mutation on two different genetic backgrounds, demonstrate alterations in the nigrostriatal dopaminergic system or in cognitive function. While both neurochemical and behavioural measures suggested that reduction of GDNF gene expression in the mutant mice does not alter the nigrostriatal dopaminergic system, it led to a significant and selective impairment of performance in the spatial version of the Morris water maze. A standard panel of blood chemistry tests and basic pathological analyses did not reveal alterations in the mutants that could account for the observed performance deficit. These results suggest that endogenous GDNF may not be critical for the development and functioning of the nigrostriatal dopaminergic system but it plays an important role in cognitive abilities. [source] A study of a single variant allele (rs1426654) of the pigmentation-related gene SLC24A5 in Greek subjectsEXPERIMENTAL DERMATOLOGY, Issue 2 2009Gerasimos Dimisianos Abstract:, The SLC24A5 gene, the human orthologue of the zebrafish golden gene, has been shown to play a key role in human pigmentation. In this study, we investigate the prevalence of the variant allele rs1426654 in a selected sample of Greek subjects. Allele-specific polymerase chain reaction was performed in peripheral blood samples from 158 attendants of a dermatology outpatient service. The results were correlated with pigmentary traits and MC1R genotype. The vast majority of subjects (99%) were homozygous for the Thr111 allele. Only two subjects from the control group (1.26%) were heterozygous for the alanine and threonine allele. Both of these Thr111/Ala111 heterozygotes carried a single polymorphism of MC1R (one with the V92M variant and another with the V60L variant). Following reports of the rs1426654 polymorphism reaching fixation in the European population, our study of Greek subjects showed a prevalence of the Thr111 allele, even among subjects with darker skin pigmentation or phototype. [source] Molecular genetics of Xeroderma pigmentosum variantEXPERIMENTAL DERMATOLOGY, Issue 5 2003Alexei Gratchev Skin abnormalities result from an inability to repair UV-damaged DNA because of defects in the nucleotide excision repair (NER) machinery. Xeroderma pigmentosum is genetically heterogeneous and is classified into seven complementation groups (XPA-XPG) that correspond to genetic alterations in one of seven genes involved in NER. The variant type of XP (XPV), first described in 1970 by Ernst G. Jung as ,pigmented xerodermoid', is caused by defects in the post replication repair machinery while NER is not impaired. Identification of the XPV gene was only achieved in 1999 by biochemical purification and sequencing of a protein from HeLa cell extracts complementing the PRR defect in XPV cells. The XPV protein, polymerase (pol),, represents a novel member of the Y family of bypass DNA polymerases that facilitate DNA translesion synthesis. The major function of pol, is to allow DNA translesion synthesis of UV-induced TT-dimers in an error-free manner; it also possesses the capability to bypass other DNA lesions in an error-prone manner. Xeroderma pigmentosum V is caused by molecular alterations in the POLH gene, located on chromosome 6p21.1,6p12. Affected individuals are homozygous or compound heterozygous for a spectrum of genetic lesions, including nonsense mutations, deletions or insertions, confirming the autosomal recessive nature of the condition. Identification of POLH as the XPV gene provides an important instrument for improving molecular diagnostics in XPV families. [source] Molecular genetics of pseudoxanthoma elasticumEXPERIMENTAL DERMATOLOGY, Issue 4 2001F. Ringpfeil Abstract: Pseudoxanthoma elasticum (PXE), a systemic heritable connective tissue disorder, is characterized by progressive calcification of elastic structures in the skin, the eyes and the cardiovascular system, with considerable intra- and interfamilial phenotypic variability. Recently, underlying genetic defects have been identified in the ABCC6 gene, which resides on the chromosomal locus 16p13.1 and encodes the MRP6 protein, a member of the ATP-binding cassette (ABC) family of proteins. The affected individuals are homozygous or compound heterozygous for a spectrum of genetic lesions, including nonsense and missense mutations, or deletions and splice-site alterations, confirming the autosomal recessive nature of this condition. Analysis of the deduced primary sequence suggests that MRP6 is a transmembrane transporter, but its function has not been delineated yet. Surprisingly, however, MRP6 is expressed primarily, if not exclusively, in the liver and the kidneys, suggesting that PXE may be a primary metabolic disorder with secondary involvement of elastic fibers. Identification of mutations in the ABCC6 gene in PXE provides a means for prenatal and presymptomatic testing in families at risk for recurrence. DNA-based analyses will also identify heterozygous carriers who may be at risk for development of limited manifestations of the disease as a result of compounding genetic factors and/or environmental modifiers. [source] Direct genotyping of the poplar leaf rust fungus, Melampsora medusae f. sp. deltoidae, using codominant PCR-SSCP markersFOREST PATHOLOGY, Issue 4 2005M. Bourassa Summary Two anonymous DNA markers that are revealed by single-strand conformational polymorphism (SSCP) analysis were developed for detection of polymorphisms in Melampsora medusae f. sp. deltoidae (Mmd). Mono-uredinial isolates of Mmd were first obtained, DNA was extracted from urediniospores and random amplified polymorphic DNA (RAPD) products of eight mono-uredinial isolates were separated on a SSCP gel to identify differences among them. Bands representing putative polymorphic loci among the eight isolates tested were excised from the SSCP gel and re-amplified by polymerase chain reaction (PCR), and then cloned and sequenced. A primer pair was designed to amplify a DNA fragment of a size suitable for SSCP analysis (<600 bp) for two out of three DNA fragments sequenced. Each set of primers amplified a PCR product for all eight isolates that were initially used to generate them and the resulting PCR products were analysed by SSCP. Polymorphisms among isolates were identified for both putative loci. The two primer pairs amplified a PCR product of the expected size on an additional 32 mono-uredinial isolates of Mmd tested. From the overall 40 mono-uredinial isolates tested, 5 and 11 alleles were detected, and 12 and 34 isolates showed to be heterozygous, as indicated by the presence of more than two bands on the SSCP gel, at loci A and B, respectively. The primer pairs were tested for specificity against 106 fungal isolates belonging to various taxa, including other rusts, and against DNA extracted from greenhouse-grown healthy poplar leaves. DNA amplification products of the expected size were obtained only when Mmd DNA was present. Optimization of PCR conditions with these two primer pairs allowed genotyping directly from single uredinia extracted from infected leaves, thus alleviating the need to culture the fungus to characterize individuals, hence making it possible to process large numbers of samples for population studies. Résumé Deux marqueurs génétiques anonymes, révélés par analyse SSCP (Single-Strand Conformational Polymorphism) ont été développés afin de détecter des polymorphismes génétiques chez le Melampsora medusae f. sp. deltoidae (Mmd). Dans un premier temps, des isolats mono-urédiniaux ont été obtenus, puis l'ADN a été extrait à partir des urédiniospores, les produits d'amplification RAPD (Random Amplified Polymorphic DNA) ont été générés à partir de huit de ces isolats mono-urédiniaux et les résultats d'amplification ont par la suite été séparés sur gel SSCP afin d'identifier des polymorphismes entre les isolats. Les bandes sur gel SSCP représentant des loci polymorphiques putatifs entre les isolats ont été prélevées du gel, ré-amplifiées par la technique d'amplification PCR (Polymerase Chain Reaction), clonées, puis séquencées. Pour deux fragments d'ADN séquencés sur un total de trois, une paire d'amorces a été développée afin de permettre l'amplification d'un fragment de taille adéquate pour analyse SSCP (<600 pb). Chaque paire d'amorces a produit un signal d'amplification positif pour chacun des huit isolats à l'origine de ces nouvelles amorces; les produits PCR ont ensuite été analysés par la technique SSCP. Les deux loci putatifs ont révélé des polymorphismes génétiques entre les isolats. Les deux paires d'amorces ont produit un fragment d'amplification de la taille attendue pour chacun des 32 isolats mono-urédiniaux supplémentaires testés. Des 40 isolats testés, 5 et 11 allèles ont été détectés, alors que 12 et 34 isolats se sont révélés hétérozygotes (tel qu'indiqué par la présence de plus de deux bandes sur gel SSCP) pour les loci A et B, respectivement. La spécificité des deux paires d'amorces a été testée à partir de 106 isolats fongiques appartenant à différents groupes taxonomiques, incluant d'autres rouilles, de même qu'à partir de l'ADN extrait de feuilles de peupliers cultivés en serre. Un signal d'amplification positif n'a été obtenu qu'en présence d'ADN du Mmd. Les conditions d'amplification PCR ont été optimisées pour les deux paires d'amorces développées afin de permettre le génotypage directement à partir d'urédinies individuelles prélevées sur des feuilles de peuplier infectées. La possibilité de génotyper directement des urédinies individuelles permet d'éviter l'obligation de cultiver le champignon pour génotyper les individus, ce qui représente un avantage important des marqueurs génétiques développés ici, puisqu'il devient dès lors possible de traiter un grand nombre d'échantillons lors de la réalisation d'études de populations. Zusammenfassung Zum Nachweis von Polymorphismen bei Melampsora medusae f. sp. deltoidae wurden zwei anonyme DNA Marker aus einer SSCP-Analyse entwickelt. Zunächst wurden Isolate aus einzelnen Uredinien gewonnen, die DNA wurde aus den Uredosporen extrahiert und polymorphe RAPD, Amplifikationsprodukte von acht Mono-Uredinium-Isolaten wurden auf einem SSCP-Gel getrennt, um Unterschiede zwischen ihnen nachzuweisen. Banden, die bei den acht geprüften Isolaten mögliche polymorphe Loci darstellten, wurden aus dem SSCP-Gel ausgeschnitten und mit PCR reamplifiziert, dann geklont und sequenziert. Für zwei von insgesamt drei sequenzierten DNA-Fragmenten wurde ein Primerpaar entwickelt, um ein in der Grösse für die SSCP-Analyse (<600 bp) geeignetes DNA-Fragment zu amplifizieren. Jedes Primerpaar amplifizierte bei allen acht ursprünglich für ihre Entwicklung verwendeten Isolaten ein PCR-Produkt, und diese wurden anschliessend mit SSCP analysiert. Für beide putativen Loci wurden bei den Isolaten Polymorphismen festgestellt. Die beiden Primerpaare amplifizierten ein PCR-Produkt der erwarteten Grösse bei allen 32 zusätzlich geprüften Mono-Uredinium-Isolaten des Pilzes. Bei den insgesamt 40 geprüften Mono-Uredinium-Isolaten wurden für die Loci A und B 5 bzw. 11 Allele gefunden, und 12 bzw. 34 Isolate erwiesen sich als heterozygot, was durch mehr als zwei Banden auf den SSCP-Gelen angezeigt wurde. Die Spezifität der Primerpaare wurden mit 106 Pilzisolaten aus verschiedenen Taxa geprüft, darunter andere Roste sowie DNA aus gesunden Pappelblättern aus Gewächshauskulturen. DNA-Amplifikationsprodukte der erwarteten Grösse wurden nur erhalten, wenn DNA von Melampsora medusae f. sp. deltoidae präsent war. Die PCR-Amplifikations-Bedingungen mit diesen beiden Primerpaaren wurde so optimiert, dass ein Genotyping direkt bei einzelnen von infizierten Blättern entnommenen Uredinien erfolgen kann und somit eine Pilzkultur zur Charakterisierung von Individuen entfällt. Dies ermöglicht grosse Probenzahlen in Populationsstudien. [source] Amygdala protein kinase C epsilon controls alcohol consumptionGENES, BRAIN AND BEHAVIOR, Issue 5 2009H. M. B. Lesscher Alcoholism is a progressive disorder that involves the amygdala. Mice lacking protein kinase C epsilon (PKC,) show reduced ethanol consumption, sensitivity and reward. We therefore investigated whether PKC, signaling in the amygdala is involved in ethanol consumption. Local knockdown of PKC, in the amygdala reduced ethanol consumption and preference in a limited-access paradigm. Further, mice that are heterozygous for the PKC, allele consume less ethanol compared with wild-type mice in this paradigm. These mice have a >50% reduction in the abundance of PKC, in the amygdala compared with wild-type mice. We conclude that amygdala PKC, is important for ethanol consumption in mice. [source] Neuronal cell adhesion molecule deletion induces a cognitive and behavioral phenotype reflective of impulsivityGENES, BRAIN AND BEHAVIOR, Issue 4 2008L. D. Matzel Cell adhesion molecules, such as neuronal cell adhesion molecule (Nr-CAM), mediate cell,cell interactions in both the developing and mature nervous system. Neuronal cell adhesion molecule is believed to play a critical role in cell adhesion and migration, axonal growth, guidance, target recognition and synapse formation. Here, wild-type, heterozygous and Nr-CAM null mice were assessed on a battery of five learning tasks (Lashley maze, odor discrimination, passive avoidance, spatial water maze and fear conditioning) previously developed to characterize the general learning abilities of laboratory mice. Additionally, all animals were tested on 10 measures of sensory/motor function, emotionality and stress reactivity. We report that the Nr-CAM deletion had no impact on four of the learning tasks (fear conditioning, spatial water maze, Lashley maze and odor discrimination). However, Nr-CAM null mice exhibited impaired performance on a task that required animals to suppress movement (passive avoidance). Although Nr-CAM mutants expressed normal levels of general activity and body weights, they did exhibit an increased propensity to enter stressful areas of novel environments (the center of an open field and the lighted side of a dark/light box), exhibited higher sensitivity to pain (hot plate) and were more sensitive to the aversive effects of foot shock (shock-induced freezing). This behavioral phenotype suggests that Nr-CAM does not play a central role in the regulation of general cognitive abilities but may have a critical function in regulating impulsivity and possibly an animal's susceptibility to drug abuse and addiction. [source] Hypolocomotion, anxiety and serotonin syndrome-like behavior contribute to the complex phenotype of serotonin transporter knockout miceGENES, BRAIN AND BEHAVIOR, Issue 4 2007A. V. Kalueff Although mice with a targeted disruption of the serotonin transporter (SERT) have been studied extensively using various tests, their complex behavioral phenotype is not yet fully understood. Here we assess in detail the behavior of adult female SERT wild type (+/+), heterozygous (+/,) and knockout (,/,) mice on an isogenic C57BL/6J background subjected to a battery of behavioral paradigms. Overall, there were no differences in the ability to find food or a novel object, nest-building, self-grooming and its sequencing, and horizontal rod balancing, indicating unimpaired sensory functions, motor co-ordination and behavioral sequencing. In contrast, there were striking reductions in exploration and activity in novelty-based tests (novel object, sticky label and open field tests), accompanied by pronounced thigmotaxis, suggesting that combined hypolocomotion and anxiety (rather than purely anxiety) influence the SERT ,/, behavioral phenotype. Social interaction behaviors were also markedly reduced. In addition, SERT ,/, mice tended to move close to the ground, frequently displayed spontaneous Straub tail, tics, tremor and backward gait , a phenotype generally consistent with ,serotonin syndrome'-like behavior. In line with replicated evidence of much enhanced serotonin availability in SERT ,/, mice, this serotonin syndrome-like state may represent a third factor contributing to their behavioral profile. An understanding of the emerging complexity of SERT ,/, mouse behavior is crucial for a detailed dissection of their phenotype and for developing further neurobehavioral models using these mice. [source] Focal 9p instability in hematologic neoplasias revealed by comparative genomic hybridization and single-nucleotide polymorphism microarray analysesGENES, CHROMOSOMES AND CANCER, Issue 4 2010Anu Usvasalo Copy number losses in chromosome arm 9p are well-known aberrations in malignancies, including leukemias. The CDKN2A gene is suggested to play a key role in these aberrations. In this study overviewing 9p losses in hematologic neoplasias, we introduce the term focal 9p instability to indicate multiple areas of copy number loss or homozygous loss within a larger heterozygous one in 9p. We have used microarray comparative genomic hybridization to study patients with acute lymphoblastic leukemia (ALL, n = 140), acute myeloid leukemia (n = 50), chronic lymphocytic leukemia (n = 20), and myelodysplastic syndromes (n = 37). Our results show that 9p instability is restricted to ALL. In total, 58/140 (41%) patients with ALL had a loss in 9p. The 9p instability was detected in 19% of the patients with ALL and always included homozygous loss of CDKN2A along with loss of CDKN2B. Other possibly important genes included MTAP, IFN, MLLT3, JAK2, PTPLAD2, and PAX5. 13/27 (48%) patients with the instability had the BCR/ABL1 fusion gene or other oncogene-activating translocation or structural aberrations. Two patients had homozygous loss of hsa-mir ,31, a microRNA known to regulate IKZF1. IKZF1 deletion at 7p12.1 was seen in 10 (37%) patients with the 9p instability. These findings suggest that, in ALL leukemogenesis, loss of CDKN2A and other target genes in the instability region is frequently associated with BCR/ABL1 and IKZF1 dysfunction. The multiple mechanisms leading to 9p instability including physical or epigenetic loss of the target genes, loss of the microRNA cluster, and the role of FRA9G fragile site are discussed. © 2009 Wiley-Liss, Inc. [source] AML1/RUNX1 mutations are infrequent, but related to AML-M0, acquired trisomy 21, and leukemic transformation in pediatric hematologic malignanciesGENES, CHROMOSOMES AND CANCER, Issue 1 2003Takeshi Taketani AML1/RUNX1, located on chromosome band 21q22, is one of the most important hematopoietic transcription factors. AML1 is frequently affected in leukemia and myelodysplastic syndrome with 21q22 translocations. Recently, AML1 mutations were found in adult hematologic malignancies, especially acute myeloid leukemia (AML),M0 or leukemia with acquired trisomy 21, and familial platelet disorder with a predisposition toward AML. Through the use of polymerase chain reaction,single-strand conformation polymorphism analysis, we examined the AML1 gene for mutations in 241 patients with pediatric hematologic malignancies, and we detected AML1 mutations in seven patients (2.9%). Deletion was found in one patient, and point mutations in four patients, including three missense mutations, two silent mutations, and one mutation within an intron resulting in an abnormal splice acceptor site. All of the mutations except for one were heterozygous. Mutations within the runt domain were found in six of seven patients. Six of seven patients with AML1 mutations were diagnosed with AML, and one had acute lymphoblastic leukemia. In three of these seven patients, AML evolved from other hematologic disorders. AML1 mutations were found in two of four AML-M0 and two of three patients with acquired trisomy 21. Patients with AML1 mutations tended to be older children. Three of four patients with AML1 mutations who received stem cell transplantation (SCT) are alive, whereas the remaining three patients with mutations without SCT died. These results suggest that AML1 mutations in pediatric hematologic malignancies are infrequent, but are possibly related to AML-M0, acquired trisomy 21, and leukemic transformation. These patients may have a poor clinical outcome. © 2003 Wiley-Liss, Inc. [source] An essential gene mutagenesis screen across the highly conserved piebald deletion region of mouse chromosome 14GENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 6 2009James A. Hagarman Piebald (left) and piebald deletion heterozygous (right) mice, showing altered pigmentation. An ENU mutagenesis screen for essential genes was performed using piebald deletion complex alleles. See the paper by Hagarman and O'Brien in this issue. [source] Stable generation of serum- and feeder-free embryonic stem cell-derived mice with full germline-competency by using a GSK3 specific inhibitorGENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 6 2009Hiromu Sato Abstract C57BL/6 (B6)-derived embryonic stem (ES) cells are not widely used to generate knockout mice despite the advantage of a well-defined genetic background because of poor developmental potential. We newly established serum- and feeder-free B6 ES cells with full developmental potential by using leukemia inhibitory factor (LIF) and 6-bromoindirubin-3,-oxime (BIO), a glycogen synthase kinase-3 (GSK3) inhibitor. BIO treatment significantly increased the expression levels of 364 genes including pluripotency markers such as Nanog and Klf family. Unexpectedly, by aggregating or microinjecting those ES cells to each eight-cell-stage diploid embryo, we stably generated germline-competent ES-derived mice. Furthermore, founder mice completely derived from female XO, heterozygous, or homozygous mutant B6 ES cells were directly available for intercross breeding and phenotypic analysis. We hereby propose that serum- and feeder-free B6 ES cells stimulated with LIF plus GSK3 inhibitor are valuable for generating mouse models on B6 background. genesis 47:414,422, 2009. © 2009 Wiley-Liss, Inc. [source] High-resolution mapping of the Gli3 deletion in the mouse extra-toesH mutantGENESIS: THE JOURNAL OF GENETICS AND DEVELOPMENT, Issue 3 2007Matthieu Genestine Abstract Extra-toes is a semidominant mutation that affects the Gli3 gene and provokes limb and brain abnormalities. Among the different alleles of this mutation, XtH is due to a deletion that has not yet been fully characterized. Using a PCR-based strategy, we undertook a high-resolution mapping of this deletion and confirmed that XtH is a null allele of Gli3. We further designed a PCR test to identify unequivocally heterozygous and homozygous embryos from their wild-type littermates. Despite the length of the XtH deletion, available data on the mouse genome indicate that no genes other than Gli3 are deleted in XtH mutants. Thus, the XtH mutation can be used as a model for studying the effects that absence of Gli3 function has during development. genesis 45:107,112, 2007. © 2007 Wiley-Liss, Inc. [source] | |||||||||||||||||||||||||||||||||||||