A RAB3GAP1 SINE Insertion in Alaskan Huskies with Polyneuropathy, Ocular Abnormalities and Neuronal Vacuolation (POANV) Resembling Human Warburg Micro Syndrome 1 (WARBM1)

We observed a hereditary phenotype in Alaskan Huskies, which was characterized by polyneuropathy with ocular abnormalities and neuronal vacuolation (POANV). The affected dogs developed a progressive severe ataxia, which led to euthanasia between 8 and 16 months of age. The pedigrees were consistent with a monogenic autosomal recessive inheritance. We localized the causative genetic defect to a 4 Mb interval on chromosome 19 by a combined linkage and homozygosity mapping approach. Whole genome sequencing of one affected dog, an obligate carrier and an unrelated control revealed a 218 bp SINE insertion into exon 7 of the RAB3GAP1 gene. The SINE insertion was perfectly associated with the disease phenotype in a cohort of 43 Alaskan Huskies and it was absent from 541 control dogs of diverse other breeds. The SINE insertion induced aberrant splicing and led to a transcript with a greatly altered exon 7. RAB3GAP1 loss-of-function variants in humans cause Warburg Micro Syndrome 1 (WARBM1), which is characterized by additional developmental defects compared to canine POANV, whereas Rab3gap1 deficient mice have a much milder phenotype than either humans or dogs. Thus the RAB3GAP1 mutant Alaskan Huskies provide an interesting intermediate phenotype that may help to better understand the function of RAB3GAP1 in development. Furthermore, the identification of the presumed causative genetic variant will enable genetic testing to avoid the non-intentional breeding of affected dogs.

Life-long growth of Quercus ilex L. at natural CO2 springs acclimates sulphur, nitrogen and carbohydrate metabolism of the progeny to elevated pCO2

The aim of the present study was to analyse whether offspring of mature Quercus ilex trees grown under life-long elevated pCO2 show alterations in the physiological response to elevated pCO2 in comparison with those originating from mature trees grown at current ambient pCO2. To investigate changes in C- (for changes in photosynthesis, biomass and lignin see Polle, McKee & Blaschke Plant, Cell and Environment 24, 1075–1083, 2001), N-, and S-metabolism soluble sugar, soluble non-proteinogenic nitrogen compounds (TSNN), nitrate reductase (NR), thiols, adenosine 5′-phosphosulphate (APS) reductase, and anions were analysed. For this purpose Q. ilex seedlings were grown from acorns of mother tree stands at a natural spring site (elevated pCO2) and a control site (ambient pCO2) of the Laiatico spring, Central Italy. Short-term elevated pCO2 exposure of the offspring of control oaks lead to higher sugar contents in stem tissues, to a reduced TSNN content in leaves, and basipetal stem tissues, to diminished thiol contents in all tissues analysed, and to reduced APS reductase activity in both, leaves and roots. Most of the components of C-, N- and S-metabolism including APS reductase activity which were reduced due to short-term elevated pCO2 exposure were recovered by life-long growth under elevated pCO2 in the offspring of spring oaks. Still TSNN contents in phloem exudates increased, nitrate contents in lateral roots and glutathione in leaves and phloem exudates remained reduced in these plants. The present results demonstrated that metabolic adaptations of Q. ilex mother trees to elevated pCO2 can be passed to the next generation. Short- and long-term effects on source-to-sink relation and physiological and genetic acclimation to elevated pCO2 are discussed.

Visual Hallucinations in Eye Disease and Lewy Body Disease

Objective: Visual hallucinations (VH) most commonly occur in eye disease (ED), Parkinson’s disease (PD), and Lewy body dementia (LBD). The phenomenology of VH is likely to carry important information about the brain areas within the visual system generating them. Methods: Data from five controlled cross-sectional VH studies (164 controls, 135 ED, 156 PD, 79 (PDD 48 + DLB 31) LBD) were combined and analysed. The prevalence, phenomenology, frequency, duration, and contents of VH were compared across diseases and gender. Results: Simple VH were most common in ED patients (ED 65% vs. LBD 22% vs. PD 9%, Chi-square [χ2] test: χ2=31.43, df=2, p<0.001), whilst complex VH were more common in LBD (LBD 76% vs. ED 38%, vs PD 28%, Chi-square test: χ2=96.80, df=2, p<0.001). The phenomenology of complex VH was different across diseases and gender. ED patients reported more “flowers” (ED 21% vs. LBD 6% vs. PD 0%, Chi-square test: χ2=10.04, df=2, p=0.005) and “body parts” (ED 40% vs. LBD 17% vs. PD 13%, Chi-square test: χ2=11.14, df=2, p=0.004); in contrast LBD patients reported “people” (LBD 85% vs. ED 67% vs. PD 63%, Chi-square test: χ2=6.20, df=2, p=0.045) and “animals/insects” (LBD 50% vs. PD 42% vs. ED 21%, Chi-square test: χ2=9.76, df=2, p=0.008). Males reported more “machines” (13 % vs. 2%, Chi-square test: χ2=6.94, df=1, p=0.008), whilst females reported more “family members/children” (48% vs. 29%, Chi-square test: χ2=5.10, df=1, p=0.024). Conclusions: The phenomenology of VH is likely related to disease specific dysfunctions within the visual system and to past, personal experiences.

Bleeding on probing as it relates to smoking status in patients enrolled in supportive periodontal therapy for at least 5 years

AIM To relate the mean percentage of bleeding on probing (BOP) to smoking status in patients enrolled in supportive periodontal therapy (SPT). MATERIALS AND METHODS Retrospective data on BOP from 8'741 SPT visits were related to smoking status among categories of both periodontal disease severity and progression (instability) in patients undergoing dental hygiene treatment at the Medi School of Dental Hygiene (MSDH), Bern, Switzerland 1985-2011. RESULTS A total of 445 patients were identified with 27.2% (n = 121) being smokers, 27.6% (n = 123) former smokers and 45.2% (n = 201) non-smokers. Mean BOP statistically significantly increased with disease severity (p = 0.0001) and periodontal instability (p = 0.0115) irrespective of the smoking status. Periodontally stable smokers (n = 30) categorized with advanced periodontal disease demonstrated a mean BOP of 16.2% compared to unstable smokers (n = 15) with a mean BOP of 22.4% (p = 0.0291). Assessments of BOP in relation to the percentage of sites with periodontal probing depths (PPD) ≥ 4 mm at patient-level yielded a statistically significantly decreased proportion of BOP in smokers compared to non-smokers and former smokers (p = 0.0137). CONCLUSIONS Irrespective of the smoking status, increased mean BOP in SPT patients relates to disease severity and periodontal instability while smokers demonstrate lower mean BOP concomitantly with an increased prevalence of residual PPDs.

Dermatomyositis in a family of Working Kelpies

Eight members of a family of Working Kelpies were presented with signs compatible with dermatomyositis. Alopecia, crusts, ulcerations of the skin, depigmentation of nasal planum and lips, onychodystrophy and atrophy of the masticatory muscles were present with varying degree. Histopathology of the skin, but not from muscles was performed in three dogs and confirmed the clinical diagnosis. Different immunomodulating drugs (steroids, cyclosporine, mycophenolate mofetil, pentoxifylline, doxycyline/niacinamid, omega-3 fatty acids and vitamin E) were used with variable success. Dermatomyositis is an immune-mediated disease and a genetic predisposition is known in humans and certain canine breeds, mainly Shetland Sheepdogs and Collies, but also for the Beauceron. The responsible genes have not been identified so far. It is assumed that the Working Kelpie derives from the Collie which could explain a hereditary predisposition in the Kelpie.

Sociodemographic, behavioral and genetic determinants of allostatic load in a Swiss population-based study.

Allostatic load (AL) is a marker of physiological dysregulation which reflects exposure to chronic stress. High AL has been related to poorer health outcomes including mortality. We examine here the association of socioeconomic and lifestyle factors with AL. Additionally, we investigate the extent to which AL is genetically determined. We included 803 participants (52% women, mean age 48±16years) from a population and family-based Swiss study. We computed an AL index aggregating 14 markers from cardiovascular, metabolic, lipidic, oxidative, hypothalamus-pituitary-adrenal and inflammatory homeostatic axes. Education and occupational position were used as indicators of socioeconomic status. Marital status, stress, alcohol intake, smoking, dietary patterns and physical activity were considered as lifestyle factors. Heritability of AL was estimated by maximum likelihood. Women with a low occupational position had higher AL (low vs. high OR=3.99, 95%CI [1.22;13.05]), while the opposite was observed for men (middle vs. high OR=0.48, 95%CI [0.23;0.99]). Education tended to be inversely associated with AL in both sexes(low vs. high OR=3.54, 95%CI [1.69;7.4]/OR=1.59, 95%CI [0.88;2.90] in women/men). Heavy drinking men as well as women abstaining from alcohol had higher AL than moderate drinkers. Physical activity was protective against AL while high salt intake was related to increased AL risk. The heritability of AL was estimated to be 29.5% ±7.9%. Our results suggest that generalized physiological dysregulation, as measured by AL, is determined by both environmental and genetic factors. The genetic contribution to AL remains modest when compared to the environmental component, which explains approximately 70% of the phenotypic variance.

The TREAT-NMD DMD Global Database: analysis of more than 7,000 Duchenne muscular dystrophy mutations.

Analyzing the type and frequency of patient-specific mutations that give rise to Duchenne muscular dystrophy (DMD) is an invaluable tool for diagnostics, basic scientific research, trial planning, and improved clinical care. Locus-specific databases allow for the collection, organization, storage, and analysis of genetic variants of disease. Here, we describe the development and analysis of the TREAT-NMD DMD Global database (http://umd.be/TREAT_DMD/). We analyzed genetic data for 7,149 DMD mutations held within the database. A total of 5,682 large mutations were observed (80% of total mutations), of which 4,894 (86%) were deletions (1 exon or larger) and 784 (14%) were duplications (1 exon or larger). There were 1,445 small mutations (smaller than 1 exon, 20% of all mutations), of which 358 (25%) were small deletions and 132 (9%) small insertions and 199 (14%) affected the splice sites. Point mutations totalled 756 (52% of small mutations) with 726 (50%) nonsense mutations and 30 (2%) missense mutations. Finally, 22 (0.3%) mid-intronic mutations were observed. In addition, mutations were identified within the database that would potentially benefit from novel genetic therapies for DMD including stop codon read-through therapies (10% of total mutations) and exon skipping therapy (80% of deletions and 55% of total mutations).

Knockdown of Kiaa0319 Reduces Dendritic Spine Density

Developmental Dyslexia is a reading disorder that affects individuals that possess otherwise normal intelligence. Until the four candidate dyslexia susceptibility genes were discovered, the cause of cortical malformations found in post mortem dyslexic brains was unclear. Normal brain development is crucial for the proper wiring of the neural circuitry that allow an individual to perform cognitive tasks like reading. For years, familial and twin studies have suggested that there was a genetic basis to the causation of dyslexia. Kiaa0319 was among the candidate dyslexia susceptibility genes that were ascertained. KIAA0319 is located on Chromosome 6p22.2-22.3 and has been found to exhibit differential spatial-temporal expression patterns in the brain throughout development, which suggests that the polycystic kidney disease (PKD) domain encoded by KIAA0319 facilitates cell-cell adhesion to enable neuronal precursors to crawl up the radial glia during neuronal migration. With the knowledge of KIAA0319 involvement in early neurogenesis, we were interested in determining how different KIAA0319 expression may impact cortical neurons in layer II and III during early adulthood. We show that KIAA0319 knockdown in cortical pyramidal neurons significantly reduces the dendritic spine density. Studies have shown that changes in dendritic spine morphology and density affect properties of neural circuitry. Henceforth, this finding may reveal a link between the Kiaa0319 gene and the deficit of the neural processing task of reading due to reduced spines density. Finding a correlation between Kiaa0319 expression and its influence on dendritic spine development may lead to a greater insight of a direct link between the dyslexia susceptibility gene and the biological mechanism that causes dyslexia.

Identification of germ cell tumor modifier genes from the 129.MOLF-Chr19 consomic mouse strain

Naturally occurring genetic variants confer susceptibility to disease in the human population, including in testicular germ cell tumor development. Disease susceptibility loci for testicular germ cell tumors have been identified by genetic mapping in humans and mice. However, the identity of many of the susceptibility genes remains unclear. My study utilized a chromosome substitution strain, the 129.MOLF-Chr 19 (or M19 strain), to identify candidate testicular germ cell tumor susceptibility genes. Males of this strain have a high incidence of germ cell tumors in the testes. By forward genetic approaches, five susceptibility loci were fine-mapped and the genetic interactions were dissected. In addition, I identified three protein-coding genes and one micro-RNA as testicular tumor susceptibility genes by genomic screening. Using reverse genetic approaches, I verified one of the candidates, Splicing factor 1, as a modifier of testicular tumor. Deficiency of SF1 significantly reduces the incidence of testicular tumors in mice. This study highlights the advantage of the 129.MOLF-Chr 19 consomic strain in disease gene identification and validation. It also sets the stage to elucidate the molecular mechanisms of tumorigenesis in the testis. ^

Identification of diabetic retinopathy genes through a genome-wide association study among Mexican-Americans from Starr County, Texas

To identify genetic susceptibility loci for severe diabetic retinopathy, 286 Mexican-Americans with type 2 diabetes from Starr County, Texas completed detailed physical and ophthalmologic examinations including fundus photography for diabetic retinopathy grading. 103 individuals with moderate-to-severe non-proliferative diabetic retinopathy or proliferative diabetic retinopathy were defined as cases for this study. DNA samples extracted from study subjects were genotyped using the Affymetrix GeneChip® Human Mapping 100K Set, which includes 116,204 single nucleotide polymorphisms (SNPs) across the whole genome. Single-marker allelic tests and 2- to 8-SNP sliding-window Haplotype Trend Regression implemented in HelixTreeTM were first performed with these direct genotypes to identify genes/regions contributing to the risk of severe diabetic retinopathy. An additional 1,885,781 HapMap Phase II SNPs were imputed from the direct genotypes to expand the genomic coverage for a more detailed exploration of genetic susceptibility to diabetic retinopathy. The average estimated allelic dosage and imputed genotypes with the highest posterior probabilities were subsequently analyzed for associations using logistic regression and Fisher's Exact allelic tests, respectively. To move beyond these SNP-based approaches, 104,572 directly genotyped and 333,375 well-imputed SNPs were used to construct genetic distance matrices based on 262 retinopathy candidate genes and their 112 related biological pathways. Multivariate distance matrix regression was then used to test hypotheses with genes and pathways as the units of inference in the context of susceptibility to diabetic retinopathy. This study provides a framework for genome-wide association analyses, and implicated several genes involved in the regulation of oxidative stress, inflammatory processes, histidine metabolism, and pancreatic cancer pathways associated with severe diabetic retinopathy. Many of these loci have not previously been implicated in either diabetic retinopathy or diabetes. In summary, CDC73, IL12RB2, and SULF1 had the best evidence as candidates to influence diabetic retinopathy, possibly through novel biological mechanisms related to VEGF-mediated signaling pathway or inflammatory processes. While this study uncovered some genes for diabetic retinopathy, a comprehensive picture of the genetic architecture of diabetic retinopathy has not yet been achieved. Once fully understood, the genetics and biology of diabetic retinopathy will contribute to better strategies for diagnosis, treatment and prevention of this disease.^

Genetic variability and the hypothalamic control of energy balance

Background. Obesity is a major health problem throughout the industrialized world. Despite numerous attempts to curtail the rapid growth of obesity, its incidence continues to rise. Therefore, it is crucial to better understand the etiology of obesity beyond the concept of energy balance.^ Aims. The first aim of this study was to first investigate the relationship between eating behaviors and body size. The second goal was to identify genetic variation associated with eating behaviors. Thirdly, this study aimed to examine the joint relationships between eating behavior, body size and genetic variation.^ Methods. This study utilized baseline data ascertained in young adults from the Training Interventions and Genetics of Exercise (TIGER) Study. Variables assessed included eating behavior (Emotional Eating Scale, Eating Attitudes Test-26, and the Block98 Food Frequency Questionnaire), body size (body mass index, waist and hip circumference, waist/hip ratio, and percent body fat), genetic variation in genes implicated related to the hypothalamic control of energy balance, and appropriate covariates (age, gender, race/ethnicity, smoking status, and physical activity. For the genetic association analyses, genotypes were collapsed by minor allele frequency, and haplotypes were estimated for each gene. Additionally, Bayesian networks were constructed in order to determine the relationships between genetic variation, eating behavior and body size.^ Results. We report that the EAT-26 score, Caloric intake, percent fat, fiber intake, HEAT index, and daily servings of vegetables, meats, grains, and fats were significantly associated with at least one body size measure. Multiple SNPs in 17 genes and haplotypes from 12 genes were tested for their association with body size. Variation within both DRD4 and HTR2A was found to be associated with EAT-26 score. In addition, variation in the ghrelin gene (GHRL) was significantly associated with daily Caloric intake. A significant interaction between daily servings of grains and the HEAT index and variation within the leptin receptor gene (LEPR) was shown to influence body size.^ Conclusion. This study has shown that there is a substantial genetic component to eating behavior and that genetic variation interacts with eating behavior to influence body size.^

Genetic epidemiology of childhood brain tumors

A rare familial cancer syndrome involving childhood brain tumors (CBT), breast cancer, sarcomas and an array of other tumors has been described (Li and Fraumeni 1969, 1975, 1982, 1987). A survey of CBT identified through the Connnecticut Tumor Registry in 1984 revealed a high frequency of CBT, leukemia and other childhood cancer in siblings of CBT patients (Farwell and Flannery, 1984). Other syndromes such as neurofibromatosis and nevoid basal cell carcinoma syndrome have also been associated with CBT; however, no systematic family studies have been conducted to determine the extent to which cancer aggregates in family members of CBT patients. This family study was designed to determine the frequency of cancer aggregation overall or at specific sites, to determine the frequency of known or potentially hereditary syndromes in families of CBT patients, and to determine a genetic model to characterize familial cancer syndromes and to identify specific kindreds to which such a model(s) might apply. This study includes 244 confirmed CBT patients referred to the University of Texas M. D. Anderson Cancer Center between the years 1944 and 1983, diagnosed under the age of 15 years and resident in the U.S. or Canada. Family histories were obtained on the proband's first (parents, siblings and offspring) and second degree (proband's aunts, uncles and grandparents) relatives following sequential sampling scheme rules. To determine if cancer aggregates in families, we compared the cancer experience in the population to that expected in the general population using Connecticut Tumor Registry calendar year, age, race and sex-specific rates. The standardized incidence ratio (SIR) for cancer overall was 0.91 (41 observed (O) and 44.94 expected (E); 95% Confidence Interval (CI) = 0.65-1.24). We observed a significant excess of colon cancer among the proband's first degree relatives (O/E = 5/1.64; 95% CI = 1.01-7.65), in particular those under age 45 year. Segregation analysis showed evidence for multifactorial inheritance in the small percentage (N = 5) of the families. ^

HEMATOCRIT AND HEMOGLOBIN, ATP AND DPG CONCENTRATIONS IN ANDEAN MAN: VARIABILITY BY SEX, AGE, VILLAGE, ALTITUDE, WEIGHT, SMOKING HABITS AND GENETIC CONSTITUTION

The Departmento de Arica in northern Chile was chosen as the investigation site for a study of the role of certain hematologic and glycolytic variables in the physiological and genetic adaptation to hypoxia.^ The population studied comprised 876 individuals, residents of seven villages at three altitudes: coast (0-500m), sierra (2,500-3,500m) and altiplano (> 4,000m). There was an equal number of males and females ranging in ages from six to 90 years. Although predominantly Aymara, those of mixed or Spanish origin were also examined. The specimens were collected in heparinized vacutainers precipitated with cold trichloroacetic acid (TCA) and immediately frozen to -196(DEGREES)C. Six variables were measured. Three were hematologic: hemoglobin, hematocrit and mean cell hemoglobin concentration. The three others were glycolytic: erythrocyte 2,3-diphosphoglycerate (DPG), adenosine triphosphate (ATP) and the percentage of phosphates (DPG + ATP) in the form of DPG.^ Hemoglobin and hematocrit were measured on site. The DPG and ATP content was assayed in specimens which had been frozen at -196(DEGREES)C and transported to Houston. Structured interviews on site provided information as to lifestyle and family pedigrees.^ The following results were obtained: (1) The actual village, rather than the altitude, of examination accounted for the greatest proportion of the variance in all variables. In the coast, a large difference in levels of ionic lithium in the drinking water exists. The chemical environment of food and drink is postulated to account, in part, for the importance of geographic location in explaining the observed variance. (2) Measurements of individuals from the two extreme altitudes, coast and altiplano, did not exhibit the same relationship with age and body mass. The hematologic variables were significantly related to both age and body build in the coast. The glycolytic variables were significantly related to age and body mass in the altiplano. (3) The environment modified male values more than female values in all variables. The two sexes responded quite differently to age and changes in body mass as well. The question of differing adaptability of the two sexes is discussed. (4) Environmental factors explained a significantly higher proportion of total variability in the altiplano than in the coast for hemoglobin, hematocrit and DPG. Most of the ATP variability at both altitudes is explained by genetic factors. ^

Effects of thymus size and involution on the contribution of recent thymic emigrants to the peripheral T cell pool

The contribution of recent thymic emigrants (RTEs) to the peripheral naïve T cell population is necessary to maintain diversity of the T cell receptor (TCR) repertoire and produce immune responses against newly encountered antigens. The thymus involutes with age, after irradiation or chemotherapy, and due to severe viral infections. Thymus involution results in decreased thymopoiesis and RTE output leading to a reduced diversity of peripheral T cells. This increases susceptibility to disease and impairs immune responsiveness to vaccines. Therefore, studies aimed at maintaining or regenerating thymic function are integral for maintaining and restoring peripheral TCR diversity. Mice that express a K5.CyclinD1 transgene expression have a severely hyperplastic thymus that fails to undergo involution. Both thymocyte and TEC development appear normal in these mice. We have used the K5.CyclinD1 transgenic model to test the hypothesis that preventing thymus involution will sustain RTE output and incorporation into the peripheral T cell pool to prevent naïve T cell depletion with age. The K5.CyclinD1 transgene was crossed to the RAG2p-GFP transgenic model so that RTEs could be tracked by the intensity of the GFP signal. The frequency and number of RTEs in naïve CD4 splenic T cells was analyzed at monthly intervals to 5 months of age. Using this double transgenic approach, we determined that preventing thymus involution does maintain or enhance the number of RTEs in the peripheral T cell pool before and after thymus involution.

ATXA -dependent gene expression in Bacillus anthracis

The Bacillus anthracis toxin genes, cya, lef , and pag, can be viewed as a regulon, in which transcription of all three genes is activated in trans by the same regulatory gene, atxA, in response to the same signal, CO2. I determined that several phenotypes are associated with the atxA gene. In addition to being toxin-deficient, an atxA -null mutant grows poorly on minimal media and sporulates early compared to the parent strain. Furthermore, an atxA-null mutant has an altered 2-D gel protein profile. I used a genetic approach to find additional atxA-regulated genes. Random transcriptional lacZ fusions were generated in B. anthracis using transposon Tn 917-LTV3. Transposon-insertion libraries were screened for mutants expressing increased β-galactosidase activity in 5% CO2. Introduction of an atxA-null mutation in these mutants revealed that 79% of the CO2-regulated fusions were also atxA-dependent. DNA sequence analysis of transposon insertion sites in mutants carrying CO 2/atxA-regulated fusions revealed ten mutants harboring transposon insertions in loci distinct from the toxin genes. The majority of the tcr (toxin co-regulated) loci mapped within the pXO1 pathogenicity island. These results indicate a clear association of atxA with CO2-enhanced gene expression in B. anthracis and provide evidence that atxA regulates genes other than the structural genes for the anthrax toxin proteins. ^ Characterization of one tcr locus revealed a new regulatory gene, pagR. The pagR gene (300 nt) is located downstream of pag. pagR is cotranscribed with pag and is responsible for autogenous control of the operon. pagR also represses expression of cya and lef. Repression of toxin gene expression by pagR may be mediated by atxA. The steady state level of atxA mRNA is increased in a pagR mutant. Recombinant PagR protein purified from Escherichia coli did not specifically bind the promoter regions of pagA or atxA. An unidentified factor in B. anthracis crude extracts, however, was able to bind the atxA promoter in the absence of PagR or AtxA. These investigations increase our knowledge of virulence regulation in B. anthracis and ultimately will lead to a better understanding of anthrax disease. ^