No description available.

No description available.

No description available.

Numerous genetic variants displaying stereotypic circling behaviour have been described in rodents. The majority are recessive mutants expressing dopaminergic alterations in the striatum- often with associated vestibular defects. We describe a novel circling mouse (Cr) with intact vestibular function frequently obtained from crosses of non-transgenic wild type mice (WT) and transgenic growth hormone mice (TGM). We have characterized Cr with stereotypic circling, head bobbing, hyperkinesia, aggressiveness, and elevated dopamine when compared to the transgenic growth hormone mouse. The Cr also demonstrates self-mutilating behaviour found in mice with dysregulated striatal dopamine levels. Cr oppositely mirrors most traits of the TGM including alterations in sleep, activity, eating, and drinking. TGM displayed superior performance than WT in novel object recognition, but this decreased with aging. Comparatively, Cr performed poorly in this test. The memory of young TGM exceeded controls, whereas young Cr displayed poorer memory with an age related improvement. In a stepdown test of emotionality, TGM step down more readily, while Cr are more hesitant than WT. TGM and Cr also demonstrate opposite alterations in striatal dopamine. Further analysis demonstrated differential responsiveness of TGM and Cr under dopaminergic drugs, and potentially sexually dimorphic differences in behaviour associated with elevated GH in TGM. TGM are characterized by increased levels of circulating growth hormone and alterations in sleep and activity. We hypothesize that frequent generation of Cr may reflect unintended selection of modifier genes that counterbalance negative consequences of elevated GH in TGM. / Thesis / Master of Science (MSc)

Inbreeding results in increased levels of homozygosity for deleterious recessive alleles, leading to increased incidence of monogenic disease in inbred families. It has also been suggested that inbreeding increases the risk of diseases such as cancer and heart disease, implying a role for the combined effects of many recessive alleles distributed across the genome. A better understanding of the links between inbreeding, homozygosity and disease is therefore of interest to those concerned with understanding the genetic architecture of complex disease. A homozygous genotype is defined as autozygous if both alleles originate from the same ancestor. Quantifying inbreeding involves quantifying autozygosity. A new, observational method of quantifying autozygosity using genomic data is developed here. Based on runs of homozygosity (ROH), this approach has a sound theoretical basis in the biological processes involved in inbreeding. It is also backed by strong empirical evidence, correlating strongly with pedigree-derived estimates of inbreeding and discriminating well between populations with different demographic histories. ROH are a signature of autozygosity, but not necessarily autozygosity of recent origin. Short ROH are shown to be abundant in demonstrably outbred individuals and it is suggested that this is a source of individual genetic variation which merits investigation as a disease risk factor, although denser genotype scans than those used in the present study are required for the reliable detection of very short ROH. In the absence of such dense scans, it is suggested that ROH longer than 1 or 1.5 Mb be used to estimate the effects of inbreeding on disease or quantitative physiological traits (QT), and that a simple measure of homozygosity be used to investigate overall recessive effects. Evidence for recessive effects on 13 QT important in cardiovascular and metabolic disease was investigated in 5 European isolate populations, characterised by heightened levels of inbreeding. A significant decrease in height was associated both with increased homozygosity and (to a lesser extent) with increased ROH longer than 5 Mb (i.e. inbreeding) estimated using a 300,000 SNP panel. No evidence was found for recessive effects on any of the other QTs. Evidence for recessive effects on colorectal cancer risk were investigated in two outbred case control samples typed with a 500,000 SNP panel. Cases were significantly more homozygous and had more of their genome in short ROH than did controls. Cases were significantly more homozygous than controls even when inbred individuals were removed from the sample. There was also some evidence of an inbreeding effect, with inbred subjects having slightly significantly higher odds of colorectal cancer than outbred subjects. This study provides evidence of recessive effects on a common, complex disease in outbred populations and on height in both inbred and outbred populations and shows that such effects are not solely attributable to increased levels of homozygosity resulting from recent inbreeding. Individual variation among outbred individuals in the proportion of the genome that is homozygous may be important in disease risk. The development of denser genotype scans will facilitate better enumeration of short ROH in outbred individuals so that these can be properly enumerated and investigated as a disease risk factor.

No description available.

MSc (Med)(Genetic Counselling), Faculty of Health Sciences, University of the Witwatersrand / Autosomal recessive polycystic kidney disease (ARPKD; MIM263200) is a severe recessively inherited disease of the kidneys and biliary tract, with an incidence of approximately 1 in 20000 in non-isolated populations. It has a variable clinical spectrum from neonatal demise (in 30-50%) to survival into adulthood. ARPKD is caused by mutations at a single locus, polycystic kidney and hepatic disease 1 (PKHD1), with over 270 pathogenic mutations described to date. The high rate of compound heterozygosity in affected individuals has made genotype-phenotype correlations difficult. A common missense mutation, p.M627K, in exon 20 of PKHD1 was identified previously on the majority of ARPKD disease associated alleles in the Afrikaans population of South Africa suggesting the presence of a founder effect. The aim of this study was to describe the clinical phenotype of ARPKD in Afrikaans speaking individuals found to be homozygous for the common mutation, and to compare this phenotype to previously described cohorts of patients with ARPKD, known to harbour a spectrum of mutations. This descriptive study used retrospective data collected from records of patients with ARPKD at Johannesburg and Pretoria Academic Hospitals. Twenty seven individuals from 24 families were included in the study. Marked clinical variability was demonstrated within this subject group supporting the limitation of genotype-phenotype correlation described worldwide. ARPKD was diagnosed at a median age of 27 days, older than a North American cohort (NAC) born after 1990 (median age of 1 day). The majority (93%) of subjects in this study were diagnosed with chronic renal v insufficiency (CRI) and hypertension (HT), indicating the renal morbidities to be more common than noted in previous studies, but occurring at a later median age (1.4 years vs 13.5 days in the NAC). This may indicate a trend toward milder expression of renal morbidities in the present study. Portal hypertension was also diagnosed more frequently (81%) than in previous studies but at a younger median age (1.3 years vs 2.8 years), although with similar complication rates. Overall statistical correlation was found between the renal and hepatic related morbidities in this study, indicating that progression of the condition is not organ specific. A survival rate of 89% at one year is comparable to previous studies with similar patient ascertainment. This cohort represents the largest series of patients affected by ARPKD with a common mutation, described to date. The findings will provide for more accurate, specific and informative genetic counselling in families with ARPKD and may present a resource for future studies of modifier genes and environmental influences on the phenotypic expression of ARPKD.

>Magister Scientiae - MSc / Molecular genetics: strategies to indentify congenital cataract genes in captive-bred Vervet monkeys Zandisiwe Emilia Magwebu MSc thesis, Department of Medical Biosciences, University of the Western Cape The present study describes molecular aspects of inherited congenital cataract in captive-bred Vervet monkeys. Congenital cataracts are lens opacities that are present at birth or soon after birth and include hereditary cataracts or cataracts caused by infectious agents. The MRC Primate Unit is housing a colony of captive-bred Vervet monkeys in which 7.5% is suffering from congenital cataract. However, the parents of the affected individuals were asymptomatic. Six families within the colony have been identified to be affected by two types of morphologies (Ysutural and total cataract). Based on the evidence provided above, it was speculated that the colony was affected with autosomal recessive cataract. The main aim of this study was to facilitate a strategy for managing breeding programs by minimizing cataract occurrences in captive-bred Vervet monkeys. Integrated combination of clinical, molecular and bioinformatic strategies were used to identify and assess reciprocal candidate susceptibility genes for cataracts. The genes that are known to be responsible for most human congenital cataract cases were prioritized. The genes include Heat shock transcription factor 4 (HSF4), Crystalline Alpha A (CRYAA), glucosaminyl (N-acetyl) transferase 2 (GCNT2) and Lens intrinsic membrane protein 2 (LIM2). Twenty two subjects were selected based on their morphology (5 carriers, 5 controls and 12 cataracts). 2ml of blood was collected for Deoxyribonucleic acid (DNA) extraction. Coding exons and flanking regions were screened by polymerase chain reaction (PCR) amplification and sequenced. The CLC DNA workbench was used for results analysis. The screening of four genes revealed 20 sequence variants which were not present in the control individuals. Sequencing of HSF4 revealed three mutations: R116R, L245>L and P421>L in exon 5, 10 and 14, respectively. The coding exons for CRYAA showed two sequence variants: S134W and K166N in exon 3. Twelve mutations were identified in exon one of all three GCNT2 transcripts (A, B and C). These mutations include: G212G, H256>H, M258>V, N275>N, V16>I, Y122>F, S15>S, S24>N, S38>S, I118>I, D194>D and Y373>Y which was found in exon three of all transcripts. There were no mutations in LIM2, however, three single nucleotide polymorphisms (SNPs) were identified in exon 2 (P66>P) and 3 (I118>T and A127>T). The above mutations were conserved when aligned with other species. The sequence variations vary among the families and those individuals with the same or different cataract phenotype. Based on these findings, it can be concluded that the four candidate genes harbour mutations that are responsible for both phenotypes. The effect of these mutations in Vervet monkeys is not yet understood, however, their impact will be further investigated. For future studies, it will be of absolute importance to screen the entire family to verify that indeed cataract formation in this colony is inherited in an autosomal recessive manner.

High-throughput sequencing technologies have changed the way we identify, study and understand the role of rare variation in Mendelian diseases. Sequencing in complex diseases have proven to be more challenging to interpret, but methods and approaches are being developed to aid in our understanding of variation in these diseases.

The number of known monogenic rare diseases (~7000) exceeds the number of effective treatments (~500) by more than an order of magnitude underlining the pressing need for generalizable therapeutic approaches for this class of conditions. In this regard, the majority of recessive and x-linked recessive disorders are caused by missense mutations encoding proteins that frequently have residual function but are rapidly degraded by the 26S proteasome. Bortezomib is a small molecule that inhibits the 26S proteasome and has been approved for use in patients for an unrelated condition; multiple myeloma. Previous work has shown that, for a small number of disorders, bortezomib can inhibit the degradation of the mutant protein, thereby increasing the protein level and activity, holding out the promise of a beneficial therapeutic effect by the repurposing of this agent. We present here a high level western blot based survey of nine recessive disorders to characterize the general effectiveness of such an approach. Thirteen patient fibroblast cell lines comprising 9 different diseases with 19 known mutations were selected on the basis of missense mutations protein expression data when available. The cell lines were incubated with bortezomib (10 nM and 50 nM; 24 hrs) and levels of the mutated protein were quantified by western blot. Unfortunately, no consistent, appreciable increase was observed for any of the conditions tested. The general therapeutic value of re-purposing bortezomib for recessive and x-linked diseases appears limited at best. The few reported cases of bortezomib successfully working in increasing mutated protein levels appear to be the exceptions and not the norm. Moreover successes are more often limited to cell lines carrying a transgene expressing the mutated protein rather than endogenous mutated protein in patient cell lines.