Browsing by Author "Kinnear, Craig"
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- ItemAltered mitochondrial respiration and other features of mitochondrial function in parkin-mutant fibroblasts from parkinson’s disease patients(Hindawi Publishing Corporation, 2016) Haylett, William; Swart, Chrisna; Van der Westhuizen, Francois; Van Dyk, Hayley; Van der Merwe, Lize; Van der Merwe, Celia; Loos, Ben; Carr, Jonathan; Kinnear, Craig; Bardien, SorayaMutations in the parkin gene are the most common cause of early-onset Parkinson’s disease (PD). Parkin, an E3 ubiquitin ligase, is involved in respiratory chain function, mitophagy, and mitochondrial dynamics. Human cellular models with parkin null mutations are particularly valuable for investigating the mitochondrial functions of parkin. However, published results reporting on patient-derived parkin-mutant fibroblasts have been inconsistent. This study aimed to functionally compare parkin-mutant fibroblasts from PD patients with wild-type control fibroblasts using a variety of assays to gain a better understanding of the role of mitochondrial dysfunction in PD. To this end, dermal fibroblasts were obtained from three PD patients with homozygous whole exon deletions in parkin and three unaffected controls. Assays of mitochondrial respiration, mitochondrial network integrity, mitochondrial membrane potential, and cell growth were performed as informative markers of mitochondrial function. Surprisingly, it was found that mitochondrial respiratory rates were markedly higher in the parkin-mutant fibroblasts compared to control fibroblasts (p = 0.0093), while exhibiting more fragmented mitochondrial networks (). Moreover, cell growth of the parkin-mutant fibroblasts was significantly higher than that of controls (). These unanticipated findings are suggestive of a compensatory mechanism to preserve mitochondrial function and quality control in the absence of parkin in fibroblasts, which warrants further investigation.
- ItemExome sequencing identifies a novel MAP3K14 mutation in recessive atypical combined immunodeficiency(Frontiers, 2017-11) Schlechter, Nikola; Glanzmann, Brigitte; Hoal, Eileen Garner; Schoeman, Mardelle; Petersen, Britt-Sabina; Franke, Andre; Lau, Yu-Lung; Urban, Michael; Van Helden, Paul David; Esser, Maria Esser; Moller, Marlo; Kinnear, CraigENGLISH ABSTRACT: Primary immunodeficiency disorders (PIDs) render patients vulnerable to infection with a wide range of microorganisms and thus provide good in vivo models for the assessment of immune responses during infectious challenges. Priming of the immune system, especially in infancy, depends on different environmental exposures and medical practices. This may determine the timing and phenotype of clinical appearance of immune deficits as exemplified with early exposure to Bacillus Calmette-Guérin (BCG) vaccination and dissemination in combined immunodeficiencies. Varied phenotype expression poses a challenge to identification of the putative immune deficit. Without the availability of genomic diagnosis and data analysis resources and with limited capacity for functional definition of immune pathways, it is difficult to establish a definitive diagnosis and to decide on appropriate treatment.
- ItemExome sequencing identifies a novel TTC37 mutation in the first reported case of Trichohepatoenteric syndrome (THE-S) in South Africa(BioMed Central, 2017-03-14) Kinnear, Craig; Glanzmann, Brigitte; Banda, Eric; Schlechter, Nikola; Durrheim, Glenda; Neethling, Annika; Nel, Etienne; Schoeman, Mardelle; Johnson, Glynis; Van Helden, Paul D.; Hoal, Eileen G; Esser, Monika; Urban, Michael; Moller, MarloBackground Trichohepatoenteric syndrome (THE-S) or phenotypic diarrhoea of infancy is a rare autosomal recessive disorder characterised by severe infantile diarrhoea, facial dysmorphism, immunodeficiency and woolly hair. It was first described in 1982 in two infants with intractable diarrhoea, liver cirrhosis and abnormal hair structure on microscopy. We report on two siblings from a consanguineous family of Somali descent who, despite extensive clinical investigation, remained undiagnosed until their demise. The index patient died of fulminant cytomegalovirus pneumonitis at 3 months of age. Methods Whole exome sequencing (WES) was performed on a premortem DNA sample from the index case. Variants in a homozygous recessive state or compound heterozygous state were prioritized as potential candidate variants using TAPER™. Sanger sequencing was done to genotype the parents, unaffected sibling and a deceased sibling for the variant of interest. Results Exome sequencing identified a novel homozygous mutation (c.4507C > T, rs200067423) in TTC37 which was confirmed by Sanger sequencing in the index case. The identification of this mutation led to the diagnosis of THE-S in the proband and the same homozygous variant was confirmed in a male sibling who died 4 years earlier with severe chronic diarrhoea of infancy. The unaffected parents and sister were heterozygous for the identified variant. Conclusions WES permitted definitive genetic diagnosis despite an atypical presentation in the index case and suggests that severe infection, likely secondary to immunodeficiency, may be a presenting feature. In addition definitive molecular diagnosis allows for genetic counseling and future prenatal diagnosis, and demonstrates the value of WES for post-mortem diagnosis of disorders with a non-specific clinical presentation in which a Mendelian cause is suspected.
- ItemHuman whole genome sequencing in South Africa(Nature, 2021-01) Glanzmann, Brigitte; Jooste, Tracey; Ghoor, Samira; Gordon, Richard; Mia, Rizwana; Mao, Jun; Li, Hao; Charls, Patrick; Douman, Craig; Kotze, Maritha J.; Peeters, Armand V.; Loots, Glaudina; Esser, Monika; Tiemessen, Caroline T.; Wilkinson, Robert J.; Louw, Johan; Gray, Glenda; Warren, Robin M.; Moller, Marlo; Kinnear, CraigThe advent and evolution of next generation sequencing has considerably impacted genomic research. Until recently, South African researchers were unable to access affordable platforms capable of human whole genome sequencing locally and DNA samples had to be exported. Here we report the whole genome sequences of the first six human DNA samples sequenced and analysed at the South African Medical Research Council’s Genomics Centre. We demonstrate that the data obtained is of high quality, with an average sequencing depth of 36.41, and that the output is comparable to data generated internationally on a similar platform. The Genomics Centre creates an environment where African researchers are able to access world class facilities, increasing local capacity to sequence whole genomes as well as store and analyse the data.
- ItemIdentification of a novel functional deletion variant in the 5'-UTR of the DJ-1 gene(BioMed Central, 2009-10) Keyser, Rowena J.; Van der Merwe, Lize; Venter, Mauritz; Kinnear, Craig; Warnich, Louise; Carr, Jonathan; Bardien, SorayaBackground: DJ-1 forms part of the neuronal cellular defence mechanism against oxidative insults, due to its ability to undergo self-oxidation. Oxidative stress has been implicated in the pathogenesis of central nervous system damage in different neurodegenerative disorders including Alzheimer's disease and Parkinson's disease (PD). Various mutations in the DJ-1 (PARK7) gene have been shown to cause the autosomal recessive form of PD. In the present study South African PD patients were screened for mutations in DJ-1 and we aimed to investigate the functional significance of a novel 16 bp deletion variant identified in one patient. Methods: The possible effect of the deletion on promoter activity was investigated using a Dual- Luciferase Reporter assay. The DJ-1 5'-UTR region containing the sequence flanking the 16 bp deletion was cloned into a pGL4.10-Basic luciferase-reporter vector and transfected into HEK293 and BE(2)-M17 neuroblastoma cells. Promoter activity under hydrogen peroxide-induced oxidative stress conditions was also investigated. Computational (in silico) cis-regulatory analysis of DJ-1 promoter sequence was performed using the transcription factor-binding site database, TRANSFAC via the PATCH™ and rVISTA platforms. Results: A novel 16 bp deletion variant (g.-6_+10del) was identified in DJ-1 which spans the transcription start site and is situated 93 bp 3' from a Sp1 site. The deletion caused a reduction in luciferase activity of approximately 47% in HEK293 cells and 60% in BE(2)-M17 cells compared to the wild-type (P < 0.0001), indicating the importance of the 16 bp sequence in transcription regulation. The activity of both constructs was up-regulated during oxidative stress. Bioinformatic analysis revealed putative binding sites for three transcription factors AhR, ARNT, HIF-1 within the 16 bp sequence. The frequency of the g.-6_+10del variant was determined to be 0.7% in South African PD patients (2 heterozygotes in 148 individuals). Conclusion: This is the first report of a functional DJ-1 promoter variant, which has the potential to influence transcript stability or translation efficiency. Further work is necessary to determine the extent to which the g.-6_+10del variant affects the normal function of the DJ-1 promoter and whether this variant confers a risk for PD.
- ItemMulti-phenotype genome-wide association study of clades causing tuberculosis in a Ghanaian- and South African cohort(Elsevier Inc., 2021-04) Müller, Stephanie J.; Haiko, Schurz; Tromp, Gerard; Van der Spuy, Gian D.; Hoal, Eileen G.; Van Helden, Paul D.; Owusu-Dabo, Ellis; Meyer, Christian G.; Muntau, Birgit; Thye, Thorsten; Niemann, Stefan; Warren, Robin M.; Streicher, Elizabeth; Muller, Marlo; Kinnear, CraigDespite decades of research and advancements in diagnostics and treatment, tuberculosis remains a major public health concern. New computational methods are needed to interrogate the intersection of host- and bacterial genomes. Paired host genotype datum and infecting bacterial isolate information were analysed for associations using a multinomial logistic regression framework implemented in SNPTest. A cohort of 853 admixed South African participants and a Ghanaian cohort of 1359 participants were included. Two directly genotyped variants, namely rs529920 and rs41472447, were identified in the Ghanaian cohort as being statistically significantly associated with risk for infection with strains of different members of the MTBC. Thus, a multinomial logistic regression using paired host-pathogen data may prove valuable for investigating the complex relationships driving infectious disease.
- ItemNeutrophils : innate effectors of TB resistance?(Frontiers Media, 2018) Kroon, Elouise E.; Coussens, Anna K.; Kinnear, Craig; Orlova, Marianna; Moller, Marlo; Seeger, Allison; Wilkinson, Robert J.; Hoal, Eileen G.; Schurr, ErwinENGLISH ABSTRACT: Certain individuals are able to resist Mycobacterium tuberculosis infection despite persistent and intense exposure. These persons do not exhibit adaptive immune priming as measured by tuberculin skin test (TST) and interferon-γ (IFN-γ) release assay (IGRA) responses, nor do they develop active tuberculosis (TB). Genetic investigation of individuals who are able to resist M. tuberculosis infection shows there are likely a combination of genetic variants that contribute to the phenotype. The contribution of the innate immune system and the exact cells involved in this phenotype remain incompletely elucidated. Neutrophils are prominent candidates for possible involvement as primers for microbial clearance. Significant variability is observed in neutrophil gene expression and DNA methylation. Furthermore, inter-individual variability is seen between the mycobactericidal capacities of donor neutrophils. Clearance of M. tuberculosis infection is favored by the mycobactericidal activity of neutrophils, apoptosis, effective clearance of cells by macrophages, and resolution of inflammation. In this review we will discuss the different mechanisms neutrophils utilize to clear M. tuberculosis infection. We discuss the duality between neutrophils' ability to clear infection and how increasing numbers of neutrophils contribute to active TB severity and mortality. Further investigation into the potential role of neutrophils in innate immune-mediated M. tuberculosis infection resistance is warranted since it may reveal clinically important activities for prevention as well as vaccine and treatment development.
- ItemObsessive-compulsive disorder : defining the role of gene-based variants and immunological factors(Stellenbosch : Stellenbosch University, 2001-03) Kinnear, Craig; Corfield, V. A.; Stellenbosch University. Faculty of Medicine & Health Sciences. Dept. of Biomedical Sciences. Molecular biology and Human Genetics.ENGLISH ABSTRACT: Please see fulltext for abstract