Browsing by Author "Marutha, Tebogo Rector"

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    Genetic variation influencing mitochondrial DNA copy number and the development of sensory neuropathy in HIV-positive patients exposed to stavudine
    (2017) Marutha, Tebogo Rector
    Antiretroviral therapy (ART) drugs such as stavudine (d4T) are known to have off-target side-effects, including the inhibition of DNA polymerase gamma which replicates mitochondrial DNA (mtDNA). ART-induced depletion of mtDNA copy number may cause mitochondrial toxicities such as sensory neuropathy (SN). Genetic variation in DNA polymerase gamma or in other nuclear genes influencing mtDNA replication and mtDNA copy number may therefore contribute to susceptibility to d4T-induced SN. DNA samples from 263 HIV-positive South African adults exposed to d4T were classified as cases with SN (n = 143) and controls without SN (n = 120). A total of 28 single nucleotide polymorphism (SNPs) were chosen in nuclear genes from the mtDNA replication pathway and from a GWAS paper examining SNP association with ART-induced SN (Leger et al. 2014). Genotyping was performed using Sequenom Mass Spectrometry. MtDNA copy number was determined using a qPCR assay. Associations between SN and genetic variants, between genetic variants and mtDNA copy number, and between mtDNA copy number and SN were evaluated in univariate and multivariate models using Plink v1.07 and GraphPad v7. Age and height were significantly different in the cases with SN vs controls without SN. In univariate analyses, three SNPs and two haplotypes were significantly associated with SN, three SNPs were associated with pain intensity and three haplotypes were significantly associated with mtDNA copy number. However, there were no significant associations with SN, pain intensity or mtDNA copy number after correction for multiple SNP testing. No significant difference in mtDNA copy number in cases vs. controls was observed. In conclusion variation in nuclear-encoded mitochondrial genes examined in the current study do not play a role in ART-related mitochondrial complications such as changes in mtDNA copy number, or occurrence of SN.
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    Quantifying drought strategies in C4 perennial Southern African grasses
    (2024) Marutha, Tebogo Rector
    Plants have a range of strategies for surviving drought associated with the morphological and physiological traits that govern water use and loss. Most research has been done on woody species and whether the research applies in C4 grasses whose above-ground biomass cures annually over a dry season is not clear. In this study I described and quantified drought strategies and recovery patterns of thirteen perennial C4 grass species found in savanna and grassland ecosystems in South Africa. An experiment was undertaken where the grasses were grown and then exposed to a drought for three months, after which they were watered for one week to simulate a post drought recovery. I monitored stomatal conductance, leaf water potential, osmotic adjustment, leaf senescence and post-drought resprouting to assess whether plants with restricted water availability retained varying proportions of live leaf material or whether a wasteful water strategy during drought was associated with increased leaf death and post-drought recovery. Based on the measurements of the thirteen species, a broad range of strategies emerged related to the trade-offs between being efficient or safe, whereby some grasses control water loss by closing stomata and sacrificing photosynthesis while others continue to lose water while they photosynthesise. Additionally, osmotic adjustment demonstrated in species such Digitaria eriantha and Panicum maximum demonstrated an alternative drought strategy that is not currently recognised in the literature. Grasses with low stomatal control cured faster than those with a high stomatal control, which might have implications for flammability and leaf phenology in seasonal ecosystems with annual drought.
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    Regulation of PXDN in eye development and PXDN gene variant screening within a South African cohort of patients presenting with anterior segment dysgenesis
    (2024) Marutha, Tebogo Rector
    Peroxidasin (PXDN) is an extracellular matrix-associated haem-peroxidase predominantly expressed in the vasculature and eye. PXDN crosslinks collagen IV through sulfilimine bond formation in the basement membrane. Aberrant PXDN expression is associated with fibrosis, heart failure and cancer, and various pathologies of the eye, where PXDN likely provides structural support via basement membrane synthesis in the cornea and lens during eye development, as well as protect the lens, trabecular meshwork and cornea against oxidative damage. Furthermore, PXDN pathogenic variants have been associated with anterior segment dysgenesis (ASD), congenital cataracts and corneal opacity. To further understand the role of PXDN in the eye, first we aimed to identify PXDN as a novel target of key transcriptional regulators of eye development, namely PAX6, FOXC1 and PITX2, and second, to screen a cohort of South African patients with ASD to look for pathogenic variants in PXDN and other ASD genes. Protein expression of PAX6, FOXC1, PITX2 and PXDN, in response to Fibroblast Growth Factor-2 (FGF-2) were quantified by western blotting and localisation visualised using immunofluorescence confocal microscopy. Chromatin immunoprecipitation-PCR and luciferase assays were employed to detect transcription factor-PXDN promoter interactions. Expression data established that PXDN, PAX6, FOXC1 and PITX2 were induced by FGF2 at varying time points. Putative binding sites for all three transcription factors were identified in the PXDN promoter and ChIP-PCR confirmed that PAX6, FOXC1 and PITX2 interact with various regions of the promoter. Luciferase reporter assays are currently underway. Next Generation Sequencing of genomic DNA from South African patients exhibiting ASD disorders. identified disease causing variants in PAX6 and GJA8. Variants of uncertain significance were identified in PXDN, BCOR, EPHA2 and LTBP2 genes and are being investigated further. In conclusion, we identified PXDN as a novel target of PAX6, FOXC1 and PITX2 that further supports for the integral role of PXDN in eye development.