The vitamin D receptor (VDR) is a transcription factor mediating genomic responses to the biologically active form of vitamin D, 1,25(OH)2D3, a key modulator of the immune system. Knowledge on how these polymorphisms modulate the vitamin D endocrine system and confer risk of disease is hindered by the fact that several of the associated allelic variants are located in introns or are synonymous and likely serve as markers within an extended haplotype covering disease-causing alleles. The functional relevance of VDR polymorphisms need to be studied in the context of the haplotype, comparing haplotypes with the process of DNA transcription, protein levels and biological function. These functional studies should be performed using techniques reflecting the in vivo, naturally occurring milieu as close as possible. VDR has several known allelic variants including a FokI restriction fragment length polymorphism in exon II, BsmI and ApaI polymorphisms in the intron VIII, and a synonymous TaqI variant in exon IX. The aim of the current study was the identification of sequence variants in VDR, to define haplotype patterns in the Caucasian population and to understand the functional consequences of single nucleotide polymorphisms (SNPs) and haplotypes. Methods: EBV transformed B-lymphocyte cell lines, from twenty-three individuals, within the Caucasian population were established. Polymorphisms and haplotypes in VDR were identified by genotyping and sequencing. Quantification of the VDR protein level measured with flow cytometry was studied together with the genotype and haplotype data to determine possible influence of genotypes or haplotype and VDR protein levels. Biological function was analysed by the percentage inhibition that each individual experienced in the presence of 1,25(OH)2D3, measured by the Alamar Blue assay and the Trypan blue dye exclusion method. Results: The results showed thatVDR genotypes and haplotypes may not influence VDR protein level although certain genotypes and haplotypes significantly influenced biological function. It was proposed that VDR variants may account for significant influences on cellular responsiveness to 1,25(OH)2D3 as mediated by VDR. Conclusions: The findings of the current study suggest that individual SNPs and haplotypes of VDR influences quality of the repose in the presence of 1,25(OH)2D3, rather than quantity of the VDR levels. This knowledge may permit a rational choice of polymorphisms to use in epidemiology studies or improve our understanding of the significance of VDR genetic polymorphisms on biological function. Keywords: functionality, polymorphism, haplotype, vitamin D receptor, VDR, 1,25(OH)2D3, structure-function analysis, biological responsiveness. / Prof. L. Bornman
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:uj/uj:7259 |
| Date | 15 May 2008 |
| Source Sets | South African National ETD Portal |
| Detected Language | English |
| Type | Thesis |
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