Expression and sequence variation of the porcine NRAMP1 gene

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Stumbaugh, Amber Nicole
Major Professor
Tuggle, Christopher K.
Committee Member
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Improving innate immunity is a natural target for marker-assisted selection in the pig because of its potential to improve herd resistance to many different pathogens, and thus to reduce production loss due to infection. The function of the NRAMP1 gene in infection in the pig is of interest to pork producers as it has been demonstrated to have a major impact on the outcome of infection with broad classes of pathogens in several host species. This thesis has undertaken the broad goal of adding to the understanding of the NRAMP1 gene structure and expression in the pig. To accomplish this goal, a comprehensive review of the literature available for the NRAMP1 gene begins the thesis. In the second chapter, the first research paper used TaqMan[Registered Trade Mark Symbol] real time RT-PCR to examine the changes in NRAMP1 expression in several porcine tissues during Salmonella choleraesuis infection, and revealed that NRAMP1 expression was upregulated in all tissues examined during infection. While the level of NRAMP1 mRNA in each tissue was correlated with several indicators of inflammation, including rectal temperature and interferon gamma concentration, there was no correlation between NRAMP1 expression levels and bacterial load. These data indicate NRAMP1 expression increased as a consequence of host response to infection rather than as a direct response to the presence of bacteria. The second research paper in this thesis examined the primary structure of the NRAMP1 gene, and confirmed the existence of six polymorphisms in the NRAMP1 mRNA sequence. In addition, two polymorphisms in the putative regulatory region of the gene were identified and confirmed by restriction enzyme digestion. One appendix to the thesis provides a multi-species alignment of the putative regulatory region in the NRAMP1 gene, and highlights the evolutionary conservation of the region adjacent to the start of the cDNA sequence, including several transcription factor binding sites. A second appendix lists the genotypic frequencies of the two non-coding region NRAMP1 polymorphisms in several populations. These papers add to the body of knowledge both about the regulation of NRAMP1 transcription in vivo during infection and about the structural variation in the gene.