Regulation of iNOS gene expression in human T cells

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Nitric oxide (NO) is a bioactive gas involved in many aspects of human physiology. It is synthesized in cells by three related enzymes: neuronal nitric oxide synthase, inducible nitric oxide synthase (iNOS), and endothelial NOS. While all of these enzymes are important in the generation of NO under different biological circumstances, I have recently identified a role for iNOS expression in human T cells in the augmentation of human immune responses and shown that the chemokine stromal cell derived factor-1alpha (SDF-1a) is a protein inducer of iNOS in human T cells. However, the intracellular mechanisms regulating iNOS expression in T cells are unknown. Also, the regulation of iNOS expression in human cells, in general, is incompletely defined and is very different than that in animals commonly used to study mammalian biology, namely mice and rats. Therefore, the goal of my proposal is to understand the regulation of SDF-1a-induced iNOS expression in human T cells, and the role this plays in human T cell biology. The first part will study the intracellular signaling pathways that regulate SDF-1a-induced iNOS expression in human T cells. This will involve the inhibition of expression of the signaling enzymes ERK1/2 and PI3K by siRNA molecules, and then examination of SDF-1a-induced iNOS expression in human T cells lacking expression of ERK1/2 and PI3K. The role of Ras in mediating a potential feedback regulatory loop will also be examined. The second part of the proposal will determine the DNA elements and related transcription factors controlling the expression of the iNOS gene in human T cells. Finally, the third part of the proposal will examine the role of SDF-1a-induced iNOS in human T cell survival and migration. I have recently found that NO can inhibit cell death of human T cells and I will examine whether NO produced from iNOS in response to SDF-1a acts in a similar manner. The described work will be of interest to cell biologists and immunologists as it will provide new information on the regulation of iNOS expression in human cells, and insight into T cell biology as it relates to human immunity.