NeST long ncRNA controls microbial susceptibility and chromatin activation at the interferon gamma locus [electronic resource]

The outcomes of most viral infections are controlled by many genetic loci whose interactions are complex and difficult to predict. To characterize the genes that predispose to viral persistence, I carried out a genetic analysis of a murine susceptibility locus termed Tmevp3. Mice that carry the SJL/J-derived Tmevp3 locus become persistently infected, whereas mice carrying the B10.S-derived locus clear the infection. My investigation revealed that a long noncoding RNA, lncRNA, transcript named NeST RNA was the most polymorphic gene in the locus. I found that a spliced version of NeST RNA was abundant in T cells of SJL/J mice and not expressed in B10.S mice. To test whether expression of this lncRNA was the cause of viral persistence, I developed transgenic mice that expressed NeST RNA in B10.S mice. Indeed, NeST RNA alone was sufficient to confer viral persistence. To investigate the function of NeST RNA in T cells, I tested whether NeST regulates its neighboring genes, IL-22 and IFN-[gamma]. CD8+ T cells from mice carrying the SJL/J-Tmevp3 locus were found to produce greater amounts of IFN-[gamma], suggesting that NeST RNA might functions as an enhancer lncRNA. Indeed, transgenic expression of NeST lncRNA induced higher of IFN-[gamma] after T cell stimulation. Results from a collaborative study revealed NeST RNA functions in trans, localize to the nucleus, associates with the histone modifying complex component WDR5, and induces accumulation of histone 3 lysine 4 trimethylation (H3K4me3) at the Ifng locus. I also found that the Tmevp3 locus controls lethality from Salmonella. However, I found that in this model the SJL/J allele, which increases viral persistence, conferred decrease susceptibility for pathogenesis. As was the case in the viral susceptibility model, transgenic NeST RNA expression was sufficient to recapitulate this susceptibility phenotype. To test for any effects of NeST RNA in the inflammatory response to pathogen antigens, I carried out septic shock assays using the bacterial component LPS. I found that mice expressing NeST RNA survived more compared to the B10.S parental strain. Take together these data suggest that during immune challenge NeST RNA regulates the inflammatory response. To further analyze the effect of NeST RNA in the control of inflammation, NeST knockout mice were utilized. Deleting a 640 base pair region in the NeST promoter was found to abrogate gene expression and increase lethality to LPS sepsis. This difference in lethality coincided with larger abundances of numerous cytokines in mice lacking NeST RNA after LPS injections. However in the absence of an immune challenge, mice with an intact NeST gene showed higher basal levels of inflammatory cytokines, suggesting the basis of NeST RNA protection is a hormetic effect. Indeed, pre-treatment with low doses of LPS phenocopies the endotoxin tolerance effect of NeST RNA. Understanding how NeST RNA controls the timing and magnitude of cytokine secretion by T cells may provide new venues for therapeutic intervention to various inflammatory diseases.

Keywords: Microbiology and Immunology.
Advisor: Kirkegaard Karla primary advisor. Thesis advisor
Brahic Michel advisor. Thesis advisor
Chang Howard
Cimprich Karlene advisor. Thesis advisor
Maecker Holden advisor. Thesis advisor