Identification and characterization of genes regulated by interferons and tumor necrosis factor

Interferons (IFNs) and tumor necrosis factor(TNF) exert their biological activities through the modulation of gene expression. This thesis focuses on identification and characterization of genes whose expression is regulated by IFNs or TNF. Our studies reveal that: (A) Expression of the gene encoding the endoplasmic reticular heat shock protein gp96 is up-regulated transcriptionally in IFN-$\alpha$ and IFN-$\gamma$ treated HeLa cells as well as IFN-$\alpha$ treated Daudi cells. The induction of the gp96 gene is dependent upon de novo protein synthesis. The level of gp96 protein in IFN treated cells was monitored by immunoprecipitation and was observed to be elevated following IFN treatment. An imperfect ISRE (Interferon Stimulated Response Element) sequence element is present in the 5$\sp\prime$ region of gp96 gene. However, its imperfect nature appears to deem this sequence unresponsive to IFN stimulation. (B) two alternative polyadenylation sites are present in the 3$\sp\prime$-untranslated region of the tryptophanyl tRNA synthetase mRNA. When the polyadenylation occurs at the distal site it results in the production of a mRNA of approximately 3.1 kb, whereas, when the polyadenylation occurs at the proximal site a mRNA of approximately 2.3 kb is generated. (C) Employing Southern blot analysis, the human tryptophanyl tRNA synthetase gene has been determined to be located on chromosome 14 and the human indoleamine 2,3-dioxygenase gene on chromosome 8. (D) The mRNA level of three genes, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), SR protein 20KDa (SRp20) and protein disulfide isomerase-Q2 (PDI-Q2) are suppressed following TNF treatment. This suppression is observed to be due to reduced transcription of these mRNAs. The kinetics of the reduction in the level of these mRNAs reveals that the suppressive effect of TNF on GAPDH and SRp20 genes is sustained, while, the effect on PDI-Q2 appears to be transient. A sequence element which appears to be responsible for TNF-mediated down-regulation of various genes is found to be present in the first intron of GAPDH gene and two similar elements are found in the 5$\sp\prime$ region of SRp20 gene. No sequence element has been observed in the gene encoding the PDI-Q2