Interferons (IFNs) are polypeptides that protect the body from microbial infection by the activation of genes that inhibit the replication of such pathogens. STAT1 is a transcription factor that is activated by IFNs and virus infection, and propagates signals to the nucleus by binding DNA enhancer elements to induce the expression of IFN-stimulated genes (ISGs). One such gene product is the double-stranded RNA (dsRNA)dependent protein kinase, PKR. PKR is an important determinant of host resistance since it has the ability to bind dsRNA, an intermediate produced during viral replication, and to phosphorylate the eukaryotic translation initiation factor-2alpha (eIF-2alpha), a modification that inhibits of protein synthesis. Additionally, recent findings have shown that PKR can modulate various signaling pathways. The objective of this research was to understand the involvement of PKR in the IFN signaling pathways. We observed that the catalytic domain of PKR interacts with the DNA-binding region of STAT1 in vitro and in vivo. Overexpression of catalytic mutants of PKR blocked STAT1 DNA-binding and ISG expression upon IFN or dsRNA treatment. The capacity of catalytic mutants of PKR to negatively regulate STAT1 DNA-binding is attributed to its ability to interact with STAT1 since in cells lacking PKR or expressing an RNA-binding mutant of PKR, unable to associate with STAT1, STAT1 displays augmented activity. A mutant of STAT1 (TM) unable to interact with PKR displayed elevated biochemical and biological activities; findings consistent with the role of PKR in regulating STAT1 function. Alternatively, the interaction between PKR and STAT1 has the ability to inhibit PKR activity in vitro and in vivo; a phenomenon not observed with STAT1 TM. / In parallel, PKR was also found to interact with TYK2, the upstream activator of STAT1. IFN treatment promoted the dissociation of PKR from TYK2 in a manner dependent on TYK2 kinase activity. Furthermore, we observed that TYK2 phosphorylates the amino terminus of PKR in vitro and in vivo on tyrosine. Taken together, PKR appears to function as an important scaffolding protein for the IFN signaling pathways. Alternatively, PKR activity itself is also regulated at multiple steps along this pathway. Thus, it appears that there is an intimate cross-talk between components of the JAK/STAT pathway and translational machinery.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.36850 |
| Date | January 2000 |
| Creators | Wong, Andrew Hoi-Tao, 1974- |
| Contributors | Koromilas, Antonis E. (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Division of Experimental Medicine.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001771089, proquestno: NQ69948, Theses scanned by UMI/ProQuest. |
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