Selection and characterization of interferon response mutants from mouse and human cell lines
Abstract
Although the events of the interferon (IFN)-mediated antiviral state have been extensively characterized, much remains to be learned about how IFN inhibits specific viruses. The isolation of cellular IFN-response mutants has been important in studying the mode of action of IFN. In this thesis, IFN-response mutants were selected from HeLa cells and mouse L 929 fibroblast cells and their specific resistance (or lack of resistance) to is-1, an IFN-sensitive mutant of mengovirus, was studied. An IFN-responsive line of HeLa cells (F-H12), which was found to be up to 100 times more responsive to IFN than the parental line as measured by cell survival, was selected using is-1. This IFN-responsiveness appears to be specific for is-1 and was not seen with three other viruses. Two distinct antiviral activities have been detected in mouse L cells pretreated with low levels of IFN, and infected with is-1. The first antiviral activity (AVA-1) consists of all the antiviral activities induced by IFN which inhibit both is-1 and is+. The second antiviral activity (AVA-2) are those activities which are specific for is-1. Only 20% of randomly tested L cell lines exhibit AVA-2. From the standard L cell subline used in our laboratory (G3), which fully expresses AVA-2, two AVA-2 non-expressive clonal sublines (TA-6 and AS-4) were selected. Two AVA-2 revertant lines (TA-6 R1 and AS-4 R1) were later isolated from TA-6 and AS-4. The kinetics of is-1 growth in the presence of IFN was found to vary in each of these 5 sublines. The phenotype of the HeLa and mouse L cell sublines cannot be accounted for by major differences in (1) their ability to bind IFN, (2) enhanced IFN-induction by is-1, or (3) variations in levels of 2,5 oligo A dependent endonuclease. Although the IFN-responsiveness in the F-H12 line is quantitatively different from that of G3, there are several qualitative similarities. Hence some function analogous to AVA-2 appears to be induced in HeLa cells. IFN-pretreated G3 cells, exposed to a variety of inhibitors of mitochondrial function prior to or following infection with is-1, fail to develop AVA-2. $\alpha$-amanitin (a highly specific inhibitor of cellular mRNA synthesis), also inhibits AVA-2 activity. Therefore, AVA-2 may be comprised of more than one component, one associated with mitochondria and one nuclear. (Abstract shortened with permission of author.)
Degree
Ph.D.
Advisors
Simon, Purdue University.
Subject Area
Microbiology
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