Extracts from interferon‐treated Ehrlich ascites tumor cells differ in various biochemical characteristics from extracts from control cells. Thus, as reported earlier, double‐stranded RNA (dsRNA) promotes the phosphorylation by ATP of at least two proteins in extracts from interferon‐treated cells, but not, or to only a lesser extent, in extracts from control cells. Moreover reovirus mRNAs are degraded faster in reaction mixtures containing extracts from interferon‐treated cells than in those containing extracts from control cells but only if the reaction mixtures are supplemented with dsRNA and ATP. The faster RNA degradation in extracts from interferon‐treated cells is due to enhanced endonuclease action. We designated the agent(s) catalyzing this as endonucleaseINT. There is some enhancement of nuclease activity by dsRNA and ATP also in extracts from control cells. The extent of this enhancement is, however, much smaller than in extracts from interferon‐treated cells. We report now that the promotion of mRNA cleavage in extracts from interferon‐treated cells by dsRNA and ATP is apparently not a consequence of a possible impairment in the attachment of the mRNA to ribosomes since (a) the protein synthesis inhibitors (sparsomycin and edeine) do not affect the degradation and (b) dsRNA and ATP enhance RNA degradation also in the 200000 × g supernatant fraction from extracts of interferon‐treated cells and this fraction is essentially free of ribosomes. (The ribosome concentration in our 200000 × g supernatant fraction is less than 1% of that in the 30000 × g extract.) GTP or the ATP analogs AMP(CH2)PP or AMP‐P(CH2)P do not substitute for ATP and dsDNA or DNA · RNA hybrids do not substitute for dsRNA in activating endonucleaseINT. The optimal concentration of ATP in activating endonucleaseINT is 1 mM or higher. The addition of dsRNA and ATP to extracts from interferon‐treated cells affects the degradation of some RNAs much more than that of others. Thus it promotes the degradation of reovirus mRNA from the large size class strongly, the degradation of those from the medium size class to an intermediate extent and that of those from the small size class the least. Furthermore it enhances the degradation of mRNA from either control or interferon‐treated Ehrlich ascites cells more than that of Ehrlich ascites cell ribosomal RNA or mouse globin mRNA. Reovirus dsRNA is not cleaved by endonucleaseINT. The partially purified mouse interferon preparation (specific activity 8 × 108 NIH reference standard units/mg protein) used in several experiments was free of nuclease activity under our experimental conditions. Moreover the addition to extracts from control cells (or extracts from interferon‐treated cells) of this interferon preparation together with dsRNA and ATP did not enhance nuclease activity.
|Number of pages||13|
|Journal||European Journal of Biochemistry|
|State||Published - Oct 1977|