rRNA cleavage as an index of ppp(A2'p)nA activity in interferon-treated encephalomyocarditis virus-infected cells

Abstract

In cell-free systems, 2-5A [ppp(A2'p)nA, n = 2 to greater than or equal to 4] activates a latent endoribonuclease, the 2-5A-dependent RNase, which cleaves rRNA in intact ribosomes into discrete and characteristic products (D. H. Wreschner et al., Nucleic Acids Res. 9:1571-1581, 1981). Here we present Northern blots which have identified the 18S or 28S origins of the cleaved products from rRNA. In addition, identical 3' termini were observed for fragments of 18S rRNA from a HeLa cell-free system incubated with 2-5A and from interferon-treated, encephalomyocarditis virus-infected HeLa cells. The previous assumption of identity of such fragments was based only on comigration on electrophoresis in agarose gels. We conclude that appropriate patterns of cleavage found in RNA isolated from intact cells are an indicator of prior 2-5A-dependent RNase activity. The assay of rRNA cleavage is relatively convenient and unambiguous. Accordingly, in the search for situations in which the 2-5A system may be active, it provides a useful alternative to the direct assay of 2-5A.