Mechanism of viroid pathogenesis: differential activation of the interferon-induced, double-stranded RNA-activated, M(r) 68,000 protein kinase by viroid strains of varying pathogenicity

Abstract

Purified potato spindle tuber viroid (PSTVd) was added to an in vitro assay system containing purified interferon-induced, dsRNA-activated protein kinase (P68). Viroid RNA activated (phosphorylated) the enzyme, although with less efficiency than did the synthetic, perfectly matched poly I-poly C. In binding experiments, RNA transcripts of the intermediate strain of PSTVd were shown to specifically bind to a P68-antibody complex. Activation of the enzyme by a strain of PSTVd that results in severe symptoms in infected tomato plants was at least ten-fold that by the mild strain. Activation by a strain that results in intermediate symptoms was quantitatively similar to activation by the severe strain. To our knowledge, this is the first demonstration of a differential effect of viroid strains inducing different levels of pathology on any biochemical or metabolic system investigated. This differential effect suggests that activation of a plant enzyme homologous to mammalian P68 protein kinase may represent the triggering event in viroid pathogenesis. Differential activation of P68 is surprising, because the primary structures of the mild and severe PSTVd strains analyzed differ by only a two-nucleotide inversion (UUC-->CUU) in the lower portion of the 'pathogenicity' region of the molecules. This change, according to thermodynamic calculations, should have only a minor effect on the secondary structure of the viroid molecule. Binding assays indicated that PSTVd specifically binds to P68.