An Interferon-Induced, Ribosome-Associated Protein Kinase Which Reduces the Activity of Initiation Factor1

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Abstract
Treatment of mouse L cells with mouse interferon resulted in an increase of ribosomal associated protein kinase activity. The increase was completely blocked by actinomycin-D or cycloheximide, and heterologous human interferon did not induce an increase of protein kinase activity in the mouse L cell system. Partial purification of protein kinases from a high salt wash ribosomal fraction from interferon-treated cells by ion exchange chromatography and glycerol density gradient centrifugation showed that the molecular weight of one of the protein kinases (designated as enzyme I) was 85,000–90,000 daltons and that of the other (designated as enzyme II) was 50,000–55,000 daltons. The specific activities of enzymes I and II from interferon-treated cells were about 2 and 9 times higher, respectively, than those from untreated cells. For optimal activities, both enzymes required divalent cations such as Mg2+ and Mn2+, and sulfhydryl reagents such as dithiothreitol (DTT). Low concentrations of dsRNA (50–100 ng/ml) markedly enhanced the activity of enzyme II, but did not affect the activity of enzyme I. Aminopurine (2 mM) and N-ethylmaleimide (5 mM) inhibited more than 90% of the activities of these enzymes. Both enzymes phosphorylated calf thymus histone, casein, or purified initiation factor (eIF-2) with [γ-32P]ATP. The eIF-2 activity (binding of eIF-2 with Met-tRNAf and GTP) was significantly inhibited when eIF-2 was preincubated with enzyme II (from interferon-treated cells) in the presence of ATP (1 mM). The reduction of eIF-2 activity by enzyme II in the presence of ATP was strongly stimulated by low concentrations of dsRNA (50–100 ng/ml), and it was completely prevented by addition of aminopurine (2 mM). These findings suggest that the reduction of eIF-2 activity by enzyme II may involve phosphorylation of eIF-2 by this interferon-induced protein kinase in the presence of ATP. The mechanism of inhibition of eIF-2 activity by interferon-induced protein kinase is similar to that of the inhibition of polypeptide chain initiation induced by HRI (cAMP independent protein kinase) in rabbit reticulocyte lysates.

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