A diminished activation capacity of the interferon-inducible protein kinase PKR in human T lymphocytes

The double-stranded (ds) RNA activated protein kinase PKR is an interferon (IFN)-inducible serine/threonine protein that regulates protein synthesis t hrough the phosphorylation of the alpha subunit of translation initiation f actor 2 (eIF-2 alpha). PKR activation in cells is induced by virus infectio n or treatment with dsRNA and is modulated by a number of viral and cellula r factors. To better understand the mechanisms of PKR action we have analyz ed and compared the mode of PKR activation in a number of cell lines of dif ferent histological origin. Here we show that PKR activation and phosphoryl ation of eIF-2 alpha are both diminished in various virus-transformed and n ontransformed human T cells. Priming of T cells with IFN does not restore P KR activation. In vitro kinase assays show that the diminished PKR activati on in T cells correlates with the presence of a 60-kDa (p60) phosphoprotein coimmunoprecipitated with PKR. P60 is absent from PKR immunoprecipitates f rom non T cells. Incubation of active PKR with T cell extracts results in i nhibition of PKR autophosphorylation, which is proportional to the amount o f phosphorylated p60 in the kinase reactions. Treatment of T cells with pro teasome inhibitors or incubation of PKR immunoprecipitates with phosphatase inhibitors does not restore PKR activation. However, phosphorylation of p6 0 is enhanced upon treatment with the phosphatase inhibitor microcystin. Th ese data show that the impaired activation capacity of PKR in human T cells is exerted at the post-translational levels in a manner that is independen t of cell transformation or virus infection.