REGULATION OF INTERFERON-INDUCED PROTEIN-KINASE PKR - MODULATION OF P58(IPK) INHIBITORY FUNCTION BY A NOVEL PROTEIN, P52(RIPK)

The cellular response to environmental signals is largely dependent up on the induction of responsive protein kinase signaling pathways, With in these pathways, distinct protein-protein interactions play a role i n determining the specificity of the response through regulation of ki nase function, The interferon-induced serine/threonine protein kinase, PKR, is activated in response to various environmental stimuli, Like many protein kinases, PKR is regulated through direct interactions wit h activator and inhibitory molecules, including P58(IPK), a cellular P KR inhibitor. P58(IPK) functions to represses PKR-mediated phosphoryla tion of the eukaryotic initiation factor 2 alpha subunit (eIF-2 alpha) through a direct interaction, thereby relieving the PKR-imposed block on mRNA translation and cell growth. To further define the molecular mechanism underlying regulation of PKR, we have utilized an interactio n cloning strategy to identify a novel cDNA encoding a P58(IPK)-intera cting protein. This protein, designated P52(rIPK), possesses limited h omology to the charged domain of Hsp90 and is expressed in a wide rang e of cell lines, p52(rIPK) and P58(IPK) interacted in a yeast two-hybr id assay and were recovered as a complex from mammalian cell extracts, When coexpressed with PKR in yeast, p58(IPK) repressed PKR-mediated e IF-2 alpha phosphorylation, inhibiting the normally toxic and growth-s uppressive effects associated with PKR function, Conversely, introduct ion of P52(rIPK) into these strains resulted in restoration of both PK R activity and eIF-2 alpha phosphorylation, concomitant with growth su ppression due to inhibition of P58(IPK) function. Furthermore, P52(rIP K) inhibited p58(IPK) function in a reconstituted in vitro PKR-regulat ory assay, Our results demonstrate that p58(IPK) is inhibited through a direct interaction with p52(rIPK) Which, in turn, results in upregul ation of PKR activity, Taken together, our data describe a novel prote in kinase-regulatory system which encompasses an intersection of inter feron-, stress-, and growth-regulatory pathways.