A cis-acting element in the 3 '-untranslated region of human TNF-alpha mRNA renders splicing dependent on the activation of protein kinase PKR

We report a role for the 3'-untranslated region in control of mRNA splicing and show that human TNF-alpha 3' UTR harbors a cis-acting element that ren ders splicing of precursor transcripts dependent on activation of PKR, the RNA-activated protein kinase that phosphorylates eukaryotic initiation fact or 2 (eIF2). When this element, designated 2-APRE, is present, splicing bec omes sensitive to inhibition by the PKR inhibitor e-aminopurine, or by coex pression of transdominant-negative mutant PKR. Our results reveal that acti vation of PKR is required for splicing of mRNA when precursor transcripts c ontain the 2-APRE and that increased expression of wild-type PKR enhances t heir splicing efficiency. Thus, PKR responds as trans-acting factor to the 2-APRE. 2-APRE RNA forms a stable, 17-bp stem-loop structure and strongly a ctivates PKR ill vitro, inducing eIF2 alpha phosphorylation. Despite its ab ility to activate PKR during splicing, the 2-APRE within the 3' UTR does no t affect translation efficiency of the resulting TNF-alpha mRNA in transfec ted cells. PKR and the 3' UTR thus interact during mRNA splicing to confer a novel type of regulation on expression of the TNF-alpha gene.