The B56alpha regulatory subunit of protein phosphatase 2A is a target for regulation by double-stranded RNA-dependent protein kinase PKR.
PKR is a cellular serine/threonine kinase that phosphorylates eukaryotic translation initiation factor 2alpha (eIF2alpha) to regulate protein synthesis. PKR also plays a role in the regulation of transcription, programmed cell death and the cell cycle, processes which likely involve other substrates. In a yeast two-hybrid screen, we isolated human protein phosphatase 2A (PP2A) regulatory subunit B56alpha as a PKR-interacting protein. The interaction between B56alpha and PKR was confirmed by in vitro binding assays as well as by in vivo coimmunoprecipitation, and this interaction is dependent on the catalytic activity of PKR. Moreover, recombinant B56alpha was efficiently phosphorylated by PKR in vitro and an isoelectric point shift in B56alpha was detected in extracts from cells induced with the PKR activator pIC. An in vitro dephosphorylation assay showed that when B56alpha was phosphorylated by PKR, the activity of PP2A trimeric holoenzyme was increased. A functional interaction between B56alpha and PKR was observed in cotransfection assays, where a B56alpha-mediated increase in luciferase expression was inhibited by cotransfection with wild-type PKR. This is likely due to a decreased level of eIF4E phosphorylation caused by an increase in PP2A activity following PKR phosphorylation of B56alpha. Taken together, our data indicate that PKR can modulate PP2A activity by phosphorylating B56alpha to regulate cellular activities.
Xu
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Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Williams
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BR
D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D039561:Eukaryotic Initiation Factor-4E; D006801:Humans; D007526:Isoelectric Point; D051379:Mice; D008810:Mice, Inbred C57BL; D010448:Peptide Initiation Factors / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D054648:Protein Phosphatase 2; D012694:Serine / Q000378:metabolism; D013379:Substrate Specificity; D013912:Threonine / Q000378:metabolism; D014162:Transfection; D020798:Two-Hybrid System Techniques; D019892:eIF-2 Kinase / Q000235:genetics / Q000378:metabolism*
D039561:Eukaryotic Initiation Factor-4E; D010448:Peptide Initiation Factors; D013912:Threonine; D012694:Serine; D019892:eIF-2 Kinase; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2