Editors' ChoiceCell Cycle

PKR, Not Just for Infected Cells

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Science Signaling  01 Jul 2014:
Vol. 7, Issue 332, pp. ec180
DOI: 10.1126/scisignal.2005642

Virally infected cells use the protein double-stranded RNA (dsRNA)–dependent protein kinase R (PKR) to sense and respond to viral infection. Upon binding dsRNA, PKR becomes autophosphorylated and phosphorylates various proteins, including the α subunit of eukaryotic initiation factor 2 (eIF2α), thereby inhibiting protein translation. By immunofluorescence analysis of both asynchronous and synchronized cells in culture and cellular fractionation, Kim et al. found that phosphorylated PKR (pPKR) was abundant and associated with chromosomes in mitotic cells, but not in cells in interphase. Furthermore, phosphorylation of the PKR targets eIF2α and c-Jun N-terminal kinase (JNK) was also increased in the nuclei of mitotic cells, and their phosphorylation was reduced in mitotic cells by pharmacological inhibition of PKR, which also prevented the decrease in translation during M phase that occurred in control cells. Knockdown of PKR or expression of a dominant-negative mutant increased the proportion of cells in G2 and M phases, increased the abundance of G2-specific cell cycle regulators that are normally degraded in M phase, and reduced the amount of phosphorylated histone 3, which occurs in M phase, suggesting that PKR activity is necessary for G2 to M progression. Inhibition of mRNA synthesis with actinomycin D or digestion of dsRNA, but not single-stranded RNA, reduced the accumulation of pPKR and phosphorylated JNK on chromosomes of mitotic cells. Alu elements are short interspersed elements in the human genome and, when present in a specific back-to-back orientation and transcribed, these can form dsRNAs called IRAlus. Analysis of an IRAlu reporter by fluorescent in situ hybridization (FISH) revealed that this transcript was primarily in the nucleus in interphase cells and became present throughout the cell during mitosis. PKR and the mRNA of the reporter coimmunoprecipitated in cells that had been exposed to formaldehyde to cross-link the protein-RNA complexes before immunoprecipitation. Endogenous mRNAs with IRAlus also coimmunoprecipitated with PKR from cells undergoing mitosis. Thus, PKR is activated by endogenous dsRNA formed by Alu sequences that escape the nucleus during mitosis and contribute to the progression through G2 into M phase of the cell cycle.

Y. Kim, J. H. Lee, J.-E. Park, J. Cho, H. Yi, V. N. Kim, PKR is activated by cellular dsRNAs during mitosis and acts as a mitotic regulator. Genes Dev. 28, 1310–1322 (2014). [Abstract] [Full Text]

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