Adenosine deaminase ADAR1 increases gene expression at the translational level by decreasing protein kinase...

ADAR1 (adenosine deaminase acting on RNA) catalyzes the deamination of adenosine to inosine on RNA substrates with double-stranded character. Here, we show that coexpression of ADAR1 in mammalian cells markedly increases plasmid-based gene expression in transfected cells. The enhanced expression was independent of the nature of the promoter (viral and cellular) used to drive gene expression, of the protein reporter (luciferase and RRP) tested, and of the human cell line examined (293T and HeLa). Exogenous protein levels were increased by approximately 20-fold to approximately 50-fold when ADAR1 was coexpressed, whereas RNA transcript levels changed by less than 2-fold. The activation of PKR (protein kinase regulated by RNA) protein kinase and the phosphorylation of translation initiation factor eIF-2alpha seen following plasmid DNA transfection were both greatly reduced in ADAR1-transfected cells. Stable knockdown of the PKR kinase increased reporter gene expression in the absence, but not in the presence, of ADAR1 coexpression. Both size forms of ADAR1-the p150-inducible form and the p110-like constitutive form-enhanced plasmid-based gene expression. Taken together, these results indicate that the ADAR1 deaminase increases exogenous gene expression at the translational level by decreasing PKR-dependent eIF-2alpha phosphorylation.