Constitutive expression of human double-stranded RNA-activated p68 kinase in murine cells mediates phosphorylation of eukaryotic initiation factor 2 and partial resistance to encephalomyocarditis virus growth.
The cDNA encoding interferon-induced human double-stranded RNA-activated p68 kinase was expressed in murine NIH 3T3 cells by using the pcDNA1/neo vector. Several stable clones were selected which expressed either the wild-type kinase or an inactive mutant possessing a single amino acid substitution in the invariant lysine 296 in the catalytic domain II. The transfected wild-type kinase showed properties similar to those of the natural kinase, such as subcellular ribosomal localization and dependence on double-stranded RNA for autophosphorylation. Upon infection with encephalomyocarditis virus (EMCV), wild-type- but not mutant-expressing clones were found to partially resist virus growth. Such natural antiviral activity was virus specific, since no inhibition was observed in the case of vesicular stomatitis virus infection. In accord with EMCV inhibition, the wild-type p68 kinase was found to be highly phosphorylated during infection. Furthermore, its natural substrate, the small subunit of protein synthesis initiation factor eIF2, was phosphorylated. These results demonstrate that p68 kinase is activated during EMCV infection, leading to reduced virus production.
Meurs
E F
EF
Unit of Virology and of Cellular Immunology (UA CNRS 1157), Institut Pasteur, Paris, France.
Watanabe
Y
Y
Kadereit
S
S
Barber
G N
GN
Katze
M G
MG
Chong
K
K
Williams
B R
BR
Hovanessian
A G
AG
D016475:3T3 Cells; D000818:Animals; D000998:Antiviral Agents; D002478:Cells, Cultured; D015180:Electrophoresis, Gel, Two-Dimensional; D004686:Encephalomyocarditis virus / Q000254:growth & development*; D004790:Enzyme Induction; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D007372:Interferons / Q000494:pharmacology; D051379:Mice; D010766:Phosphorylation; D010957:Plasmids; D011494:Protein Kinases / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014721:Vesicular stomatitis Indiana virus / Q000254:growth & development; D019892:eIF-2 Kinase
D000998:Antiviral Agents; D015852:Eukaryotic Initiation Factor-2; D007372:Interferons; D011494:Protein Kinases; D019892:eIF-2 Kinase