The double-stranded RNA-activated protein kinase mediates radiation resistance in mouse embryo fibroblasts through nuclear factor kappaB and Akt activation.
OBJECTIVE
Activation of the double-stranded RNA-activated protein kinase (PKR) leads to the induction of various pathways including the down-regulation of translation through phosphorylation of the eukaryotic translation initiation factor 2alpha (eIF-2alpha). There have been no reports to date about the role of PKR in radiation sensitivity.


METHODS
A clonogenic survival assay was used to investigate the sensitivity of PKR mouse embryo fibroblasts (MEF) to radiation therapy. 2-Aminopurine (2-AP), a chemical inhibitor of PKR, was used to inhibit PKR activation. Nuclear factor-kappaB (NF-kappaB) activation was assessed by electrophoretic mobility shift assay (EMSA). Expression of PKR and downstream targets was examined by Western blot analysis and immunofluorescence.


RESULTS
Ionizing radiation leads to dose- and time-dependent increases in PKR expression and function that contributes to increased cellular radiation resistance as shown by clonogenic survival and terminal nucleotidyl transferase-mediated nick end labeling (TUNEL) apoptosis assays. Specific inhibition of PKR with the chemical inhibitor 2-AP restores radiation sensitivity. Plasmid transfection of the PKR wild-type (wt) gene into PKR(-/-) MEFs leads to increased radiation resistance. The protective effect of PKR to radiation may be mediated in part through NF-kappaB and Akt because both NF-kappaB and Akt are activated after ionizing radiation in PKR+/+ but not PKR-/- cells.


CONCLUSIONS
We suggest a novel role for PKR as a mediator of radiation resistance modulated in part through the protective effects of NF-kappaB and Akt activation. The modification of PKR activity may be a novel strategy in the future to overcome radiation resistance.
von Holzen
Urs
U
Department of Surgical Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.
Pataer
Abujiang
A
Raju
Uma
U
Bocangel
Dora
D
Vorburger
Stephan A
SA
Liu
Yanna
Y
Lu
Xiaolin
X
Roth
Jack A
JA
Aggarwal
Bharat B
BB
Barber
Glen N
GN
Keyomarsi
Khandan
K
Hunt
Kelly K
KK
Swisher
Stephen G
SG
D000818:Animals; D003720:Densitometry; D004307:Dose-Response Relationship, Radiation; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D020287:In Situ Nick-End Labeling; D051379:Mice; D008954:Models, Biological; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011839:Radiation, Ionizing; D014162:Transfection; D019892:eIF-2 Kinase / Q000378:metabolism*
D015852:Eukaryotic Initiation Factor-2; D016328:NF-kappa B; D051057:Proto-Oncogene Proteins c-akt; D019892:eIF-2 Kinase