RAX, the PKR activator, sensitizes cells to inflammatory cytokines, serum withdrawal, chemotherapy, and viral infection.
While the interferon (IFN)-inducible double-stranded RNA (dsRNA)-dependent protein kinase PKR is reported to initiate apoptosis in some instances, the mechanism by which diverse stress stimuli activate PKR remains unknown. Now we report that RAX, the only known cellular activator for PKR, initiates PKR activation in response to a broad range of stresses including serum deprivation, cytotoxic cytokine or chemotherapy treatment, or viral infection. Thus, knock-down of RAX expression by 80% using small interfering RNA (siRNA) prevents IFNgamma/tumor necrosis factor alpha (TNFalpha)-induced PKR activation and eIF2alpha phosphorylation, IkappaB degradation, IRF-1 expression, and STAT1 phosphorylation, resulting in enhanced murine embryonic fibroblast (MEF) cell survival. In contrast, expression of exogenous RAX, but not of the nonphosphorylatable, dominant-negative RAX(S18A) mutant, sensitizes cells to IFNgamma/TNFalpha, mitomycin C (MMC), or serum deprivation in association with increased PKR activity and apoptosis. Furthermore, RAX(S18A) expression in Fanconi anemia complementation group C-null MEF cells not only prevents PKR activation but also blocks hypersensitivity to IFNgamma/TNFalpha or mitomycin C that results in enhanced apoptosis. In addition, reduced RAX expression facilitates productive viral infection with vesicular stomatitis virus (VSV) and promotes anchorage-independent colony growth of MEF cells. Collectively, these data indicate that RAX may function as a negative regulator of growth that is required to activate PKR in response to a broad range of apoptosis-inducing stress.
Bennett
Richard L
RL
Department of Immunology and Molecular Genetics, Shands Cancer Center, University of Florida, Gainesville, USA.
Blalock
William L
WL
Abtahi
Dean M
DM
Pan
Yu
Y
Moyer
Sue A
SA
May
W Stratford
WS
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D002460:Cell Line; D049109:Cell Proliferation; D016207:Cytokines / Q000494:pharmacology; D005136:Eye Proteins / Q000235:genetics / Q000502:physiology*; D005347:Fibroblasts / Q000166:cytology; D018398:Homeodomain Proteins / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007249:Inflammation; D051379:Mice; D018962:Phlebotomy; D034741:RNA, Small Interfering / Q000494:pharmacology; D016601:RNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D013312:Stress, Physiological / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014777:Virus Diseases / Q000209:etiology; D019892:eIF-2 Kinase / Q000378:metabolism*
D000970:Antineoplastic Agents; D016207:Cytokines; D005136:Eye Proteins; D018398:Homeodomain Proteins; C486393:RAX protein, human; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D014157:Transcription Factors; D019892:eIF-2 Kinase