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 PKIR 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, cytoto),Ic cytokine or chemotherapy treatment, or viral infection. Thus, knockdown of RAX expression by 80% using small interfering RNA (siRNA) prevents IFN-gamma tumor necrosis factor alpha (TNF alpha)- induced PKR activation and eIF2 alpha phosphorylation, I kappa B 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 IFN gamma/TNF alpha, 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 IFN-gamma/TNF alpha 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-i ndepen dent 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.