RAX, a Cellular Activator for Double-stranded RNA-dependent Protein Kinase during Stress Signaling

Abstract

The double-stranded (ds) RNA-dependent protein kinase (PKR) regulates protein synthesis by phosphorylating the ␣ subunit of eukaryotic initiation factor-2. PKR is activated by viral induced dsRNA and thought to be involved in the host antiviral defense mechanism. PKR is also activated by various nonviral stresses such as growth factor deprivation, although the mechanism is unknown. By screening a mouse cDNA expression library, we have identified an ubiquitously expressed PKR-associated protein, RAX. RAX has a high sequence homology to human PACT, which activates PKR in the absence of dsRNA. Although RAX also can directly activate PKR in vitro, overexpression of RAX does not induce PKR activation or inhibit growth of interleukin-3 (IL-3)-dependent cells in the presence of IL-3. However, IL-3 deprivation as well as diverse cell stress treatments including arsenite, thapsigargin, and H 2 O 2 , which are known to inhibit protein synthesis, induce the rapid phosphorylation of RAX followed by RAX-PKR association and activation of PKR. Therefore, cellular RAX may be a stress-activated, physiologic activator of PKR that couples transmembrane stress signals and protein synthesis.

Roles of the translationally controlled tumour protein (TCTP) and the double-stranded RNA-dependent protein kinase, PKR, in cellular stress responses

“…Indeed, PKR activation alone may not be enough to result in loss of TCTP, since arsenite is also a well-documented activator of PKR (Ito et al, 1999;Patel et al, 2000). Taken together, our data favour a model in which PKR activation, eIF2a phosphorylation and an additional signal related to Ca 2 þ stress are required for reduction in TCTP expression (Figure 7).…”

“…In contrast, in PKR-knockout cells TCTP levels are largely unaffected under any of these conditions, indicating that PKR is required for downregulation of TCTP levels under Ca þ þ -stress conditions. The fact that tunicamycin and sodium arsenite were unable to downregulate TCTP in wild-type cells, in spite of being activators of PKR and inducers of eIF2a phosphorylation (Ito et al, 1999;Patel et al, 2000), suggests that PKR-mediated phosphorylation of eIF2a is not sufficient to affect TCTP expression and that an additional Ca þ þ -stress-dependent step is involved.…”

PKR; a sentinel kinase for cellular stress

“…PKR activation by heparin A does not require the dsRBD and a mutant lacking the Nterminal 145 amino acids can be activated by heparin (Patel et al, 1994). Recently protein activators of PKR have been described in both human (PACT) and mouse cells (RAX) (Patel and Sen, 1998;Ito et al, 1999). PACT contains two dsRBMs and heterodimerizes with PKR via the dsRBD but activates it in the absence of dsRNA (Patel and Sen, 1998).…”

“…PACT contains two dsRBMs and heterodimerizes with PKR via the dsRBD but activates it in the absence of dsRNA (Patel and Sen, 1998). The activation of RAX and its association with PKR is stimulated by growth factor deprivation in IL3-dependent cells (Ito et al, 1999).…”

“…There are similar gaps in our knowledge of how the Jak tyrosine kinase pathway is linked to PKR. However, in a murine interleukin 3 (IL-3)-dependent cell line, IL-3 deprivation results in PKR activation that may involve a protein activator (Ito et al, 1999). Addition of IL-3 to deprived cells induces a reciprocal response characterized by the rapid dephosphorylation and inactivation of PKR.…”