TRAF family proteins link PKR with NF-kappa B activation.
The double-stranded RNA (dsRNA)-dependent protein kinase PKR activates NF-kappa B via the I kappa B kinase (IKK) complex, but little is known about additional molecules that may be involved in this pathway. Analysis of the PKR sequence enabled us to identify two putative TRAF-interacting motifs. The viability of such an interaction was further suggested by computer modeling. Here, we present evidence of the colocalization and physical interaction between PKR and TRAF family proteins in vivo, as shown by immunoprecipitation and confocal microscopy experiments. This interaction is induced upon PKR dimerization. Most importantly, we show that the binding between PKR and TRAFs is functionally relevant, as observed by the absence of NF-kappa B activity upon PKR expression in cells genetically deficient in TRAF2 and TRAF5 or after expression of TRAF dominant negative molecules. On the basis of sequence information and mutational and computer docking analyses, we favored a TRAF-PKR interaction model in which the C-terminal domain of TRAF binds to a predicted TRAF interaction motif present in the PKR kinase domain. Altogether, our data suggest that TRAF family proteins are key components located downstream of PKR that have an important role in mediating activation of NF-kappa B by the dsRNA-dependent protein kinase.
Gil
Jesús
J
Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas, Campus Universidad Autónoma, 28049 Madrid, Spain.
García
Maria Angel
MA
Gomez-Puertas
Paulino
P
Guerra
Susana
S
Rullas
Joaquín
J
Nakano
Hiroyasu
H
Alcamí
José
J
Esteban
Mariano
M
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008958:Models, Molecular; D016328:NF-kappa B / Q000378:metabolism*; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D047992:TNF Receptor-Associated Factor 2; D048015:TNF Receptor-Associated Factor 5; D019892:eIF-2 Kinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D016328:NF-kappa B; D011506:Proteins; D012330:RNA, Double-Stranded; D047992:TNF Receptor-Associated Factor 2; D048015:TNF Receptor-Associated Factor 5; D004247:DNA; D019892:eIF-2 Kinase