MyD88 adapter-like (Mal)/TIRAP interaction with TRAF6 is critical for TLR2- and TLR4-mediated NF-kappaB proinflammatory responses.
Toll/interleukin-1 (TIR)receptor-containing adapters are critical in orchestrating the different signal transduction pathways following Toll-like receptor (TLR) activation. MyD88 adapter-like (Mal), also termed TIRAP, is involved in bridging MyD88 to the receptor complex for TLR-2 and TLR4 signaling in response to bacterial infection. We have previously reported an interaction between Mal and tumor necrosis factor receptor-associated factor 6 (TRAF6) via a TRAF6-binding motif, the disruption of which inhibited TLR-mediated NF-kappaB-luciferase reporter activity. Given the recent report of intracellular TRAM localization promoting sequential signaling in TLR4 responses, we further characterized Mal interaction with TRAF6, the cellular localization, and the outcomes of disrupting this association on TLR inflammatory responses. We found that Mal and TRAF6 directly interact in response to TLR2 and TLR4 stimulation, although membrane localization is not necessary to facilitate interaction. Critically, reconstitution of murine Mal-deficient macrophages with MalE190A, containing a mutation within the TRAF6-binding motif, fails to reconstitute the proinflammatory response to TLR2 and TLR4 ligands compared with wild type Mal. Furthermore, Mal interaction with TRAF6 mediates Ser phosphorylation of the p65 subunit of NF-kappaB and thus controls transcriptional activation but not nuclear translocation of NF-kappaB. This study characterizes the novel role for Mal in facilitating the direct recruitment of TRAF6 to the plasma membrane, which is necessary for TLR2- and TLR4-induced transactivation of NF-kappaB and regulation of the subsequent pro-inflammatory response.
Verstak
Brett
B
Centre for Innate Immunity and Infectious Disease, Monash Institute of Medical Research, Monash University, Clayton, Victoria 3168, Australia.
Nagpal
Kamalpreet
K
Bottomley
Stephen P
SP
Golenbock
Douglas T
DT
Hertzog
Paul J
PJ
Mansell
Ashley
A
D020816:Amino Acid Motifs / Q000235:genetics / Q000276:immunology; D000818:Animals; D001424:Bacterial Infections / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D002461:Cell Line, Transformed; D006801:Humans; D007249:Inflammation / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D009154:Mutation; D053594:Myeloid Differentiation Factor 88 / Q000235:genetics / Q000276:immunology; D016328:NF-kappa B / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D010766:Phosphorylation / Q000235:genetics / Q000276:immunology; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D017472:Receptors, Interleukin-1 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics / Q000276:immunology; D048029:TNF Receptor-Associated Factor 6 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051195:Toll-Like Receptor 2 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015533:Transcriptional Activation / Q000235:genetics / Q000276:immunology
C507779:MYD88 protein, human; D008562:Membrane Glycoproteins; C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D017472:Receptors, Interleukin-1; C434984:TIRAP protein, human; C499045:TIRAP protein, mouse; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D048029:TNF Receptor-Associated Factor 6; C493486:Tlr2 protein, mouse; C493487:Tlr4 protein, mouse; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4