Caspase-1 targets the TLR adaptor Mal at a crucial TIR-domain interaction site.
Toll-like receptors (TLRs) are crucial components of innate immunity, ensuring efficient responses against invading pathogens. After ligand binding, TLR signaling is initiated by recruitment of adaptor molecules, a step mediated by homotypic Toll-IL-1 receptor (TIR) domain interactions. Four TIR-containing TLR adaptor molecules are described, all of which are susceptible to modification and strict regulation. For example, caspase-1 is reported to cleave the TLR adaptor Mal at position D198, an event that is indispensible for Mal function. In this report, we use the mammalian two-hybrid technique MAPPIT to study the implications of Mal cleavage. We show that a Mal mutant, which mimics caspase-1 cleavage and a caspase-1-uncleavable MalD198A mutant, are abrogated in their bridging function and lose the ability to activate NF-kappaB. A MalD198E mutant is still fully functional, suggesting that caspase-1 cleavage of Mal is not necessary for Mal-mediated signaling. D198 of Mal is conserved in MyD88 and TLR4 TIR domains and the negatively charged amino acid at this position is crucial for the interactions and function of Mal, MyD88 and TLR4 TIR. Our data suggest an inhibitory, rather than an activating role for caspase-1 in Mal regulation, and show that the caspase-1 cleavage site in Mal is part of a TIR-domain interaction site.
Ulrichts
Peter
P
Department of Medical Protein Research, VIB, B-9000 Ghent, Belgium.
Bovijn
Celia
C
Lievens
Sam
S
Beyaert
Rudi
R
Tavernier
Jan
J
Peelman
Frank
F
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D024342:Amino Acids, Acidic / Q000378:metabolism; D001665:Binding Sites; D020170:Caspase 1 / Q000378:metabolism*; D002460:Cell Line; D017124:Conserved Sequence; D006801:Humans; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism*; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D053594:Myeloid Differentiation Factor 88 / Q000235:genetics / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D017472:Receptors, Interleukin-1 / Q000737:chemistry* / Q000378:metabolism*; D016415:Sequence Alignment; D051197:Toll-Like Receptor 4 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051193:Toll-Like Receptors / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D024342:Amino Acids, Acidic; D008562:Membrane Glycoproteins; D050505:Mutant Proteins; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D017472:Receptors, Interleukin-1; C434984:TIRAP protein, human; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D020170:Caspase 1