Identification and analysis of dominant negative mutants of RAIDD and PIDD.
Caspases are cysteine proteases that are essential during the initiation and execution of apoptosis and inflammation. The formation of large oligomeric protein complexes is critical to the activation of caspases in apoptotic and inflammatory signaling pathways. These oligomeric protein complexes function as a platform to recruit caspases, which leads to caspase activation via a proximity-induced mechanism. One well-known oligomeric caspase-activating complex is the PIDDosome for caspase-2 activation, which is composed of 3 protein components, PIDD, RAIDD and Caspase-2. Despite the significant role that caspase-2 activated by PIDDosome plays during genotoxic stress-induced apoptosis, the oligomerization mechanism and the method by which the caspase-activating process is mediated by the formation of PIDDosome is currently not well understood. Here, we show that the assembly mechanism of the core of PIDDosome is time-dependent and salt concentration-dependent. In addition, we demonstrate that point mutations on RAIDD (R147E) and on PIDD (Y814A) exert a dominant negative effect on the formation of the PIDDosome, and that this effect cannot be applied after the PIDDosome has been formed.
Jang
Tae-Ho
TH
Department of Biochemistry, School of Biotechnology, Yeungnam University, Gyeongsan, South Korea.
Bae
Ju Young
JY
Park
Ok Kyoung
OK
Kim
Ji Hoe
JH
Cho
Kyung-Hyun
KH
Jeon
Ju-Hong
JH
Park
Hyun Ho
HH
D000595:Amino Acid Sequence; D017209:Apoptosis; D053420:CRADD Signaling Adaptor Protein / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics*; D020169:Caspases / Q000378:metabolism; D057056:Cysteine Proteases / Q000737:chemistry; D053418:Death Domain Receptor Signaling Adaptor Proteins; D004789:Enzyme Activation; D005799:Genes, Dominant; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009153:Mutagens / Q000737:chemistry; D017354:Point Mutation; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction
D053420:CRADD Signaling Adaptor Protein; C104252:CRADD protein, human; D002352:Carrier Proteins; D053418:Death Domain Receptor Signaling Adaptor Proteins; D009153:Mutagens; C485896:PIDD1 protein, human; D057056:Cysteine Proteases; D020169:Caspases