Sequestosome-1/p62 is the key intracellular target of innate defense regulator peptide.
Innate defense regulator-1 (IDR-1) is a synthetic peptide with no antimicrobial activity that enhances microbial infection control while suppressing inflammation. Previously, the effects of IDR-1 were postulated to impact several regulatory pathways including mitogen-activated protein kinase (MAPK) p38 and CCAAT-enhancer-binding protein, but how this was mediated was unknown. Using a combined stable isotope labeling by amino acids in cell culture-proteomics methodology, we identified the cytoplasmic scaffold protein p62 as the molecular target of IDR-1. Direct IDR-1 binding to p62 was confirmed by several biochemical binding experiments, and the p62 ZZ-type zinc finger domain was identified as the IDR-1 binding site. Co-immunoprecipitation analysis of p62 molecular complexes demonstrated that IDR-1 enhanced the tumor necrosis factor alpha-induced p62 receptor-interacting protein 1 (RIP1) complex formation but did not affect tumor necrosis factor alpha-induced p62-protein kinase zeta complex formation. In addition, IDR-1 induced p38 MAPK activity in a p62-dependent manner and increased CCAAT-enhancer-binding protein beta activity, whereas NF-kappaB activity was unaffected. Collectively, these results demonstrate that IDR-1 binding to p62 specifically affects protein-protein interactions and subsequent downstream events. Our results implicate p62 in the molecular mechanisms governing innate immunity and identify p62 as a potential therapeutic target in both infectious and inflammatory diseases.
Yu
Hong Bing
HB
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia V6T 1Z4.
Kielczewska
Agnieszka
A
Inimex Pharmaceuticals, Vancouver, British Columbia V5A 4T8.
Rozek
Annett
A
Inimex Pharmaceuticals, Vancouver, British Columbia V5A 4T8.
Takenaka
Shunsuke
S
Inimex Pharmaceuticals, Vancouver, British Columbia V5A 4T8.
Li
Yuling
Y
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia V6T 1Z4.
Thorson
Lisa
L
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia V6T 1Z4.
Hancock
Robert E W
REW
Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia V6T 1Z4.
Guarna
M Marta
MM
Department of Biochemistry and Molecular Biology, Centre for High-Throughput Biology, Vancouver, British Columbia V6T 1Z4, Canada.
North
John R
JR
Inimex Pharmaceuticals, Vancouver, British Columbia V5A 4T8.
Foster
Leonard J
LJ
Department of Biochemistry and Molecular Biology, Centre for High-Throughput Biology, Vancouver, British Columbia V6T 1Z4, Canada.
Donini
Oreola
O
Inimex Pharmaceuticals, Vancouver, British Columbia V5A 4T8.
Finlay
B Brett
BB
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia V6T 1Z4; Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia V6T 1Z4. Electronic address: bfinlay@interchange.ubc.ca.
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000818:Animals; D001665:Binding Sites / Q000235:genetics / Q000276:immunology; D022762:CCAAT-Enhancer-Binding Proteins / Q000276:immunology / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007113:Immunity, Innate / Q000187:drug effects* / Q000235:genetics; D007155:Immunologic Factors / Q000494:pharmacology*; D007249:Inflammation / Q000188:drug therapy / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D017433:Protein Structure, Secondary / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000235:genetics; D000071456:Sequestosome-1 Protein; D048051:p38 Mitogen-Activated Protein Kinases / Q000276:immunology / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D022762:CCAAT-Enhancer-Binding Proteins; D006360:Heat-Shock Proteins; D007155:Immunologic Factors; D016328:NF-kappa B; D010455:Peptides; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse; D048051:p38 Mitogen-Activated Protein Kinases