Involvement of caspase-1 and caspase-3 in the production and processing of mature human interleukin 18 in monocytic THP.1 cells.
Recently, human interleukin 18 (hIL-18) cDNA was cloned, and the recombinant protein with a tentatively assigned NH2-terminal amino acid sequence was generated. However, natural hIL-18 has not yet been isolated, and its cellular processing is therefore still unclear. To clarify this, we purified natural hIL-18 from the cytosolic extract of monocytic THP.1 cells. Natural hIL-18 exhibited a molecular mass of 18.2 kDa, and the NH2-terminal amino acid was Tyr37. Biological activities of the purified protein were identical to those of recombinant hIL-18 with respect to the enhancement of natural killer cell cytotoxicity and interferon-gamma production by human peripheral blood mononuclear cells. We also found two precursor hIL-18 (prohIL-18)-processing activities in the cytosol of THP.1 cells. These activities were blocked separately by the caspase inhibitors Ac-YVAD-CHO and Ac-DEVD-CHO. Further analyses of the partially purified enzymes revealed that one is caspase-1, which cleaves prohIL-18 at the Asp36-Tyr37 site to generate the mature hIL-18, and the other is caspase-3, which cleaves both precursor and mature hIL-18 at Asp71-Ser72 and Asp76-Asn77 to generate biologically inactive products. These results suggest that the production and processing of natural hIL-18 are regulated by two processing enzymes, caspase-1 and caspase-3, in THP.1 cells.
Akita
K
K
Fujisaki Institute, Hayashibara Biochemical Laboratories, Inc., 675-1 Fujisaki, Okayama 702, Japan.
Ohtsuki
T
T
Nukada
Y
Y
Tanimoto
T
T
Namba
M
M
Okura
T
T
Takakura-Yamamoto
R
R
Torigoe
K
K
Gu
Y
Y
Su
M S
MS
Fujii
M
M
Satoh-Itoh
M
M
Yamamoto
K
K
Kohno
K
K
Ikeda
M
M
Kurimoto
M
M
D000595:Amino Acid Sequence; D000818:Animals; D020170:Caspase 1; D053148:Caspase 3; D020169:Caspases; D002460:Cell Line; D002853:Chromatography, Liquid; D003546:Cysteine Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D016207:Cytokines / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D020382:Interleukin-18; D051379:Mice; D008969:Molecular Sequence Data; D009000:Monocytes / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000235:genetics; D017386:Sequence Homology, Amino Acid
D016207:Cytokines; D020382:Interleukin-18; D012333:RNA, Messenger; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D020170:Caspase 1