| RRC ID |
39905
|
| 著者 |
Kanazawa T, Watanabe M, Matsushima-Hibiya Y, Kono T, Tanaka N, Koyama K, Sugimura T, Wakabayashi K.
|
| タイトル |
Distinct roles for the N- and C-terminal regions in the cytotoxicity of pierisin-1, a putative ADP-ribosylating toxin from cabbage butterfly, against mammalian cells.
|
| ジャーナル |
Proc Natl Acad Sci U S A
|
| Abstract |
Pierisin-1 is an 850-aa cytotoxic protein found in the cabbage butterfly, Pieris rapae, and has been suggested to consist of an N-terminal region with ADP-ribosyltransferase domain and of a C-terminal region that might have a receptor-binding domain. To elucidate the role of each region, we investigated the functions of various fragments of pierisin-1. In vitro expressed polypeptide consisting of amino acid residues 1-233 or 234-850 of pierisin-1 alone did not show cytotoxicity against human cervical carcinoma HeLa cells. However, the presence of both polypeptides in the culture medium showed some of the original cytotoxic activity. Introduction of the N-terminal polypeptide alone by electroporation also induced cell death in HeLa cells, and even in the mouse melanoma MEB4 cells insensitive to pierisin-1. Thus, the N-terminal region has a principal role in the cytotoxicity of pierisin-1 inside mammalian cells. Analyses of incorporated pierisin-1 indicated that the entire protein, regardless of whether it consisted of a single polypeptide or two separate N- and C-terminal polypeptides, was incorporated into HeLa cells. However, neither of the terminal polypeptides was incorporated when each polypeptide was present separately. These findings indicate that the C-terminal region is important for the incorporation of pierisin-1. Moreover, presence of receptor for pierisin-1 in the lipid fraction of cell membrane was suggested. The cytotoxic effects of pierisin-1 were enhanced by previous treatment with trypsin, producing "nicked" pierisin-1. Generation of the N-terminal fragment in HeLa cells was detected after application of intact entire molecule of pierisin-1. From the above observations, it is suggested that after incorporation of pierisin-1 into the cell by interaction of its C-terminal region with the receptor in the cell membrane, the entire protein is cleaved into the N- and C-terminal fragments with intracellular protease, and the N-terminal fragment then exhibits cytotoxicity.
|
| 巻・号 |
98(5)
|
| ページ |
2226-31
|
| 公開日 |
2001-2-27
|
| DOI |
10.1073/pnas.051628898
|
| PII |
98/5/2226
|
| PMID |
11226221
|
| PMC |
PMC30120
|
| MeSH |
ADP Ribose Transferases
ADP-Ribosylation Factors / chemistry
ADP-Ribosylation Factors / pharmacology*
Amino Acid Sequence
Animals
Apoptosis / drug effects
Base Sequence
Butterflies
Cell Survival / drug effects*
Cytotoxins / chemistry
Cytotoxins / pharmacology*
DNA Primers
Fluorescent Dyes
Humans
Insect Proteins / chemistry
Insect Proteins / pharmacology*
Tumor Cells, Cultured
|
| IF |
9.412
|
| 引用数 |
28
|
|
WOS 分野
|
BIOCHEMISTRY & MOLECULAR BIOLOGY
|
| リソース情報 |
| ヒト・動物細胞 |
HeLa(RCB0007)
MEB4(RCB1027) |
