Splicing factor SF3b as a target of the antitumor natural product pladienolide.
Pladienolide is a naturally occurring antitumor macrolide that was discovered by using a cell-based reporter gene expression assay controlled by the human vascular endothelial growth factor promoter. Despite the unique mechanisms of action and prominent antitumor activities of pladienolides B and D in diverse in vitro and in vivo systems, their target protein has remained unclear. We used 3H-labeled, fluorescence-tagged and photoaffinity/biotin (PB)-tagged 'chemical probes' to identify a 140-kDa protein in splicing factor SF3b as the binding target of pladienolide. Immunoblotting of an enhanced green fluorescent protein fusion protein of SF3b subunit 3 (SAP130) revealed direct interaction between the PB probe and SAP130. The binding affinities of pladienolide derivatives to the SF3b complex were highly correlated with their inhibitory activities against reporter gene expression and cell proliferation. Furthermore, pladienolide B impaired in vivo splicing in a dose-dependent manner. Our results demonstrate that the SF3b complex is a pharmacologically relevant protein target of pladienolide and suggest that this splicing factor is a potential antitumor drug target.
Kotake
Yoshihiko
Y
Tsukuba Research Laboratories, Eisai Co., Ltd., 5-1-3 Tokodai, Tsukuba, Ibaraki 300-2635, Japan.
Sagane
Koji
K
Owa
Takashi
T
Mimori-Kiyosue
Yuko
Y
Shimizu
Hajime
H
Uesugi
Mai
M
Ishihama
Yasushi
Y
Iwata
Masao
M
Mizui
Yoshiharu
Y
D000970:Antineoplastic Agents / Q000494:pharmacology*; D049109:Cell Proliferation / Q000187:drug effects; D016503:Drug Delivery Systems; D004852:Epoxy Compounds / Q000494:pharmacology*; D017930:Genes, Reporter; D006801:Humans; D018942:Macrolides / Q000494:pharmacology*; D011485:Protein Binding; D012326:RNA Splicing / Q000187:drug effects; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins / Q000037:antagonists & inhibitors*; D017413:Ribonucleoprotein, U2 Small Nuclear
D000970:Antineoplastic Agents; D004852:Epoxy Compounds; D018942:Macrolides; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; D017413:Ribonucleoprotein, U2 Small Nuclear; C486349:SF3B3 protein, human; C522342:pladienolide B; C522343:pladienolide D