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Inhibition of proteasome‐dependent degradation of Wee1 in G 2 ‐arrested Hep3B cells by TGFβ1
Author(s) -
Hashimoto Osamu,
Ueno Takato,
Kimura Rina,
Ohtsubo Motoaki,
Nakamura Toru,
Koga Hironori,
Torimura Takuji,
Uchida Sanae,
Yamashita Katsumi,
Sata Michio
Publication year - 2003
Publication title -
molecular carcinogenesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.254
H-Index - 97
eISSN - 1098-2744
pISSN - 0899-1987
DOI - 10.1002/mc.10111
Subject(s) - wee1 , cyclin dependent kinase 1 , biology , cyclin dependent kinase , cyclin b1 , cell cycle , cell cycle checkpoint , kinase , hyperphosphorylation , phosphorylation , microbiology and biotechnology , cdk inhibitor , proteasome , apoptosis , biochemistry
Transforming growth factor β1 (TGFβ1)‐induced G 2 arrest was observed when a proliferation inhibitory function of the retinoblastoma protein (Rb) was compromised, but the mechanism underlying the G 2 arrest was poorly characterized compared with that of G 1 arrest. In the present study, we characterized G 2 arrest induced by TGFβ1 (1 ng/mL) in the Rb‐negative hepatoma cell line (Hep3B) and compared with G 1 arrest in the Rb‐positive hepatoma cell line (Huh7). Activities of cyclin‐dependent kinases (CDK) 2 and cell division cycle (CDC) 2 were markedly decreased at 24 h, the time when cell‐cycle arrest became apparent in both cell lines. However, considerable amounts of inactive CDC2‐cyclinB1 complexes were present in the nucleus of G 2 ‐arrested Hep3B but were not present in G 1 ‐arrested Huh7. The inhibitory phosphorylation of CDC2 on Tyr‐15 was significantly elevated at 12–24 h, and its levels gradually declined during G 2 arrest in Hep3B. In particular, augmentation of CDK inhibitors p21 cip1 and p27 kip1 and Wee1 kinase and diminution of CDC25C phosphatase coincided with induced Tyr‐15 phosphorylation and inhibition of CDC2. Wee1 in Hep3B was unstable and was degraded in a proteasome‐dependent manner, but it became substantially stabilized within 6 h of TGFβ1 treatment. Moreover, a Wee1 inhibitor, PD0166285, abrogated the TGFβ1‐induced G 2 arrest in Hep3B. These findings suggest that TGFβ1 induced G 2 arrest in Hep3B at least in part through stabilization of Wee1 and subsequent increase in Tyr‐15 phosphorylation and inhibition of CDC2. © 2003 Wiley‐Liss, Inc.