title: Control of a tumor suppressor PDCD4: Degradation mechanisms of the protein in hepatocellular carcinoma cells.
abstract: In this study, we demonstrate that EGF inhibits the TGF-β1-induced apoptosis of Huh7 cells. TGF-β1 up-regulates the expression of PDCD4 causing apoptosis, by stimulating the synthesis of PDCD4 mRNA via the Smad signaling pathway. TGF-β1 also inhibits the activation of S6 kinase 1 which phosphorylates the serine 67 residue of PDCD4 and leads to the phosphorylation of serine 71 and serine 76 in the β-TRCP binding sequence. This phosphorylation sequence causes the protein to be degraded in the ubiquitin-proteasome system. EGF activates S6 kinase 1 via the PI3K-Akt-mTOR signaling pathway and stimulates the degradation of PDCD4. EGF also suppresses PDCD4 mRNA levels. As the mTOR inhibitor rapamycin up-regulated PDCD4 mRNA levels, the PI3K-Akt-mTOR signaling pathway may control the transcription of the PDCD4 gene as well as the degradation of the protein. TPA also inhibited the TGF-β1-induced apoptosis of Huh7 cells, stimulating the degradation of the PDCD4-protein. Analyses using PDCD4 mutants with changes of serines 67, 71 and 76 to alanine revealed that the phosphorylation of serine 67 is not essential for the TPA-induced suppression of the protein. The mitogens could not suppress the PDCD4-mutant proteins with changes of serine 71 and/or serine 76 to alanine, however, indicating that phosphorylations at these residues are necessary for the proteasome-mediated degradation of PDCD4. The phosphor-mimic S71/D and S76/D mutants were able to be degraded in the ubiquitin-proteasome system unlike the mutants with changes of serine to alanine. The expression of S71/D mutant was suppressed with EGF but that of S76/D mutant was not indicating that at least partly the phosphorylation of both sites was mediated by different enzymes.
authors: Department of Internal Medicine, Saga Medical School, Saga University, Japan. Electronic address: matsuha2@edu.cc.saga-u.ac.jp.
authors: Sachiko
authors: S
authors: Matsuhashi
authors: Organization for Cooperation with Industry and Regional Community, Saga University, Japan.
authors: Hiroshi
authors: H
authors: Hamajima
authors: Department of Internal Medicine, Saga Medical School, Saga University, Japan.
authors: Jinghe
authors: J
authors: Xia
authors: Department of Surgery, First Affiliated Hospital, China Medical University, Shenyang, People's Republic of China.
authors: Hao
authors: H
authors: Zhang
authors: Department of Internal Medicine, Saga Medical School, Saga University, Japan.
authors: Toshihiko
authors: T
authors: Mizuta
authors: Department of Internal Medicine, Saga Medical School, Saga University, Japan.
authors: Keizo
authors: K
authors: Anzai
authors: Department of Internal Medicine, Saga Medical School, Saga University, Japan; Health Administration Centre, Saga Medical School, Saga University, Saga 849-8501, Japan.
authors: Iwata
authors: I
authors: Ozaki
mesh_terms: D017209:Apoptosis / Q000235:genetics*; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D045744:Cell Line, Tumor; D004815:Epidermal Growth Factor / Q000494:pharmacology; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism*; D009154:Mutation; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors; D012333:RNA, Messenger / Q000096:biosynthesis; D016601:RNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D020123:Sirolimus / Q000494:pharmacology; D051785:Smad Proteins / Q000235:genetics; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology
chemical_list: D051017:Apoptosis Regulatory Proteins; C114487:PDCD4 protein, human; D000081082:Phosphoinositide-3 Kinase Inhibitors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D051785:Smad Proteins; D053773:Transforming Growth Factor beta1; D004815:Epidermal Growth Factor; C546842:MTOR protein, human; D051057:Proto-Oncogene Proteins c-akt; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; C448901:ribosomal protein S6 kinase, 70kD, polypeptide 1; D020123:Sirolimus
keywords: Apoptosis; EGF; Huh7 hepatoma cells; PDCD4; TGF-β1; TPA