title: Proteasome-mediated degradation of cell division cycle 25C and cyclin-dependent kinase 1 in phenethyl isothiocyanate-induced G2-M-phase cell cycle arrest in PC-3 human prostate cancer cells.
abstract: Phenethyl isothiocyanate (PEITC), a constituent of many cruciferous vegetables, offers significant protection against cancer in animals induced by a variety of carcinogens. The present study demonstrates that PEITC suppresses proliferation of PC-3 cells in a dose-dependent manner by causing G(2)-M-phase cell cycle arrest and apoptosis. Interestingly, phenyl isothiocyanate (PITC), which is a structural analogue of PEITC but lacks the -CH(2) spacers that link the aromatic ring to the -N=C=S group, neither inhibited PC-3 cell viability nor caused cell cycle arrest or apoptosis. These results indicated that even a subtle change in isothiocyanate (ITC) structure could have a significant impact on its biological activity. The PEITC-induced cell cycle arrest was associated with a >80% reduction in the protein levels of cyclin-dependent kinase 1 (Cdk1) and cell division cycle 25C (Cdc25C; 24 h after treatment with 10 micro M PEITC), which led to an accumulation of Tyr(15) phosphorylated (inactive) Cdk1. On the other hand, PITC treatment neither reduced protein levels of Cdk1 or Cdc25C nor affected Cdk1 phosphorylation. The PEITC-induced decline in Cdk1 and Cdc25C protein levels and cell cycle arrest were significantly blocked on pretreatment of PC-3 cells with proteasome inhibitor lactacystin. A 24 h exposure of PC-3 cells to 10 micro M PEITC, but not PITC, resulted in about 56% and 44% decrease in the levels of antiapoptotic proteins Bcl-2 and Bcl-X(L), respectively. However, ectopic expression of Bcl-2 failed to alter sensitivity of PC-3 cells to growth inhibition or apoptosis induction by PEITC. Treatment of cells with PEITC, but not PITC, also resulted in cleavage of procaspase-3, procaspase-9, and procaspase-8. Moreover, the PEITC-induced apoptosis was significantly attenuated in the presence of general caspase inhibitor and specific inhibitors of caspase-8 and caspase-9. In conclusion, our data indicate that PEITC-induced cell cycle arrest in PC-3 cells is likely due to proteasome-mediated degradation of Cdc25C and Cdk1, and ectopic expression of Bcl-2 fails to confer resistance to PEITC-induced apoptosis. Furthermore, the results of the present study point toward involvement of both caspase-8- and caspase-9-mediated pathways in apoptosis induction by PEITC.
authors.affiliation: Department of Pharmacology and University of Pittsburgh Cancer Institute, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, USA.
authors.forename: Dong
authors.initials: D
authors.lastname: Xiao
authors.forename: Candace S
authors.initials: CS
authors.lastname: Johnson
authors.forename: Donald L
authors.initials: DL
authors.lastname: Trump
authors.forename: Shivendra V
authors.initials: SV
authors.lastname: Singh
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D016588:Anticarcinogenic Agents / Q000037:antagonists & inhibitors / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D020169:Caspases / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D016195:G2 Phase / Q000187:drug effects*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D017879:Isothiocyanates / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D008297:Male; D011471:Prostatic Neoplasms / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D051020:bcl-X Protein; D020687:cdc25 Phosphatases / Q000378:metabolism*
chemical_list: D016588:Anticarcinogenic Agents; C494811:BCL2L1 protein, human; D018797:Cell Cycle Proteins; D015853:Cysteine Proteinase Inhibitors; D017879:Isothiocyanates; D019253:Proto-Oncogene Proteins c-bcl-2; D051020:bcl-X Protein; C067713:lactacystin; C058305:phenethyl isothiocyanate; D016203:CDC2 Protein Kinase; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases; D020169:Caspases; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine