Caspase activation and changes in Bcl-2 family member protein expression associated with E2F-1-mediated apoptosis in human esophageal cancer cells.
The prognosis for patients with esophageal cancer remains poor, prompting the search for new treatment strategies. Overexpression of E2F-1 has been shown to induce apoptosis in several cancer cell types. In the present study, the effect of adenovirus-mediated E2F-1 overexpression on human esophageal cancer cell lines Yes-4 and Yes-6 was evaluated. Cells were treated by mock infection, infection with an adenoviral vector expressing beta-galactosidase (Ad5CMV-LacZ), or E2F-1 (Ad5CMVE2F-1). Western blot analysis confirmed marked overexpression of E2F-1 in Ad5CMVE2F-1-infected cells. Overexpression of E2F-1 resulted in marked growth inhibition and rapid loss of cell viability due to apoptosis, although Yes-6 cells were somewhat more resistant to E2F-1-mediated growth inhibition than Yes-4 cells. Cell cycle analysis revealed that overexpression of E2F-1 led to G2 arrest, followed by apoptotic cell death. p53 expression remained undetectable in both cell lines after E2F-1 overexpression. The apoptosis inhibitor proteins of the Bcl-2 gene family, Bcl-2, Mcl-1, and BcI-XL, decreased at 48 h after infection in Yes-4 cells, but remained unchanged in Yes-6 cells. Levels of retinoblastoma gene product (pRb) declined at 48 h after E2F-1 infection in Yes-4 cells, at which apoptosis predominated, whereas pRb expression remained constant in Yes-6 cells. Expression of p14ARF did not change after E2F-1 infection in either cell line. Involvement of caspase 3 and caspase 6 in E2F-1-mediated apoptosis was demonstrated by cleavage of caspase 3/CPP32 and poly-ADP-ribose polymerase, as well as fragmentation of the caspase 6 substrate, lamin B. These results indicate that the sensitivity of esophageal cancer cells to E2F-1-mediated apoptosis may be related to differential expression of Bcl-2 family member proteins and suggest that the adenovirus-mediated E2F-1 gene therapy may be a promising treatment strategy for the treatment of this disease.
Yang
H L
HL
Department of Surgery, University of Louisville, James Graham Brown Cancer Center, Kentucky 40202, USA.
Dong
Y B
YB
Elliott
M J
MJ
Liu
T J
TJ
McMasters
K M
KM
D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis; D002352:Carrier Proteins; D020169:Caspases / Q000378:metabolism*; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D016923:Cell Death; D002455:Cell Division; D002470:Cell Survival; D004274:DNA, Recombinant / Q000235:genetics; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D004789:Enzyme Activation; D004938:Esophageal Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D015972:Gene Expression Regulation, Neoplastic; D005822:Genetic Vectors; D006801:Humans; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins / Q000096:biosynthesis; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis*; D016160:Retinoblastoma Protein / Q000096:biosynthesis; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics; D051020:bcl-X Protein
C494811:BCL2L1 protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004274:DNA, Recombinant; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D016160:Retinoblastoma Protein; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D051020:bcl-X Protein; D020169:Caspases