Expression of human papilloma virus E7 protein causes apoptosis and inhibits DNA synthesis in primary hepatocytes via increased expression of p21(Cip-1/WAF1/MDA6).
The impact of human papilloma virus (HPV16) E7 proteins and retinoblastoma (RB) antisense oligonucleotides upon mitogen-activated protein kinase (MAPK)-mediated inhibition of DNA synthesis via p21(Cip-1/WAF1/MDA6) (p21) was determined in primary hepatocytes. Prolonged activation of the MAPK pathway in p21(+/+) or p21(-/-) hepatocytes caused a large decrease and increase, respectively, in DNA synthesis. Either transfection with RB antisense oligonucleotides, expression of wild type E7, or RB binding mutant E7 (C24S) proteins increased p21 levels and reduced DNA synthesis in p21(+/+) hepatocytes. RB antisense oligonucleotides and E7 proteins increased apoptosis in p21(+/+), but not p21(-/-), hepatocytes. Expression of wild type E7 increased DNA synthesis above control levels in p21(-/-) cells, which was additive with prolonged MAPK activation. In contrast, expression of mutant E7 did not alter DNA synthesis above control levels in p21(-/-) cells and was supra-additive with prolonged MAPK activation. Antisense ablation of RB in p21(-/-) hepatocytes had a weak stimulatory effect upon DNA synthesis itself but enhanced the capacity of mutant E7 protein to stimulate DNA synthesis to the same level observed using wild type E7. The ability of prolonged MAPK activation to stimulate DNA synthesis in the presence of mutant E7 and antisense RB was additive. Collectively, the present data demonstrate that loss of RB function together with loss of p21 function plays an important role in the E7- and MAPK-dependent modulation of apoptosis and DNA synthesis in primary hepatocytes.
Park
J S
JS
Department of Radiation Oncology, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia 23298, USA.
Boyer
S
S
Mitchell
K
K
Gilfor
D
D
Birrer
M
M
Darlington
G
G
El Deiry
W
W
Firestone
G L
GL
Munger
K
K
Band
V
V
Fisher
P B
PB
Dent
P
P
D000818:Animals; D017209:Apoptosis; D042846:CDC2-CDC28 Kinases; D002455:Cell Division; D002472:Cell Transformation, Viral; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D016213:Cyclins / Q000096:biosynthesis* / Q000235:genetics; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D020935:MAP Kinase Signaling System; D051379:Mice; D008817:Mice, Mutant Strains; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D027383:Papillomaviridae; D050725:Papillomavirus E7 Proteins; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins; D019908:Proto-Oncogene Proteins c-raf / Q000235:genetics / Q000378:metabolism; D011960:Receptors, Estrogen / Q000235:genetics / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000096:biosynthesis / Q000235:genetics
C493957:Cdkn1a protein, mouse; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D016376:Oligonucleotides, Antisense; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D011518:Proto-Oncogene Proteins; D011960:Receptors, Estrogen; D016160:Retinoblastoma Protein; C059731:oncogene protein E7, Human papillomavirus type 16; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D042846:CDC2-CDC28 Kinases; C495893:Cdk2 protein, mouse; C495902:Cdk4 protein, mouse; D051357:Cyclin-Dependent Kinase 2; D051358:Cyclin-Dependent Kinase 4; D018844:Cyclin-Dependent Kinases