Human papillomavirus-16 E7 interacts with Siva-1 and modulates apoptosis in HaCaT human immortalized keratinocytes.
The viral factor E7 plays a key role in the well-established association between "high-risk" Human Papillomavirus (HPV) infection and the development of epithelial malignant tumors, as uterine cervix and ano-genital cancer. To delve into the molecular mechanisms of HPV-mediated cell transformation, we searched for novel potential cellular targets of the HPV-16 E7 oncoprotein, by means of the yeast two-hybrid technique, identifying a protein-protein interaction between HPV-16 E7 and the pro-apoptotic cellular factor Siva-1. Using co-precipitation assays and the "PepSets" technique, we confirmed this physical interaction and mapped accurately, for both proteins, the amino acid residues involved. Additionally, we found that HPV-16 E7 competed in vitro with the binding of the Bcl-X(L) anti-apoptotic factor to Siva-1, an interaction that has a major inference in UV radiation-induced apoptosis. In HaCaT immortalized human keratinocytes, forced HPV-16 E7 expression by retroviral infection caused Siva-1 transcript up-regulation, detected by cDNA macroarray hybridization and real-time quantitative PCR, paralleled by an increased amount of protein. Confirming the anti-apoptotic role of HPV-16 E7 in the HaCaT cellular model, evaluated by nuclear morphology, we also found that Siva-1 expression produced a significant increase of the apoptotic rate in UV radiation-exposed HaCaT cells, and that this effect resulted explicitly counteracted by HPV-16 E7. Being apoptosis a key physiological process for the elimination of irreversibly injured cells, the anti-apoptotic role of HPV-16 E7, performed at least by its interference with Siva-1, can be considered an additional mechanism for the survival of damaged, potentially transforming, cell clones.
Severino
Anna
A
Department for the Development of Therapeutic Programs, Regina Elena Cancer Institute, Rome, Italy.
Abbruzzese
Claudia
C
Manente
Lucrezia
L
Valderas
Alvaro Avivar
AA
Mattarocci
Stefano
S
Federico
Antonio
A
Starace
Giuseppe
G
Chersi
Alberto
A
Mileo
Anna Maria
AM
Paggi
Marco G
MG
D017209:Apoptosis / Q000502:physiology; D051017:Apoptosis Regulatory Proteins; D002460:Cell Line; D015723:Gene Library; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D015603:Keratinocytes / Q000378:metabolism*; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D050725:Papillomavirus E7 Proteins; D011485:Protein Binding; D051020:bcl-X Protein / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; C494811:BCL2L1 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; C106549:SIVA1 protein, human; D051020:bcl-X Protein; C056651:oncogene protein E4, human papilloma virus type 1; C059731:oncogene protein E7, Human papillomavirus type 16